Pancreatic β-Cell Electrical Activity and Insulin Secretion: Of Mice and Men

Only a small fraction of secretory granules (<1%/h) undergo exocytosis even at high glucose concentrations.

“Only a small fraction of the secretory granules (<1%/h) undergo exocytosis even at high glucose concentrations (571).”

• cites [571] Rorsman P Renstrom E Insulin granule dynamics in pancreatic beta cells Diabetologia 2003 46 1029 1045 12879249 sourceDisputed

  The Ultrastructural correlates of biphasic insulin secretion section (page unavailable) describes sustained glucose-stimulated secretion at 5 granules per minute per beta cell. The same section reports about 10,000 granules per mouse beta cell, making that rate approximately 3% per hour, not less than 1% per hour. (LLM, conf=0.86)

  Checked by a reasoning model — may be wrong.

  Experimental — accuracy not yet measured

In most mouse β-cells, TTX-sensitive Na+ channels are not involved in action potential firing.

“For example, activation of TTX-sensitive Na+ channels plays a prominent role in human β-cell action potential firing at low glucose but not at high glucose (90, 521) - and in most mouse β-cells it is not involved in action potential firing at all (761).”

• cites [761] Zhang Q Chibalina MV Bengtsson M Groschner LN Ramracheya R Rorsman NJ Leiss V Nassar MA Welling A Gribble FM Reimann F Na+ current properties in islet alpha- and beta-cells reflect cell-specific Scn3a and Scn9a expression J Physiol 2014 592 4677 4696 25172946 PMC4253470 sourceDisputed

  The Results section ('Role of Na+ channels in islet cell electrical activity: effects of TTX'; page unavailable) reports that TTX markedly reduced action-potential amplitude in three of six mouse β-cells at 8 mM glucose and significantly reduced peak voltage at 20 mM glucose. Although it barely affected the other three cells tested at 8 mM, the Discussion ('Role of Na+ channels in β-cells'; page unavailable) concludes that a TTX-sensitive Na+ current likely contributes to action-potential firing. The source therefore does not support saying these channels are uninvolved in most mouse β-cells at all. (LLM, conf=0.89)

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  Experimental — accuracy not yet measured

During membrane depolarization, β-cell Ca2+ currents undergo both Ca2+- and voltage-dependent inactivation.

“During membrane depolarization, β-cell Ca2+ currents undergo both Ca2+- and voltage-dependent inactivation (515).”

• cites [515] Plant TD Properties and calcium-dependent inactivation of calcium currents in cultured mouse pancreatic B-cells J Physiol 1988 404 731 747 2855352 PMC1190852 sourceDisputed

  The paper’s summary (points 4 and 7; page not supplied) says inactivation depended on the amount of Ca2+ entering, independently of pre-pulse potential. It describes the mechanism as almost purely Ca2+-dependent, contrary to the claim that voltage-dependent inactivation also occurs. (LLM, conf=0.88)

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  Experimental — accuracy not yet measured

Action potential frequency was paradoxically reduced in β-cells from Knck3-deficient mice.

“Action potential frequency was paradoxically reduced (135), but this may be a consequence of voltage-dependent inactivation of other currents involved in action potential generation.”

• cites [135] Dadi PK Luo B Vierra NC Jacobson DA TASK-1 Potassium Channels Limit Pancreatic alpha-Cell Calcium Influx and Glucagon Secretion Mol Endocrinol 2015 29 777 787 25849724 PMC4415209 sourceDisputed

  In “TASK-1 currents regulate pancreatic α-cell Δp” (page not available), the paper reports greater action-potential firing in TASK-1-deficient α-cells than in controls under elevated glucose, not reduced firing. The experiment concerns α-cells, whereas the claim specifies β-cells. (LLM, conf=0.85)

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  Experimental — accuracy not yet measured

The reduction of Kslow in dispersed β-cells may account for the loss of normal oscillatory electrical activity seen in this preparation.

“which may account for the loss of normal oscillatory electrical activity seen in this preparation (but see (217)).”

• cites [223] Gopel SO Kanno T Barg S Eliasson L Galvanovskis J Renstrom E Rorsman P Activation of Ca(2+)-dependent K(+) channels contributes to rhythmic firing of action potentials in mouse pancreatic beta cells J Gen Physiol 1999 114 759 770 10578013 PMC2230648 sourceSupported
• cites [217] Goforth PB Bertram R Khan FA Zhang M Sherman A Satin LS Calcium-activated K+ channels of mouse beta-cells are controlled by both store and cytoplasmic Ca2+: experimental and theoretical studies J Gen Physiol 2002 120 307 322 12198088 PMC2229522 sourceDisputed

  The Discussion describes the proposal that dispersed β-cells have less Kslow and that this explains their altered electrical activity, but rejects that explanation. It concludes that differences in Kslow amplitude are not sufficient to explain the electrical differences between dispersed and in situ β-cells. (LLM, conf=0.90)

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The enhanced GIIS in Kcnk16/TALK1-deficient mice correlates with a moderate improvement of glucose tolerance in vivo.

“This correlates with a moderate improvement of glucose tolerance in vivo (708).”

• cites [708] Vierra NC Dadi PK Jeong I Dickerson M Powell DR Jacobson DA Type 2 Diabetes-Associated K+ Channel TALK-1 Modulates beta-Cell Electrical Excitability, Second-Phase Insulin Secretion, and Glucose Homeostasis Diabetes 2015 64 3818 3828 26239056 PMC4613978 sourceDisputed

  The Results section (‘TALK-1 Channels Are Critical for Maintaining Fasting Glycemia’; page not provided) explicitly reports no altered glucose tolerance in chow-fed TALK-1 knockout mice. Although the paper reports increased glucose-stimulated insulin secretion in isolated knockout islets and lower fasting glucose after a high-fat diet, it does not report the claimed improvement in glucose tolerance in vivo. (LLM, conf=0.87)

  Checked by a reasoning model — may be wrong.

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The non-vesicular mode of GABA release is remarkably effective, as the rate of release of GABA into culture medium suggests that GABA stores in β-cells turn over 30 times during 24 hours.

“The non-vesicular mode of release is remarkably effective, as the rate of release of GABA into the culture medium suggests that the GABA stores in β-cells turn over 30 times during 24 hours (629).”

• cites [629] Smismans A Schuit F Pipeleers D Nutrient regulation of gamma-aminobutyric acid release from islet beta cells Diabetologia 1997 40 1411 1415 9447948 sourceDisputed

  The Results and Discussion report that rat β-cells released about five times their cellular GABA content over 24 hours, not 30 times. The Discussion also describes direct, non-vesicular transport as a hypothetical route rather than an established explanation for that release rate. (LLM, conf=0.90)

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The interaction of Munc18 with syntaxin plays a key role in priming insulin granules for release.

“The interaction of Munc18 with syntaxin plays a key role in priming insulin granules for release (196).”

• cites [196] Gandasi NR Barg S Contact-induced clustering of syntaxin and munc18 docks secretory granules at the exocytosis site Nat Commun 2014 5 3914 24835618 sourceDisputed

  The ‘Clustering of other proteins at the docking site’ section says Munc18 binds syntaxin and is required for granule docking. The Discussion distinguishes this early recruitment from the later recruitment of Munc13 and SNAP25, whose timing is consistent with a role in priming. The paper therefore assigns the Munc18–syntaxin interaction a docking role, rather than the claimed priming role. (LLM, conf=0.83)

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Stimulation of basal secretion by miR-124 may be due to an increase in [Ca2+]i resulting from decreased KATP channel activity and stimulation of electrical activity.

“Stimulation of basal secretion may be due to an increase in [Ca2+]i resulting from decreased KATP channel activity and stimulation of electrical activity (53).”

• cites [53] Baroukh N Ravier MA Loder MK Hill EV Bounacer A Scharfmann R Rutter GA Van Obberghen E MicroRNA-124a regulates Foxa2 expression and intracellular signaling in pancreatic beta-cell lines J Biol Chem 2007 282 19575 19588 17462994 sourceDisputed

  The ‘Impact of miR-124a2 on Stimulus-dependent Ca²⁺’ section proposes that increased basal Ca²⁺ might activate insulin release, but then reports that the Ca²⁺ changes produced no difference in basal or stimulated hormone release. Its Conclusions likewise state that the authors could not detect a change in regulated insulin secretion. The proposed mechanism therefore does not support the claim that miR-124 stimulated basal secretion. (LLM, conf=0.88)

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Phosphorylation of glucose by glucokinase is the rate-limiting step in insulin secretion.

“Phosphorylation of glucose is the rate-limiting step in insulin secretion and loss-of-function mutations in GCK impair GIIS, with heterozygous mutations leading to maturity–onset-diabetes-of-the-young (MODY) and homozygous mutations to neonatal diabetes (214).”

• cites [214] Gloyn AL Odili S Zelent D Buettger C Castleden HA Steele AM Stride A Shiota C Magnuson MA Lorini R d'Annunzio G Insights into the structure and regulation of glucokinase from a novel mutation (V62M), which causes maturity-onset diabetes of the young J Biol Chem 2005 280 14105 14113 15677479 sourcePartly supported

  The introductory passage (page unavailable) calls glucokinase the pancreatic β-cell glucose sensor and says its kinetics let β-cells change glucose phosphorylation rates across physiological glucose concentrations. It also links inactivating GCK mutations to MODY, and the figure caption identifies the affected family members as heterozygous. The copy does not establish that phosphorylation is the rate-limiting step in insulin secretion or support the sentence’s claim about homozygous mutations causing neonatal diabetes. (LLM, conf=0.83)

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β-cell mass in T2DM may have been underestimated due to equating a reduction in insulin staining/content with a reduction in β-cell number.

“Furthermore, there is evidence that β-cell mass in T2DM may have been underestimated due to equating a reduction in insulin staining/content with a reduction in β-cell number (97, 429, 667).”

• cites [97] Brereton MF Rohm M Shimomura K Holland C Tornovsky-Babeay S Dadon D Iberl M Chibalina MV Lee S Glaser B Dor Y Hyperglycaemia induces metabolic dysfunction and glycogen accumulation in pancreatic beta-cells Nat Commun 2016 7 13496 27882918 PMC5123088 sourcePartly supported

  The Results (‘Diabetes duration impacts β-cell function’; page unavailable) define β-cell mass in diabetic mice by the extent of insulin staining, while the Discussion reports similar changes in insulin content or staining in T2DM. This supports the concern that staining need not directly measure cell number, but the paper does not establish that β-cell number or mass in people with T2DM has been underestimated. (LLM, conf=0.82)

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• cites [429] Marselli L Suleiman M Masini M Campani D Bugliani M Syed F Martino L Focosi D Scatena F Olimpico F Filipponi F Are we overestimating the loss of beta cells in type 2 diabetes? Diabetologia 2014 57 362 365 24233056 sourceSupported
• cites [667] Talchai C Xuan S Lin HV Sussel L Accili D Pancreatic beta cell dedifferentiation as a mechanism of diabetic beta cell failure Cell 2012 150 1223 1234 22980982 PMC3445031 sourcePartly supported

  The Results sections on cell-autonomous dedifferentiation and relevance to type 2 diabetes (pages unavailable) show that cells descended from β-cells can lose insulin staining while endocrine mass persists in diabetic mice. This supports the proposed reason an insulin-based estimate could be too low, but the paper does not establish that β-cell mass estimates in people with T2DM have been underestimated. (LLM, conf=0.82)

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Almost all glucose entering glycolysis proceeds into the Krebs cycle in β-cells.

“An important aspect of glucose metabolism in the β-cell is that almost all the glucose entering glycolysis proceeds into the Krebs cycle (602) (520).”

• cites [602] Schuit F De Vos A Farfari S Moens K Pipeleers D Brun T Prentki M Metabolic fate of glucose in purified islet cells. Glucose-regulated anaplerosis in beta cells J Biol Chem 1997 272 18572 18579 9228023 sourceSupported
• cites [520] Prentki M Matschinsky FM Madiraju SR Metabolic signaling in fuel-induced insulin secretion Cell Metab 2013 18 162 185 23791483 sourcePartly supported

  The passage labeled Abstract says that glucose carbons are largely converted to pyruvate, which is primarily used in the citric acid cycle. This supports the pathway described, but does not establish the stronger claim that almost all glucose entering glycolysis proceeds into that cycle. (LLM, conf=0.84)

  Experimental — accuracy not yet measured

The KATP channel complex has SUR1 subunits arranged around a central tetrameric Kir6.2 pore.

“The structure of the KATP channel complex reveals the SUR1 subunits are arranged around a central tetrameric Kir6.2 pore (393, 432, 450) (Figure 7B,C).”

• cites [393] Li N Wu JX Ding D Cheng J Gao N Chen L Structure of a Pancreatic ATP-Sensitive Potassium Channel Cell 2017 168 101 110 e110 28086082 sourceSupported
• cites [432] Martin GM Yoshioka C Rex EA Fay JF Xie Q Whorton MR Chen JZ Shyng SL Cryo-EM structure of the ATP-sensitive potassium channel illuminates mechanisms of assembly and gating Elife 2017 6 PMC5344670 28092267 sourceSupported
• cites [450] Mikhailov MV Campbell JD de Wet H Shimomura K Zadek B Collins RF Sansom MS Ford RC Ashcroft FM 3-D structural and functional characterization of the purified KATP channel complex Kir6.2-SUR1 EMBO J 2005 24 4166 4175 16308567 PMC1356316 sourceSupported

The number of KATP channels per β-cell is not known with certainty because open probability varies with measurement conditions.

“However, the open probability varies considerably according to how it is measured: in cell-attached patches it varies with the intracellular ATP concentration, while in inside-out patches it diminishes with time after patch excision (rundown; (526). Accordingly, the number of KATP channels per β-cell is not known with certainty.”

• cites [526] Proks P Puljung MC Vedovato N Sachse G Mulvaney R Ashcroft FM Running out of time: the decline of channel activity and nucleotide activation in adenosine triphosphate-sensitive K-channels Philos Trans R Soc Lond B Biol Sci 2016 371 PMC4938026 27377720 sourceSupported

Both Kir6.2 and SUR1 subunits are required for correct trafficking of the KATP channel to the plasma membrane.

“Both subunits are also required for correct trafficking of the channel to the plasma membrane (757).”

• cites [757] Zerangue N Schwappach B Jan YN Jan LY A new ER trafficking signal regulates the subunit stoichiometry of plasma membrane K(ATP) channels Neuron 1999 22 537 548 10197533 sourceSupported

Canine and porcine β-cells show no obvious oscillatory electrical activity in membrane potential recordings.

“Membrane potential recordings have also been performed on canine and porcine β-cells (454, 455). These revealed no obvious oscillatory electrical activity but as these measurements were made in dispersed β-cells, it remains unclear whether this reflects a true species difference or if it is simply a consequence of cell isolation (as in the mouse).”

• cites [454] Misler S Silva AM Barnett D Dickey AS Phasic and tonic modes of depolarization-exocytosis coupling in beta-cells of porcine islets of Langerhans Channels (Austin) 2009 3 101 109 19242115 sourcePartly supported

  The Results (pp. 5–6) report membrane-potential recordings from porcine β-cells showing irregular action potentials and intermittent plateau depolarizations, rather than describing a regular oscillatory pattern. The Discussion (p. 8) confirms that recordings were made from isolated cells and cautions against extrapolating to intact islets. The paper does not explicitly conclude that porcine cells show no obvious oscillatory electrical activity; the canine portion is not the part this citation appears to support. (LLM, conf=0.76)

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• cites [455] Misler S Zhou Z Dickey AS Silva AM Pressel DM Barnett DW Electrical activity and exocytotic correlates of biphasic insulin secretion from beta-cells of canine islets of Langerhans: contribution of tuning two modes of Ca2+ entry-dependent exocytosis to two modes of glucose-induced electrical activity Channels (Austin) 2009 3 181 193 19535907 sourceNot checked — no full text of the source

Measurements of channel activity in cell-attached patches before and after permeabilization with α-toxin, followed by dialysis with different ATP concentrations, yield submembrane ATP values of ~1 mM in glucose-free solution and ~3 mM in 11 mM glucose.

“However, measurements of channel activity in cell-attached patches before and after permeabilization of the plasma membrane with α-toxin, followed by dialysis with different ATP concentrations, yield values of ~1 mM in glucose-free solution and ~3 mM in 11 mM glucose (674).”

• cites [674] Tarasov AI Girard CA Ashcroft FM ATP sensitivity of the ATP-sensitive K+ channel in intact and permeabilized pancreatic beta-cells Diabetes 2006 55 2446 2454 16936192 sourceSupported

SUR1 regulates the activity of the Kir6.2 pore, endowing it with sensitivity to inhibitory sulfonylurea drugs, K+ channel openers such as diazoxide, and stimulatory effects of Mg-nucleotides.

“Instead, it regulates the activity of the Kir6.2 pore, endowing it with sensitivity to inhibitory sulfonylurea drugs (hence its name), to K+ channel openers such as diazoxide, and to the stimulatory effects of Mg-nucleotides (5, 232, 621, 698).”

• cites [5] Aguilar-Bryan L Nichols CG Wechsler SW Clement JPt Boyd AE 3rd Gonzalez G Herrera-Sosa H Nguy K Bryan J Nelson DA Cloning of the beta cell high-affinity sulfonylurea receptor: a regulator of insulin secretion Science 1995 268 423 426 7716547 sourceNot checked — no full text of the source
• cites [232] Gribble FM Tucker SJ Ashcroft FM The essential role of the Walker A motifs of SUR1 in K-ATP channel activation by Mg-ADP and diazoxide EMBO J 1997 16 1145 1152 9135131 PMC1169713 sourceSupported
• cites [621] Shyng S Ferrigni T Nichols CG Regulation of KATP channel activity by diazoxide and MgADP. Distinct functions of the two nucleotide binding folds of the sulfonylurea receptor J Gen Physiol 1997 110 643 654 9382893 PMC2229399 sourceSupported

Metabolic regulation of the KATP channel is mediated by changes in intracellular concentrations of ATP and MgADP, which inhibit and stimulate the channel, respectively.

“Metabolic regulation of the channel is mediated by changes in the intracellular concentrations of ATP and MgADP, which inhibit and stimulate the channel, respectively (127, 163, 328).”

• cites [127] Cook DL Hales CN Intracellular ATP directly blocks K+ channels in pancreatic B-cells Nature 1984 311 271 273 6090930 sourceNot checked — no full text of the source
• cites [163] Dunne MJ Petersen OH Intracellular ADP activates K+ channels that are inhibited by ATP in an insulin-secreting cell line FEBS Lett 1986 208 59 62 2429868 sourcePartly supported

  The Results report that ADP activated ATP-inhibited K+ channels in permeabilized RINmSF cells and that ATP closed ADP-activated channels. The Discussion proposes that changes in the ATP/ADP ratio regulate channel opening after metabolic stimulation. The paper identifies the activating substance as ADP, however, and does not establish that MgADP specifically mediates the effect. (LLM, conf=0.84)

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• cites [328] Kakei M Kelly RP Ashcroft SJ Ashcroft FM The ATP-sensitivity of K+ channels in rat pancreatic B-cells is modulated by ADP FEBS Lett 1986 208 63 66 2429869 sourceNot checked — no full text of the source

The islet's response to glucose is also due to recruitment of an increasing number of β-cells.

“Thus, although every β-cell has the capacity to respond to glucose in a graded fashion, the islet’s response to glucose is also due to recruitment of an increasing number of β-cells (283, 402).”

• cites [283] Hiriart M Matteson DR Na channels and two types of Ca channels in rat pancreatic B cells identified with the reverse hemolytic plaque assay J Gen Physiol 1988 91 617 639 2458427 PMC2216150 sourceSupported
• cites [402] Low JT Mitchell JM Do OH Bax J Rawlings A Zavortink M Morgan G Parton RG Gaisano HY Thorn P Glucose principally regulates insulin secretion in mouse islets by controlling the numbers of granule fusion events per cell Diabetologia 2013 56 2629 2637 23995471 PMC3825531 sourceSupported

At 20 mM glucose, the time spent above -20 mV (the voltage where insulin granule exocytosis first becomes detectable) is ~30 ms.

“Its width is somewhat variable but, at 20 mM glucose, the time spent above -20 mV (the voltage where insulin granule exocytosis first becomes detectable (222)) is ~30 ms (288).”

• cites [222] Gopel S Zhang Q Eliasson L Ma XS Galvanovskis J Kanno T Salehi A Rorsman P Capacitance measurements of exocytosis in mouse pancreatic alpha-, beta- and delta-cells within intact islets of Langerhans J Physiol 2004 556 711 726 14966302 PMC1664984 sourceNot checked — no full text of the source
• cites [288] Hoppa MB Collins S Ramracheya R Hodson L Amisten S Zhang Q Johnson P Ashcroft FM Rorsman P Chronic palmitate exposure inhibits insulin secretion by dissociation of Ca(2+) channels from secretory granules Cell Metab 2009 10 455 465 19945403 PMC2814048 sourcePartly supported

  The Results section, under “Pharmacological Correction of the Inhibitory Action of Palmitate Exposure on Insulin Secretion” (page unavailable), reports action potentials averaging 31 ± 1 ms at 20 mM glucose, supporting the approximately 30 ms figure. It does not specify that this duration was measured as time above −20 mV; the citing sentence attaches a separate citation to the exocytosis-threshold explanation. (LLM, conf=0.85)

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Amino acids, neurotransmitters and hormones can stimulate electrical activity and insulin release in human β-cells only in the presence of a permissive concentration of glucose.

“Amino acids (arginine and glycine)(744), neurotransmitters (like adenine nucleotides, GABA and acetylcholine) (89, 92, 347, 568) and hormones (618) can also stimulate electrical activity and insulin release but only in the presence of a ‘permissive’ concentration of glucose.”

• cites [744] Yan-Do R Duong E Manning Fox JE Dai X Suzuki K Khan S Bautista A Ferdaoussi M Lyon J Wu X Cheley S A Glycine-Insulin Autocrine Feedback Loop Enhances Insulin Secretion From Human beta-Cells and Is Impaired in Type 2 Diabetes Diabetes 2016 65 2311 2321 27207556 sourceNot checked — no full text of the source
• cites [89] Braun M Ramracheya R Bengtsson M Clark A Walker JN Johnson PR Rorsman P Gamma-aminobutyric acid (GABA) is an autocrine excitatory transmitter in human pancreatic beta-cells Diabetes 2010 59 1694 1701 20413510 PMC2889769 sourceSupported
• cites [618] Shigeto M Ramracheya R Tarasov AI Cha CY Chibalina MV Hastoy B Philippaert K Reinbothe T Rorsman N Salehi A Sones WR GLP-1 stimulates insulin secretion by PKC-dependent TRPM4 and TRPM5 activation J Clin Invest 2015 125 4714 4728 26571400 PMC4665783 sourceSupported

T2DM human islets accumulate large amounts of glycogen, like diabetic mouse β-cells.

“This idea is supported by the fact that like diabetic mouse β-cells (97), T2DM human islets accumulate large amounts of glycogen (97, 685).”

• cites [97] Brereton MF Rohm M Shimomura K Holland C Tornovsky-Babeay S Dadon D Iberl M Chibalina MV Lee S Glaser B Dor Y Hyperglycaemia induces metabolic dysfunction and glycogen accumulation in pancreatic beta-cells Nat Commun 2016 7 13496 27882918 PMC5123088 sourceSupported
• cites [685] Toreson WE Glycogen infiltration (so-called hydropic degeneration) in the pancreas in human and experimental diabetes mellitus Am J Pathol 1951 27 327 347 14819264 PMC1937312Not checked — no full text of the source

Glucose-induced insulin secretion follows a biphasic time course in both mouse and human islets.

“As will be discussed at length below (Section IX), glucose-induced insulin secretion (GIIS) follows a biphasic time course in both mouse and human islets.”

• cites [473] Nesher R Cerasi E Modeling phasic insulin release: immediate and time-dependent effects of glucose Diabetes 2002 51 Suppl 1 S53 59 11815459 sourcePartly supported

  The opening passage in Section 2 describes a biphasic response to a square-wave glucose stimulus in perifused rat islets, perfused rat pancreas, and humans. It does not establish that the response was observed in mouse islets or isolated human islets, as the claim specifies. The section on biphasic insulin release also notes that clear phasicity is not detected after oral glucose. (LLM, conf=0.87)

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Loss-of-function mutations in KATP channel genes cause congenital hyperinsulinism because they result in permanent membrane depolarization and persistent insulin release.

“Loss-of-function mutations cause congenital hyperinsulinism because they result in permanent membrane depolarization and persistent insulin release (479, 644, 681).”

• cites [479] Nichols CG Remedi MS The diabetic beta-cell: hyperstimulated vs. hyperexcited Diabetes Obes Metab 2012 14 Suppl 3 129 135 22928573 PMC3712635 sourcePartly supported

  The Human Diseases section (page unavailable) identifies loss-of-function mutations in KATP genes as major causes of congenital hyperinsulinism. The Glucose Stimulation section links KATP inhibition to membrane depolarization and insulin secretion, but the Chronic Hyperexcitability section reports that severe or prolonged loss of KATP activity can instead reduce insulin secretion. Thus, permanent depolarization with persistent insulin release is too categorical. (LLM, conf=0.88)

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• cites [644] Stanley CA Perspective on the Genetics and Diagnosis of Congenital Hyperinsulinism Disorders J Clin Endocrinol Metab 2016 101 815 826 26908106 PMC4803157 sourcePartly supported

  The KATP-HI form section says inactivating KATP-channel mutations cause congenital hyperinsulinism. The Pathways of β-Cell Insulin Secretion section explains that KATP-channel closure depolarizes the membrane and triggers insulin release. The paper does not establish permanent depolarization for all such mutations; it describes some dominant mutations with residual channel activity. (LLM, conf=0.83)

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• cites [681] Thomas PM Cote GJ Wohllk N Haddad B Mathew PM Rabl W Aguilar-Bryan L Gagel RF Bryan J Mutations in the sulfonylurea receptor gene in familial persistent hyperinsulinemic hypoglycemia of infancy Science 1995 268 426 429 7716548 sourceNot checked — no full text of the source

Sulfonylurea drugs provide an effective therapy in >90% of patients with neonatal diabetes due to KATP mutations.

“Sulfonylurea drugs, which bypass metabolism and block the channel directly, provide an effective therapy in >90% of patients (44, 504).”

• cites [44] Babiker T Vedovato N Patel K Thomas N Finn R Mannikko R Chakera AJ Flanagan SE Shepherd MH Ellard S Ashcroft FM Successful transfer to sulfonylureas in KCNJ11 neonatal diabetes is determined by the mutation and duration of diabetes Diabetologia 2016 59 1162 1166 27033559 PMC4869695 sourcePartly supported

  The Introduction says sulfonylureas directly close KATP channels, and the Discussion reports that most patients with KCNJ11-related neonatal diabetes achieved near-normal glycaemia after switching. But the reported success rate is 88%, not >90%, and the study does not establish that rate for all KATP mutations. (LLM, conf=0.90)

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• cites [504] Pearson ER Flechtner I Njolstad PR Malecki MT Flanagan SE Larkin B Ashcroft FM Klimes I Codner E Iotova V Slingerland AS Switching from insulin to oral sulfonylureas in patients with diabetes due to Kir6.2 mutations N Engl J Med 2006 355 467 477 16885550 sourcePartly supported

  The Abstract reports that 44 of 49 patients (90%) with Kir6.2 mutations successfully discontinued insulin after receiving sulfonylureas. It also says sulfonylureas close KATP channels by an ATP-independent route. The study therefore supports the mechanism and effectiveness for Kir6.2 mutations, but not the claim of effectiveness in >90% of patients with KATP mutations generally. (LLM, conf=0.88)

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In both whole-cell and perforated-patch preparations, glucose produces a concentration-dependent reduction of whole-cell KATP conductance that is half-maximal at 3-4 mM glucose.

“In both preparations, glucose produces a concentration-dependent reduction of the whole-cell conductance which, when expressed as a fraction of that in the absence of glucose, is half-maximal at 3-4 mM glucose (26, 212, 584, 674, 762).”

• cites [26] Ashcroft FM Ashcroft SJ Harrison DE Properties of single potassium channels modulated by glucose in rat pancreatic beta-cells J Physiol 1988 400 501 527 2458459 PMC1191821 sourcePartly supported

  The paper’s numbered findings report a reversible, dose-dependent decrease in G-channel activity with glucose, accompanied by increased cell input resistance. But the experiments described use single-channel recordings from cell-attached patches; our copy does not establish the claimed result in both whole-cell and perforated-patch preparations or a half-maximal concentration of 3–4 mM. (LLM, conf=0.86)

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• cites [212] Girard CA Wunderlich FT Shimomura K Collins S Kaizik S Proks P Abdulkader F Clark A Ball V Zubcevic L Bentley L Expression of an activating mutation in the gene encoding the KATP channel subunit Kir6.2 in mouse pancreatic beta cells recapitulates neonatal diabetes J Clin Invest 2009 119 80 90 19065048 PMC2613450 sourcePartly supported

  The results (p. 3) report half-maximal glucose inhibition at 3 mM in wild-type β cells, and Figure 6 (p. 7) shows those whole-cell currents were recorded using perforated patches and expressed relative to glucose-free current. The paper does not establish the same value in a separate whole-cell preparation; it reports an IC50 above 20 mM for mutant β cells. (LLM, conf=0.88)

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• cites [584] Sakura H Ashcroft SJ Terauchi Y Kadowaki T Ashcroft FM Glucose modulation of ATP-sensitive K-currents in wild-type, homozygous and heterozygous glucokinase knock-out mice Diabetologia 1998 41 654 659 9662046 sourcePartly supported

  The Results report concentration-dependent inhibition of KATP current in neonatal wild-type beta cells, with an EC50 of 3.2 mmol/l; the Statistical analysis section says conductance was expressed relative to glucose-free controls (pages unavailable in our copy). The paper also describes both recording configurations, but it does not report a 3–4 mmol/l EC50 for standard whole-cell recordings. Its EC50 values differ for heterozygous and older cells, so the quoted range cannot be applied to all cells studied. (LLM, conf=0.86)

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Coexpressing SUR1 with Kir6.2 carrying a mutation that renders it ATP-insensitive yields EC50 of 8 μM for MgADP and 112 μM for MgATP.

“However, it can be estimated by coexpressing SUR1 with Kir6.2 carrying a mutation that renders it ATP-insensitive. This yields EC50 of 8 µM for MgADP and 112 µM for MgATP (523): it is believed that the lower affinity of MgATP reflects the fact that it must be hydrolyzed to MgADP before it can stimulate channel opening.”

• cites [523] Proks P de Wet H Ashcroft FM Activation of the K(ATP) channel by Mg-nucleotide interaction with SUR1 J Gen Physiol 2010 136 389 405 20876358 PMC2947056 sourceSupported

A cocktail of isradipine, SNX482, ω-contoxin and ω-agatoxin almost completely inhibits the β-cell whole-cell Ca2+ current.

“A cocktail of isradipine, SNX482, ω-contoxin and ω-agatoxin almost completely inhibits the β-cell whole-cell Ca2+ current (603).”

• cites [603] Schulla V Renstrom E Feil R Feil S Franklin I Gjinovci A Jing XJ Laux D Lundquist I Magnuson MA Obermuller S Impaired insulin secretion and glucose tolerance in beta cell-selective Ca(v)1.2 Ca2+ channel null mice EMBO J 2003 22 3844 3854 12881419 PMC169062 sourcePartly supported

  The Results report that isradipine reduced β-cell Ca2+ current by 53 ± 6%, and the Figure 2 caption describes adding SNX482 after isradipine. Our copy does not report testing ω-conotoxin or ω-agatoxin in that combination, or show that a four-drug cocktail almost completely inhibited the whole-cell current. (LLM, conf=0.85)

  Experimental — accuracy not yet measured

Parallel measurements of the cytosolic ATP/ADP-ratio and KATP channel activity have provided direct evidence for an inverse correlation between these two parameters.

“Indeed, parallel measurements of the cytosolic ATP/ADP-ratio and KATP channel activity have provided direct evidence for an inverse correlation between these two parameters (675).”

• cites [675] Tarasov AI Griffiths EJ Rutter GA Regulation of ATP production by mitochondrial Ca(2+) Cell Calcium 2012 52 28 35 22502861 PMC3396849 sourcePartly supported

  The pancreatic islet β-cell section (page unavailable) says that increases in cytosolic ATP/ADP ratio block KATP channels, supporting an inverse relationship. It does not report parallel measurements of the ratio and channel activity as direct evidence; the combined measurements it describes are of ATP/ADP and cytosolic Ca2+. (LLM, conf=0.84)

  Experimental — accuracy not yet measured

The KATP current of dispersed β-cells is not fully inhibited until glucose reaches ~20 mM.

“Similarly, the KATP current of dispersed β-cells is not fully inhibited until glucose reaches ~20 mM (212, 674) (Figure 2C).”

• cites [212] Girard CA Wunderlich FT Shimomura K Collins S Kaizik S Proks P Abdulkader F Clark A Ball V Zubcevic L Bentley L Expression of an activating mutation in the gene encoding the KATP channel subunit Kir6.2 in mouse pancreatic beta cells recapitulates neonatal diabetes J Clin Invest 2009 119 80 90 19065048 PMC2613450 sourcePartly supported

  Figure 6 (p. 7) reports glucose-dependent KATP currents in isolated mouse β-cells. The accompanying text (p. 3) says the mutant cells’ glucose-inhibition IC50 was greater than 20 mM, versus 3 mM in wild-type cells. Thus the paper supports incomplete inhibition at 20 mM in mutant cells, but not the claim that dispersed β-cell currents generally become fully inhibited at about 20 mM. (LLM, conf=0.86)

  Experimental — accuracy not yet measured
• cites [674] Tarasov AI Girard CA Ashcroft FM ATP sensitivity of the ATP-sensitive K+ channel in intact and permeabilized pancreatic beta-cells Diabetes 2006 55 2446 2454 16936192 sourceSupported

Experiments on Kcnb1-deficient mice make it clear that Kv2.1 channels account for most of the delayed outward current in mouse β-cells.

“However, experiments on Kcnb1-deficient mice make it clear that these channels account for most of the delayed outward current in mouse β-cells (310).”

• cites [310] Jacobson DA Kuznetsov A Lopez JP Kash S Ammala CE Philipson LH Kv2.1 ablation alters glucose-induced islet electrical activity, enhancing insulin secretion Cell Metab 2007 6 229 235 17767909 PMC2699758 sourceSupported

There is evidence that the T2DM risk associated with the KCNQ1 locus is mediated through another gene in the imprinted 11p15.5 region of the chromosome.

“There is now evidence that the T2DM risk associated with the KCNQ1 locus is mediated through another gene in the imprinted 11p15.5 region of the chromosome (692).”

• cites [692] Travers ME Mackay DJ Dekker Nitert M Morris AP Lindgren CM Berry A Johnson PR Hanley N Groop LC McCarthy MI Gloyn AL Insights into the molecular mechanism for type 2 diabetes susceptibility at the KCNQ1 locus from temporal changes in imprinting status in human islets Diabetes 2013 62 987 992 23139357 PMC3581222 sourceSupported

The mouse β-cell Na+ channel is highly unusual in the voltage dependence of its inactivation; in most β-cells it inactivates at such negative membrane potentials that little or no Na+ current remains within the physiological range.

“The mouse (and rat) β-cell Na+ channel is highly unusual in the voltage dependence of its inactivation. In most β-cells, it inactivates at such negative membrane potentials that little or no Na+ current remains within the physiological range of membrane potentials (221, 514, 761) (Figure 10).”

• cites [221] Gopel S Kanno T Barg S Galvanovskis J Rorsman P Voltage-gated and resting membrane currents recorded from B-cells in intact mouse pancreatic islets J Physiol 1999 521 Pt 3 717 728 10601501 PMC2269694 sourceNot checked — no full text of the source
• cites [514] Plant TD Na+ currents in cultured mouse pancreatic B-cells Pflugers Arch 1988 411 429 435 2456517 sourceNot checked — no full text of the source
• cites [761] Zhang Q Chibalina MV Bengtsson M Groschner LN Ramracheya R Rorsman NJ Leiss V Nassar MA Welling A Gribble FM Reimann F Na+ current properties in islet alpha- and beta-cells reflect cell-specific Scn3a and Scn9a expression J Physiol 2014 592 4677 4696 25172946 PMC4253470 sourceSupported

In contrast to mouse β-cells, the delayed rectifying K+ current in human β-cells is carried principally by Kv2.2 (KCNB2) and Kv1.6 (KCNA6) channels.

“In contrast to mouse β-cells, the delayed rectifying K+ current in human β-cells is carried principally by Kv2.2 (KCNB2) and Kv1.6 (KCNA6) channels (Appendix Figure 5A), which are expressed at high levels in the human β-cell and encode inactivating TEA-sensitive voltage-gated K+ channels (68, 475).”

• cites [68] Blodgett DM Nowosielska A Afik S Pechhold S Cura AJ Kennedy NJ Kim S Kucukural A Davis RJ Kent SC Greiner DL Novel Observations From Next-Generation RNA Sequencing of Highly Purified Human Adult and Fetal Islet Cell Subsets Diabetes 2015 64 3172 3181 25931473 PMC4542439 sourceCould not verify

  The Results section on enriched transcripts reports β-cell expression of KCNQ2, but does not address Kv2.2 (KCNB2) or Kv1.6 (KCNA6). Searches for both genes and for delayed-rectifier currents and TEA sensitivity found no passage establishing which channels principally carry the human β-cell current or comparing it with mouse β-cells. Page numbers were not available in our copy. (LLM, conf=0.76)

  Experimental — accuracy not yet measured
• cites [475] Nica AC Ongen H Irminger JC Bosco D Berney T Antonarakis SE Halban PA Dermitzakis ET Cell-type, allelic, and genetic signatures in the human pancreatic beta cell transcriptome Genome Res 2013 23 1554 1562 23716500 PMC3759730 sourceCould not verify

  The Results (page unavailable) describe gene-expression differences among human β-cells, islets, and non-β-cells, but do not identify which channels carry the delayed rectifying K+ current. Searches for KCNB2, KCNA6, the channel names, and a human–mouse comparison found no passage addressing the claim. (LLM, conf=0.76)

  Experimental — accuracy not yet measured

Syntaxin-1 interferes with the trafficking of Kv2.1 to the plasma membrane.

“Syntaxin-1 also interferes with the trafficking of Kv2.1 to the plasma membrane (386).”

• cites [386] Leung YM Kang Y Gao X Xia F Xie H Sheu L Tsuk S Lotan I Tsushima RG Gaisano HY Syntaxin 1A binds to the cytoplasmic C terminus of Kv2.1 to regulate channel gating and trafficking J Biol Chem 2003 278 17532 17538 12621036 sourceSupported

Human β-cells mainly express SCN8A (Nav1.6), accounting for 40% of Na+ channel transcripts, with the remainder being SCN9A (Nav1.7), SCN3A (Nav1.3) and SCN2A (Nav1.2).

“Whereas mouse β-cells mainly express Scn9a (Nav1.7), SCN8A (Nav1.6) accounts for 40% of Na+ channel transcripts in human β-cells (68, 475) with the remainder being SCN9A (Nav1.7), SCN3A (Nav1.3) and SCN2A (Nav1.2) (see Appendix Figure 4).”

• cites [68] Blodgett DM Nowosielska A Afik S Pechhold S Cura AJ Kennedy NJ Kim S Kucukural A Davis RJ Kent SC Greiner DL Novel Observations From Next-Generation RNA Sequencing of Highly Purified Human Adult and Fetal Islet Cell Subsets Diabetes 2015 64 3172 3181 25931473 PMC4542439 sourceCould not verify

  The Results section on enriched α- and β-cell transcripts reports β-cell gene expression but does not give the claimed sodium-channel breakdown. Searches for SCN8A, Nav1.6, and the 40% figure found no passage in our copy stating the proportions or identifying SCN9A, SCN3A, and SCN2A as the remainder; page numbers were not available. (LLM, conf=0.72)

  Experimental — accuracy not yet measured
• cites [475] Nica AC Ongen H Irminger JC Bosco D Berney T Antonarakis SE Halban PA Dermitzakis ET Cell-type, allelic, and genetic signatures in the human pancreatic beta cell transcriptome Genome Res 2013 23 1554 1562 23716500 PMC3759730 sourceCould not verify

  The Results passages in our copy (page not provided) describe human β-cell transcript profiles but do not give a breakdown of sodium-channel transcripts. Searches for SCN8A, SCN9A, Nav1.6, and the claimed 40% figure found no passage addressing the claim. (LLM, conf=0.75)

  Experimental — accuracy not yet measured

TALK-1 stabilizes the interburst intervals.

“Collectively, these observations suggest that TALK-1 stabilizes the interburst intervals.”

• cites [708] Vierra NC Dadi PK Jeong I Dickerson M Powell DR Jacobson DA Type 2 Diabetes-Associated K+ Channel TALK-1 Modulates beta-Cell Electrical Excitability, Second-Phase Insulin Secretion, and Glucose Homeostasis Diabetes 2015 64 3818 3828 26239056 PMC4613978 sourceSupported

[Ca2+]i buffering plays an important role in Kslow regulation.

“This suggests [Ca2+]i buffering plays an important role in Kslow regulation.”

• cites [217] Goforth PB Bertram R Khan FA Zhang M Sherman A Satin LS Calcium-activated K+ channels of mouse beta-cells are controlled by both store and cytoplasmic Ca2+: experimental and theoretical studies J Gen Physiol 2002 120 307 322 12198088 PMC2229522 sourceSupported

Following cessation of electrical activity, Kslow deactivates with a time course similar to the slow depolarization between two successive bursts of action potentials.

“Following cessation of electrical activity, the current deactivates with a time course similar to the slow depolarization between two successive bursts of action potentials (223).”

• cites [223] Gopel SO Kanno T Barg S Eliasson L Galvanovskis J Renstrom E Rorsman P Activation of Ca(2+)-dependent K(+) channels contributes to rhythmic firing of action potentials in mouse pancreatic beta cells J Gen Physiol 1999 114 759 770 10578013 PMC2230648 sourceSupported

β-cells from Knck3-deficient mice are slightly more depolarized in the presence of glucose than their wild-type counterparts.

“Despite the small size of the current, β-cells from Knck3-deficient mice were slightly more depolarized in the presence of glucose than their wild-type counterparts.”

• cites [135] Dadi PK Luo B Vierra NC Jacobson DA TASK-1 Potassium Channels Limit Pancreatic alpha-Cell Calcium Influx and Glucagon Secretion Mol Endocrinol 2015 29 777 787 25849724 PMC4415209 sourcePartly supported

  The section “TASK-1 currents regulate pancreatic α-cell Δp” reports that KCNK3/TASK-1-deficient cells were more depolarized than controls under glucose conditions. But the measured cells were α-cells, not β-cells; our copy does not establish the claimed β-cell result. No page number is available in the indexed passage. (LLM, conf=0.89)

  Experimental — accuracy not yet measured

The modulation of Kslow by glucose may underlie the shortening of the electrically silent inter-burst intervals that occurs with increasing glucose concentrations.

“The modulation of Kslow by glucose is of functional importance as it may underlie the shortening of the electrically silent inter-burst intervals that occurs with increasing glucose concentrations.”

• cites [217] Goforth PB Bertram R Khan FA Zhang M Sherman A Satin LS Calcium-activated K+ channels of mouse beta-cells are controlled by both store and cytoplasmic Ca2+: experimental and theoretical studies J Gen Physiol 2002 120 307 322 12198088 PMC2229522 sourcePartly supported

  The Results ('Modulation of Kslow by Glucose'; page unavailable) show that raising glucose from 5 to 10 mM reduces Kslow current density. The Introduction says increasing glucose shortens the silent phase, and the Results ('The Role of Kslow in Islet Bursting'; page unavailable) propose a role for Kslow in electrical oscillations. The paper does not specifically propose or test whether glucose's effect on Kslow accounts for that shortening. (LLM, conf=0.83)

  Experimental — accuracy not yet measured

The Kslow current is reduced by 50% when glucose is increased from 5 to 10 mM.

“The Kslow current is reduced by 50% when glucose is increased from 5 to 10 mM (217).”

• cites [217] Goforth PB Bertram R Khan FA Zhang M Sherman A Satin LS Calcium-activated K+ channels of mouse beta-cells are controlled by both store and cytoplasmic Ca2+: experimental and theoretical studies J Gen Physiol 2002 120 307 322 12198088 PMC2229522 sourceSupported

A polymorphism in KCNJ15 (encoding Kir4.2) increases the risk of T2DM more than 1.7-fold.

“In addition, a polymorphism in KCNJ15 (the gene encoding Kir4.2) increases the risk of T2DM >1.7-fold (522).”

• cites [522] Prokopenko I McCarthy MI Lindgren CM Type 2 diabetes: new genes, new understanding Trends Genet 2008 24 613 621 18952314 PMC7116807 sourceCould not verify

  The section “A leap forward in complex trait genetics” discusses KCNJ11 and a 15–20% increase in T2D risk per susceptibility allele, not KCNJ15 or Kir4.2. Searches for KCNJ15, Kir4.2, and a greater-than-1.7-fold risk found no passage in our copy addressing the claimed association. (LLM, conf=0.76)

  Experimental — accuracy not yet measured

Like their mouse counterparts, human β-cells predominantly express SK3 (KCNN3), with other SKs being expressed at 10-fold lower levels.

“Like their mouse counterparts, human β-cells predominantly express SK3 (KCNN3) with other SKs being expressed at 10-fold lower levels (68, 475).”

• cites [68] Blodgett DM Nowosielska A Afik S Pechhold S Cura AJ Kennedy NJ Kim S Kucukural A Davis RJ Kent SC Greiner DL Novel Observations From Next-Generation RNA Sequencing of Highly Purified Human Adult and Fetal Islet Cell Subsets Diabetes 2015 64 3172 3181 25931473 PMC4542439 sourceCould not verify

  The Results section on enriched α- and β-cell transcripts names KCNQ2 and KCND3, but does not report KCNN3 or compare expression of SK channels. Searches of our copy for KCNN3, SK3, and the claimed 10-fold difference found no passage addressing that comparison. (LLM, conf=0.72)

  Experimental — accuracy not yet measured
• cites [475] Nica AC Ongen H Irminger JC Bosco D Berney T Antonarakis SE Halban PA Dermitzakis ET Cell-type, allelic, and genetic signatures in the human pancreatic beta cell transcriptome Genome Res 2013 23 1554 1562 23716500 PMC3759730 sourceCould not verify

  The Results describe gene-expression profiles of human β-cells, but the passages in our copy do not compare SK-channel expression. Searches for KCNN3, SK3, and related channel terms found no passage establishing that SK3 predominates or that other SKs are expressed at 10-fold lower levels. (LLM, conf=0.75)

  Experimental — accuracy not yet measured

The effect of KCNJ15 on T2DM risk is probably mediated via other cell types.

“As KCNJ15 is expressed at low, albeit detectable, levels in human β-cells (Appendix Figure 7), and there is no evidence its expression is increased in diabetic β-cells (606), its effect on T2DM risk is probably mediated via other cell types.”

• cites [606] Segerstolpe A Palasantza A Eliasson P Andersson EM Andreasson AC Sun X Picelli S Sabirsh A Clausen M Bjursell MK Smith DM Single-Cell Transcriptome Profiling of Human Pancreatic Islets in Health and Type 2 Diabetes Cell Metab 2016 24 593 607 27667667 PMC5069352 sourceCould not verify

  The citation is attached to the statement that there is no evidence of increased KCNJ15 expression in diabetic β-cells; the proposed effect through other cell types is the citing authors’ inference. The Results section on gene-expression alterations in T2D discusses other β-cell genes but does not address KCNJ15. Searches for KCNJ15 and its expression in diabetic β-cells found no relevant passage in our copy; page numbers were unavailable. (LLM, conf=0.72)

  Experimental — accuracy not yet measured

Kslow is reduced in dispersed β-cells maintained in tissue culture.

“It has been reported that Kslow is reduced in dispersed β-cells maintained in tissue culture (223) which may account for the loss of normal oscillatory electrical activity seen in this preparation (but see (217)).”

• cites [223] Gopel SO Kanno T Barg S Eliasson L Galvanovskis J Renstrom E Rorsman P Activation of Ca(2+)-dependent K(+) channels contributes to rhythmic firing of action potentials in mouse pancreatic beta cells J Gen Physiol 1999 114 759 770 10578013 PMC2230648 sourceSupported

The SERCA inhibitor thapsigargin exerts a dual effect on Kslow, transiently increasing and then reducing its amplitude.

“It is interesting that the SERCA inhibitor thapsigargin exerts a dual effect on Kslow, transiently increasing and then reducing its amplitude (217).”

• cites [217] Goforth PB Bertram R Khan FA Zhang M Sherman A Satin LS Calcium-activated K+ channels of mouse beta-cells are controlled by both store and cytoplasmic Ca2+: experimental and theoretical studies J Gen Physiol 2002 120 307 322 12198088 PMC2229522 sourceSupported

The time and voltage-dependence of activation of Kslow echoes the changes in [Ca2+]i associated with simulated electrical activity.

“The time and voltage-dependence of activation of Kslow echoes the changes in [Ca2+]i associated with (simulated) electrical activity (223).”

• cites [223] Gopel SO Kanno T Barg S Eliasson L Galvanovskis J Renstrom E Rorsman P Activation of Ca(2+)-dependent K(+) channels contributes to rhythmic firing of action potentials in mouse pancreatic beta cells J Gen Physiol 1999 114 759 770 10578013 PMC2230648 sourceSupported

TASK1 activates around -30 mV in human β-cells.

“TASK1 activates around -30 mV in human β-cells (135).”

• cites [135] Dadi PK Luo B Vierra NC Jacobson DA TASK-1 Potassium Channels Limit Pancreatic alpha-Cell Calcium Influx and Glucagon Secretion Mol Endocrinol 2015 29 777 787 25849724 PMC4415209 sourceCould not verify

  The section ‘TASK-1 channels are functionally expressed in mouse and human α-cells’ reports functional TASK-1-like currents in human α-cells, not β-cells. A search for β-cell activation and −30 mV found no passage establishing the claimed activation voltage in human β-cells; the reported voltage ramp from −120 to 60 mV is not an activation threshold. Page numbers were unavailable in our copy. (LLM, conf=0.76)

  Experimental — accuracy not yet measured

In the presence of cAMP, activation of HCN channels is shifted to more positive membrane potentials, with half-maximal activation at -74 mV in the presence of forskolin and IBMX.

“In the presence of cAMP, however, activation of HCN channels is shifted to more positive membrane potentials (half maximal at -74 mV in the presence of forskolin and IBMX).”

• cites [168] El-Kholy W MacDonald PE Fox JM Bhattacharjee A Xue T Gao X Zhang Y Stieber J Li RA Tsushima RG Wheeler MB Hyperpolarization-activated cyclic nucleotide-gated channels in pancreatic beta-cells Mol Endocrinol 2007 21 753 764 17158221 sourceSupported

Kcnk16 (TALK1) gives rise to an outwardly rectifying current that becomes detectable at voltages above -45 mV.

“Kcnk16 (TALK1) gives rise to an outwardly rectifying current that becomes detectable at voltages above -45 mV (708).”

• cites [708] Vierra NC Dadi PK Jeong I Dickerson M Powell DR Jacobson DA Type 2 Diabetes-Associated K+ Channel TALK-1 Modulates beta-Cell Electrical Excitability, Second-Phase Insulin Secretion, and Glucose Homeostasis Diabetes 2015 64 3818 3828 26239056 PMC4613978 sourcePartly supported

  The Introduction (page unavailable) describes TALK-1 channels as producing outwardly rectifying K+ currents, and the Results section on functional expression (page unavailable) reports such a current in mouse β-cells that is reduced in TALK-1 knockouts. Neither passage establishes that the current becomes detectable specifically above −45 mV. (LLM, conf=0.87)

  Experimental — accuracy not yet measured

HCN channels may therefore contribute to the reported shortening of the interburst interval observed in β-cells of mouse islets exposed to high (50 nM) concentrations of GLP-1.

“HCN channels may therefore contribute to the reported shortening of the interburst interval observed in β-cells (211) of mouse islets exposed to high (50 nM) concentrations of GLP-1 (238).”

• cites [211] Gilon P Yakel J Gromada J Zhu Y Henquin JC Rorsman P G protein-dependent inhibition of L-type Ca2+ currents by acetylcholine in mouse pancreatic B-cells J Physiol 1997 499 Pt 1 65 76 9061640 PMC1159337 sourceCould not verify

  The indexed paper’s title and selected references concern acetylcholine and calcium currents in mouse pancreatic B-cells. Searches for GLP-1, 50 nM, and interburst interval found no passage in our copy reporting the claimed shortening of the interburst interval. (LLM, conf=0.72)

  Experimental — accuracy not yet measured
• cites [238] Gromada J Ding WG Barg S Renstrom E Rorsman P Multisite regulation of insulin secretion by cAMP-increasing agonists: evidence that glucagon-like peptide 1 and glucagon act via distinct receptors Pflugers Arch 1997 434 515 524 9242714 sourceNot checked — no full text of the source

The single-channel conductance of Kslow recorded with symmetrical 130 mM K+ solutions is 39 pS.

“The single-channel conductance recorded with symmetrical 130 mM K+ solutions is 39 pS (156).”

• cites [156] Dufer M Gier B Wolpers D Krippeit-Drews P Ruth P Drews G Enhanced glucose tolerance by SK4 channel inhibition in pancreatic beta-cells Diabetes 2009 58 1835 1843 19401418 PMC2712794 sourceSupported

Despite the high expression of TALK1, the TALK1-dependent current is small and limited to about 1 pA/pF at 0 mV.

“Despite the high expression, the TALK1-dependent current is small and limited to ~1 pA/pF at 0 mV.”

• cites [708] Vierra NC Dadi PK Jeong I Dickerson M Powell DR Jacobson DA Type 2 Diabetes-Associated K+ Channel TALK-1 Modulates beta-Cell Electrical Excitability, Second-Phase Insulin Secretion, and Glucose Homeostasis Diabetes 2015 64 3818 3828 26239056 PMC4613978 sourcePartly supported

  The Introduction describes KCNK16 as the most abundant K⁺-channel transcript in mouse and human β-cells, and the Results report TALK-1-dependent currents. Figure 1C quantifies current density at 0 mV, but the text of our copy does not give a value of approximately 1 pA/pF there or characterize that current as small. (LLM, conf=0.82)

  Experimental — accuracy not yet measured

With a reversal potential of -50 mV, the TALK1 current corresponds to an estimated membrane conductance of 0.02 nS/pF.

“With a reversal potential of -50 mV (708), this corresponds to an estimated membrane conductance of 0.02 nS/pF.”

• cites [708] Vierra NC Dadi PK Jeong I Dickerson M Powell DR Jacobson DA Type 2 Diabetes-Associated K+ Channel TALK-1 Modulates beta-Cell Electrical Excitability, Second-Phase Insulin Secretion, and Glucose Homeostasis Diabetes 2015 64 3818 3828 26239056 PMC4613978 sourceCould not verify

  The citation is attached to the −50 mV reversal potential. The Results report TALK-1 currents and β-cell membrane potentials, but do not state a TALK-1 reversal potential of −50 mV; the reported values near −50 mV are plateau membrane potentials, not reversal potentials. Searches for the reversal potential and the 0.02 nS/pF conductance estimate found neither in our copy. (LLM, conf=0.75)

  Experimental — accuracy not yet measured

Action potential height is also reduced in human β-cells when TASK1 is blocked, presumably due to depolarization-induced inactivation of ion channels involved in action potential firing.

“Action potential height is also reduced, presumably due to depolarization-induced inactivation of ion channels involved in action potential firing (see Section V.B.2.B.2).”

• cites [135] Dadi PK Luo B Vierra NC Jacobson DA TASK-1 Potassium Channels Limit Pancreatic alpha-Cell Calcium Influx and Glucagon Secretion Mol Endocrinol 2015 29 777 787 25849724 PMC4415209 sourcePartly supported

  The introductory passage (page unavailable) says TASK-1 inhibition depolarizes the membrane potential in pancreatic β-cells. The Discussion (page unavailable) proposes voltage-dependent channel inactivation after TASK-1 loss, but for α-cells. Our copy does not report reduced action-potential height in human β-cells when TASK-1 is blocked. (LLM, conf=0.78)

  Experimental — accuracy not yet measured

β-cells in islets from Kcnk16/TALK1-deficient mice are more depolarized than wild-type β-cells, during both the depolarized plateau from which action potentials arise and the silent interburst intervals.

“β-cells in islets from Kcnk16/TALK1-deficient mice are more depolarized than wild-type β-cells, during both the depolarized plateau from which action potentials arise, and the silent interburst intervals (708).”

• cites [708] Vierra NC Dadi PK Jeong I Dickerson M Powell DR Jacobson DA Type 2 Diabetes-Associated K+ Channel TALK-1 Modulates beta-Cell Electrical Excitability, Second-Phase Insulin Secretion, and Glucose Homeostasis Diabetes 2015 64 3818 3828 26239056 PMC4613978 sourceSupported

Kslow likely contributes to the pacemaker current.

“Thus Kslow likely contributes to the pacemaker current.”

• cites [223] Gopel SO Kanno T Barg S Eliasson L Galvanovskis J Renstrom E Rorsman P Activation of Ca(2+)-dependent K(+) channels contributes to rhythmic firing of action potentials in mouse pancreatic beta cells J Gen Physiol 1999 114 759 770 10578013 PMC2230648 sourceSupported

An increased risk of T2DM has been linked to a coding sequence polymorphism in TALK1 (rws1535500) that increases TALK1 channel activity.

“An increased risk of T2DM has been linked to a coding sequence polymorphism in TALK1 (rws1535500) (708) that increases TALK1 channel activity.”

• cites [708] Vierra NC Dadi PK Jeong I Dickerson M Powell DR Jacobson DA Type 2 Diabetes-Associated K+ Channel TALK-1 Modulates beta-Cell Electrical Excitability, Second-Phase Insulin Secretion, and Glucose Homeostasis Diabetes 2015 64 3818 3828 26239056 PMC4613978 sourceSupported

In human β-cells, ITPR3 is expressed at particularly high levels, approximately 10-fold greater than ITPR1 or ITPR2.

“In human β-cells, ITPR3 is expressed at particularly high levels (≈10-fold greater than ITPR1 or ITPR2) (68, 475).”

• cites [68] Blodgett DM Nowosielska A Afik S Pechhold S Cura AJ Kennedy NJ Kim S Kucukural A Davis RJ Kent SC Greiner DL Novel Observations From Next-Generation RNA Sequencing of Highly Purified Human Adult and Fetal Islet Cell Subsets Diabetes 2015 64 3172 3181 25931473 PMC4542439 sourceCould not verify

  The Results section on enriched β-cell transcripts lists several genes but does not mention ITPR3, ITPR1, or ITPR2. Searches for those genes and for inositol trisphosphate receptor comparisons found no passage in our copy supporting the approximately 10-fold expression difference. (LLM, conf=0.72)

  Experimental — accuracy not yet measured
• cites [475] Nica AC Ongen H Irminger JC Bosco D Berney T Antonarakis SE Halban PA Dermitzakis ET Cell-type, allelic, and genetic signatures in the human pancreatic beta cell transcriptome Genome Res 2013 23 1554 1562 23716500 PMC3759730 sourceCould not verify

  The Results describe gene expression in human β-cells, but the passages in our copy do not report expression of ITPR3 relative to ITPR1 or ITPR2. Searches for the gene names, inositol trisphosphate receptors, and a tenfold comparison found no passage addressing the claim; page numbers were not available. (LLM, conf=0.72)

  Experimental — accuracy not yet measured

GIIS is enhanced in Kcnk16/TALK1-deficient mice, especially at glucose concentrations that are just above the threshold for stimulation of insulin release.

“GIIS is enhanced, especially at glucose concentrations that are just above the threshold for stimulation of insulin release.”

• cites [708] Vierra NC Dadi PK Jeong I Dickerson M Powell DR Jacobson DA Type 2 Diabetes-Associated K+ Channel TALK-1 Modulates beta-Cell Electrical Excitability, Second-Phase Insulin Secretion, and Glucose Homeostasis Diabetes 2015 64 3818 3828 26239056 PMC4613978 sourcePartly supported

  The Results (TALK-1 Regulates β-Cell Electrical Excitability and Ca2+ Entry; page not provided) report increased insulin secretion from TALK-1-deficient islets at 7 and 14 mmol/L glucose, and increased second-phase secretion at 11 mmol/L. The Discussion calls 7 mmol/L a basal glucose level in mice, but the paper does not identify it as just above the threshold for insulin release or establish that the enhancement is especially pronounced there. (LLM, conf=0.84)

  Experimental — accuracy not yet measured

Provided KATP channel activity is low, as with >3 mM glucose, the arginine/lysine current is capable of stimulating β-cell electrical activity.

“Provided KATP channel activity is low (as is the case in the presence of >3 mM glucose), this current is capable of stimulating β-cell electrical activity (278, 499).”

• cites [278] Hermans MP Schmeer W Henquin JC The permissive effect of glucose, tolbutamide and high K+ on arginine stimulation of insulin release in isolated mouse islets Diabetologia 1987 30 659 665 3308605 sourcePartly supported

  In the Results (page not available), arginine induced β-cell electrical activity at 7 mmol/L glucose; the Discussion says it triggered spikes when the membrane was depolarised by glucose or tolbutamide. The paper does not establish an arginine/lysine current or show that any glucose concentration above 3 mmol/L suffices: it reports 7 mmol/L as the threshold condition tested for this effect. (LLM, conf=0.86)

  Experimental — accuracy not yet measured
• cites [499] Pace CS Goldsmith KT Effect of substitution of a permeable weak acid for the permissive role of glucose in amino acid-induced electrical activity in B-cells Endocrinology 1986 119 2433 2438 3536443 sourceNot checked — no full text of the source

In islets expressing non-functional KATP channels in 70% of β-cells, normal KATP channel activity in the remaining 30% of β-cells is sufficient to ensure nearly normal [Ca2+]i oscillations across the entire islet in response to glucose.

“In islets expressing non-functional KATP channels in 70% of β-cells, normal KATP channel activity in the remaining 30% of β-cells is sufficient to ensure nearly normal [Ca2+]i oscillations across the entire islet in response to glucose (554).”

• cites [554] Rocheleau JV Remedi MS Granada B Head WS Koster JC Nichols CG Piston DW Critical role of gap junction coupled KATP channel activity for regulated insulin secretion PLoS Biol 2006 4 e26 16402858 PMC1334237 sourceSupported

Electrogenic uptake of positively charged arginine or lysine evokes a small depolarizing current.

“Electrogenic uptake of positively charged arginine (or lysine) evokes a small depolarizing current (636).”

• cites [636] Smith PA Sakura H Coles B Gummerson N Proks P Ashcroft FM Electrogenic arginine transport mediates stimulus-secretion coupling in mouse pancreatic beta-cells J Physiol 1997 499 Pt 3 625 635 9130159 PMC1159281 sourcePartly supported

  The Abstract reports that L-arginine produces an inward current carried by the amino acid and links its electrogenic uptake to beta-cell depolarization. It reports a similar calcium response to L-lysine, but does not say that lysine evokes an inward current or describe the current as small. (LLM, conf=0.85)

  Experimental — accuracy not yet measured

Alanine stimulates 45Ca efflux from islets preloaded with the isotope in a Na+-dependent fashion.

“Alanine has been reported to stimulate 45Ca efflux from islets preloaded with the isotope in a Na+-dependent fashion (114).”

• cites [114] Charles S Henquin JC Distinct effects of various amino acids on 45Ca2+ fluxes in rat pancreatic islets Biochem J 1983 214 899 907 6414462 PMC1152330 sourceSupported

Coordinated [Ca2+]i oscillations are observed in human islets in the presence of the incretin hormones GLP-1 and GIP.

“However, although [Ca2+]i oscillations elicited by high glucose appear to be stochastic in human islets, coordinated oscillations are observed in the presence of the incretin hormones GLP-1 and GIP (284).”

• cites [284] Hodson DJ Mitchell RK Bellomo EA Sun G Vinet L Meda P Li D Li WH Bugliani M Marchetti P Bosco D Lipotoxicity disrupts incretin-regulated human beta cell connectivity J Clin Invest 2013 123 4182 4194 24018562 PMC4382273 sourcePartly supported

  The section “GLP-1 activates a coordinated subpopulation of β cells” reports stochastic Ca²⁺ oscillations under elevated glucose and markedly more coordinated Ca²⁺ activity after GLP-1 application in human islets. The copy identifies GIP as an incretin in the Introduction, but the passages found do not show coordinated oscillations in human islets in response to GIP. (LLM, conf=0.88)

  Experimental — accuracy not yet measured

GIIS is little affected in Tpcn1/Tpcn2 double knockout mice.

“However, GIIS is little affected in Tpcn1/Tpcn2 double knockout mice (618), which suggests two-pore channels may not play an important role in glucose-induced electrical activity, at least in mouse β-cells.”

• cites [618] Shigeto M Ramracheya R Tarasov AI Cha CY Chibalina MV Hastoy B Philippaert K Reinbothe T Rorsman N Salehi A Sones WR GLP-1 stimulates insulin secretion by PKC-dependent TRPM4 and TRPM5 activation J Clin Invest 2015 125 4714 4728 26571400 PMC4665783 sourcePartly supported

  In the section ‘GLP-1 increases [Ca²⁺]i in hyperpolarized β cells’ (page not indicated), the paper says Tpc1/Tpc2 double-knockout islets remained responsive to 20 mM glucose. That supports a retained glucose response, but the text does not establish that glucose-induced insulin secretion was ‘little affected’ relative to wild-type islets; its statement of full responsiveness concerns GLP-1. (LLM, conf=0.82)

  Experimental — accuracy not yet measured

Ryrs are only expressed at low levels in mouse β-cells.

“However, Ryrs are only expressed at low levels in mouse β-cells (56).”

• cites [56] Benner C van der Meulen T Caceres E Tigyi K Donaldson CJ Huising MO The transcriptional landscape of mouse beta cells compared to human beta cells reveals notable species differences in long non-coding RNA and protein-coding gene expression BMC Genomics 2014 15 620 25051960 PMC4124169 sourceCould not verify

  The Results sections on mouse β-cell gene expression discuss other receptors and ion channels, but do not state the expression level of Ryrs. Searches for Ryr, Ryr1–3, and ryanodine receptors found no passage addressing the claim in our copy. (LLM, conf=0.70)

  Experimental — accuracy not yet measured

Connexin-36 is distinguished by its low unitary conductance (10-15 pS).

“Connexin-36 is distinguished by its low unitary conductance (10-15 pS) (643, 680).”

• cites [643] Srinivas M Rozental R Kojima T Dermietzel R Mehler M Condorelli DF Kessler JA Spray DC Functional properties of channels formed by the neuronal gap junction protein connexin36 J Neurosci 1999 19 9848 9855 10559394 PMC6782942 sourceSupported
• cites [680] Teubner B Degen J Sohl G Guldenagel M Bukauskas FF Trexler EB Verselis VK De Zeeuw CI Lee CG Kozak CA Petrasch-Parwez E Functional expression of the murine connexin 36 gene coding for a neuron-specific gap junctional protein J Membr Biol 2000 176 249 262 10931976 PMC3659790 sourceSupported

Arginine and lysine enter the β-cell via the y+ family of cationic amino acid transporters.

“They enter the β-cell via the y+ family of cationic amino acid transporters (636).”

• cites [636] Smith PA Sakura H Coles B Gummerson N Proks P Ashcroft FM Electrogenic arginine transport mediates stimulus-secretion coupling in mouse pancreatic beta-cells J Physiol 1997 499 Pt 3 625 635 9130159 PMC1159281 sourcePartly supported

  The Abstract (page not specified in our copy) says that arginine enters mouse β-cells by electrogenic transport mediated by mCAT2A. It reports a similar calcium response to lysine, but does not establish that lysine enters β-cells through a y+ transporter. A cited reference identifies mCAT genes with y+-type transport. (LLM, conf=0.82)

  Experimental — accuracy not yet measured

Two-pore channels may not play an important role in glucose-induced electrical activity, at least in mouse β-cells.

“which suggests two-pore channels may not play an important role in glucose-induced electrical activity, at least in mouse β-cells.”

• cites [618] Shigeto M Ramracheya R Tarasov AI Cha CY Chibalina MV Hastoy B Philippaert K Reinbothe T Rorsman N Salehi A Sones WR GLP-1 stimulates insulin secretion by PKC-dependent TRPM4 and TRPM5 activation J Clin Invest 2015 125 4714 4728 26571400 PMC4665783 sourcePartly supported

  In “GLP-1 increases [Ca²⁺]i in hyperpolarized β cells” (page unavailable), the paper reports that Tpc1/Tpc2 double-knockout islets remained responsive to 20 mM glucose. Figure 10C also shows GLP-1-stimulated electrical activity in knockout β-cells. Neither passage directly establishes that glucose-induced electrical activity is unaffected by loss of the two-pore channels, so the claim is a plausible but more specific inference. (LLM, conf=0.82)

  Experimental — accuracy not yet measured

This would explain why glucose-induced [Ca2+]i responses in some β-cells (~40%) from mice expressing a gain-of-function KATP channel mutation were not synchronized across the islet.

“This would explain why although some β-cells (~40%) in islets from mice expressing a gain-of-function KATP channel mutation displayed glucose-induced [Ca2+]i responses, these were not synchronized across the islet (212).”

• cites [212] Girard CA Wunderlich FT Shimomura K Collins S Kaizik S Proks P Abdulkader F Clark A Ball V Zubcevic L Bentley L Expression of an activating mutation in the gene encoding the KATP channel subunit Kir6.2 in mouse pancreatic beta cells recapitulates neonatal diabetes J Clin Invest 2009 119 80 90 19065048 PMC2613450 sourceSupported

Zn2+ uptake into insulin granules is mediated by the ZnT8 transporter.

“Zn2+ uptake into the insulin granules is mediated by the ZnT8 (Slc30a8/SLC30A8) transporter (117).”

• cites [117] Chimienti F Devergnas S Pattou F Schuit F Garcia-Cuenca R Vandewalle B Kerr-Conte J Van Lommel L Grunwald D Favier A Seve M In vivo expression and functional characterization of the zinc transporter ZnT8 in glucose-induced insulin secretion J Cell Sci 2006 119 4199 4206 16984975 sourcePartly supported

  The Discussion says the results strongly suggest that ZnT8 enhances zinc storage in insulin granules and that its membrane topology is consistent with transport from the cytoplasm into insulin-containing vesicles. Those passages support a role for ZnT8, but present granule uptake as an inference rather than establishing the unqualified claim that ZnT8 mediates it. (LLM, conf=0.85)

  Experimental — accuracy not yet measured

vNUT is expressed in mouse β-cells.

“which is expressed in mouse β-cells(3, 146).”

• cites [3] Adriaenssens AE Svendsen B Lam BY Yeo GS Holst JJ Reimann F Gribble FM Transcriptomic profiling of pancreatic alpha, beta and delta cell populations identifies delta cells as a principal target for ghrelin in mouse islets Diabetologia 2016 59 2156 2165 27390011 PMC5016554 sourceCould not verify

  The Methods (Islet isolation and FACS; page unavailable) describe isolating mouse β-cells, and the Discussion (page unavailable) describes comparing their transcriptomes with those of other islet cells. Neither passage reports vNUT expression in β-cells; searches for vNUT, vesicular nucleotide transporter, and Slc17a9 found no passage addressing it. (LLM, conf=0.72)

  Experimental — accuracy not yet measured
• cites [146] DiGruccio MR Mawla AM Donaldson CJ Noguchi GM Vaughan J Cowing-Zitron C van der Meulen T Huising MO Comprehensive alpha, beta and delta cell transcriptomes reveal that ghrelin selectively activates delta cells and promotes somatostatin release from pancreatic islets Mol Metab 2016 5 449 458 27408771 PMC4921781 sourceCould not verify

  The Results section reports transcriptomes of mouse β-cells but does not identify vNUT as an expressed gene. Searches for vNUT, Slc17a9, and vesicular nucleotide transporter found no passage addressing its expression; page numbers are unavailable in our copy. (LLM, conf=0.72)

  Experimental — accuracy not yet measured

Nucleotide uptake into insulin secretory vesicles is mediated by vNUT (Slc17a9/SLC17A9).

“Nucleotide uptake into insulin secretory vesicles is mediated by the vesicular nucleotide transporter (vNUT, encoded by Slc17a9/SLC17A9) (596), which is expressed in mouse β-cells(3, 146).”

• cites [596] Sawada K Echigo N Juge N Miyaji T Otsuka M Omote H Yamamoto A Moriyama Y Identification of a vesicular nucleotide transporter Proc Natl Acad Sci U S A 2008 105 5683 5686 18375752 PMC2311367 sourcePartly supported

  The paper identifies SLC17A9 as VNUT and reports nucleotide uptake by the purified transporter (Identification of a vesicular nucleotide transporter; Results and Discussion, page numbers unavailable). It examines chromaffin granules and PC12-cell secretory granules, but does not establish that VNUT mediates uptake into insulin secretory vesicles. (LLM, conf=0.86)

  Experimental — accuracy not yet measured

GABA transporters GAT1-3 are only expressed at low levels in mouse and human β-cells.

“However, these are only expressed at low levels in mouse and human β-cells alike (3, 68, 146, 475).”

• cites [3] Adriaenssens AE Svendsen B Lam BY Yeo GS Holst JJ Reimann F Gribble FM Transcriptomic profiling of pancreatic alpha, beta and delta cell populations identifies delta cells as a principal target for ghrelin in mouse islets Diabetologia 2016 59 2156 2165 27390011 PMC5016554 sourceCould not verify

  The Results and Discussion passages describe low or negligible expression of the ghrelin receptor Ghsr in mouse β-cells, not GABA transporters. Searches for GABA, GAT1–3, and their gene names found no passage in our copy addressing transporter expression in mouse or human β-cells; no page numbers were provided. (LLM, conf=0.76)

  Experimental — accuracy not yet measured
• cites [68] Blodgett DM Nowosielska A Afik S Pechhold S Cura AJ Kennedy NJ Kim S Kucukural A Davis RJ Kent SC Greiner DL Novel Observations From Next-Generation RNA Sequencing of Highly Purified Human Adult and Fetal Islet Cell Subsets Diabetes 2015 64 3172 3181 25931473 PMC4542439 sourceCould not verify

  The Results section on established and enriched transcripts in purified α- and β-cells lists several β-cell genes, but does not give expression levels for GAT1–3. Searches of our copy for GABA transporters, GAT1–3, and their gene names found no passage addressing their expression in human or mouse β-cells. (LLM, conf=0.72)

  Experimental — accuracy not yet measured
• cites [146] DiGruccio MR Mawla AM Donaldson CJ Noguchi GM Vaughan J Cowing-Zitron C van der Meulen T Huising MO Comprehensive alpha, beta and delta cell transcriptomes reveal that ghrelin selectively activates delta cells and promotes somatostatin release from pancreatic islets Mol Metab 2016 5 449 458 27408771 PMC4921781 sourceCould not verify

  The Results section discusses GPCR expression in islet cells, not GABA transporter expression. Searches for GAT1–3, their gene names, and low expression in mouse and human β-cells found no passage in our copy addressing the claim. (LLM, conf=0.72)

  Experimental — accuracy not yet measured
• cites [475] Nica AC Ongen H Irminger JC Bosco D Berney T Antonarakis SE Halban PA Dermitzakis ET Cell-type, allelic, and genetic signatures in the human pancreatic beta cell transcriptome Genome Res 2013 23 1554 1562 23716500 PMC3759730 sourceCould not verify

  The Results describe transcript expression in purified human β-cells, but the passages found do not report expression levels for GAT1–3 or their gene names. Searches for GABA transporters, SLC6A1/SLC6A12/SLC6A13, and mouse and human β-cells found no passage addressing the claimed low expression in both species. (LLM, conf=0.72)

  Experimental — accuracy not yet measured

Data from patients with truncating gene variants suggest that reduced activity of ZnT8 is protective against T2DM.

“Although it was originally predicted that this variant increases T2DM risk, data from patients with truncating gene variants suggest that reduced activity of ZnT8 is actually protective (187).”

• cites [187] Flannick J Thorleifsson G Beer NL Jacobs SB Grarup N Burtt NP Mahajan A Fuchsberger C Atzmon G Benediktsson R Blangero J Loss-of-function mutations in SLC30A8 protect against type 2 diabetes Nat Genet 2014 46 357 363 24584071 PMC4051628 sourceSupported

The Cl- equilibrium potential in β-cells is around -35 mV.

“Because pancreatic β-cells have a high intracellular Cl- concentration (89), an increased Cl- conductance will depolarise the β-cells towards the Cl- equilibrium potential (ECl), which is around -35 mV (89).”

• cites [89] Braun M Ramracheya R Bengtsson M Clark A Walker JN Johnson PR Rorsman P Gamma-aminobutyric acid (GABA) is an autocrine excitatory transmitter in human pancreatic beta-cells Diabetes 2010 59 1694 1701 20413510 PMC2889769 sourceSupported

In both rodent and human β-cells, GABA is released both by a vesicular route (via exocytosis of insulin-containing granules) and a non-vesicular route.

“However, it is now clear that in both rodent and human β-cells, GABA is released both by a vesicular route (via exocytosis of the insulin-containing granules) and a non-vesicular route (89, 95).”

• cites [89] Braun M Ramracheya R Bengtsson M Clark A Walker JN Johnson PR Rorsman P Gamma-aminobutyric acid (GABA) is an autocrine excitatory transmitter in human pancreatic beta-cells Diabetes 2010 59 1694 1701 20413510 PMC2889769 sourceSupported
• cites [95] Braun M Wendt A Karanauskaite J Galvanovskis J Clark A MacDonald PE Rorsman P Corelease and differential exit via the fusion pore of GABA, serotonin, and ATP from LDCV in rat pancreatic beta cells J Gen Physiol 2007 129 221 231 17296927 PMC2151613 sourcePartly supported

  The Discussion (Concluding Remarks; page not available) says GABA can leave β-cells through vesicular and nonvesicular pathways, with vesicular release involving insulin-containing granules. The Methods (Islet Cell Preparation; page not available) identify the experimental cells as rat cells. This source supports the rodent portion of the claim but does not establish the same routes in human β-cells. (LLM, conf=0.90)

  Experimental — accuracy not yet measured

In human islets, spontaneous activity of endogenous GABAA receptor Cl- channels is observed.

“In human islets, spontaneous activity of endogenous GABAA receptor Cl- channels is observed (670), possibly reflecting the presence of an ambient GABA concentration sufficient to result in receptor activation.”

• cites [670] Taneera J Jin Z Jin Y Muhammed SJ Zhang E Lang S Salehi A Korsgren O Renstrom E Groop L Birnir B gamma-Aminobutyric acid (GABA) signalling in human pancreatic islets is altered in type 2 diabetes Diabetologia 2012 55 1985 1994 22538358 PMC3369140 sourceSupported

GABAB antagonists such as CGP55485 stimulate insulin secretion in human islets.

“GABAB antagonists such as CGP55485 stimulate insulin secretion in human islets (670).”

• cites [670] Taneera J Jin Z Jin Y Muhammed SJ Zhang E Lang S Salehi A Korsgren O Renstrom E Groop L Birnir B gamma-Aminobutyric acid (GABA) signalling in human pancreatic islets is altered in type 2 diabetes Diabetologia 2012 55 1985 1994 22538358 PMC3369140 sourcePartly supported

  The Results (Fig. 5c) report that the GABAB antagonist CGP55845 increased insulin release in human islets at 16.7 mmol/l glucose, but not at 1 mmol/l. The claim gives a different compound number, CGP55485, and omits the high-glucose condition. (LLM, conf=0.90)

  Experimental — accuracy not yet measured

GIIS is reduced in both human and mouse β-cells when purinergic receptor activation is prevented.

“Indeed, GIIS is reduced in both human and mouse β-cells when purinergic receptor activation is prevented (202, 312, 347, 737).”

• cites [202] Geisler JC Corbin KL Li Q Feranchak AP Nunemaker CS Li C Vesicular nucleotide transporter-mediated ATP release regulates insulin secretion Endocrinology 2013 154 675 684 23254199 PMC3548185 sourcePartly supported

  The section ‘VNUT overexpression enhances insulin secretion by activating P2X purinergic receptors’ reports that blocking P2X receptors significantly reduced glucose-stimulated insulin secretion in control mouse islets. The paper does not report the same receptor-blockade result in human β-cells; its Discussion mentions human islets only in connection with insulin secretion stimulated by added ATP. Page numbers are unavailable in our copy. (LLM, conf=0.86)

  Experimental — accuracy not yet measured
• cites [312] Jacques-Silva MC Correa-Medina M Cabrera O Rodriguez-Diaz R Makeeva N Fachado A Diez J Berman DM Kenyon NS Ricordi C Pileggi A ATP-gated P2X3 receptors constitute a positive autocrine signal for insulin release in the human pancreatic beta cell Proc Natl Acad Sci U S A 2010 107 6465 6470 20308565 PMC2851966 sourcePartly supported

  The Results (pp. 2–3) show that blocking certain P2 receptors reduced glucose-induced insulin secretion in human islets. The paper does not show the same effect in mouse β-cells; its Discussion (p. 5) instead emphasizes species differences and says ATP did not evoke insulin responses in mouse islets at low glucose. (LLM, conf=0.90)

  Experimental — accuracy not yet measured
• cites [347] Khan S Yan-Do R Duong E Wu X Bautista A Cheley S MacDonald PE Braun M Autocrine activation of P2Y1 receptors couples Ca (2+) influx to Ca (2+) release in human pancreatic beta cells Diabetologia 2014 57 2535 2545 25208758 sourcePartly supported

  The section on autocrine P2Y1 activation reports that the antagonist MRS-2500 reduced the glucose-induced insulin secretory response in human islets by 35%. The Discussion mentions effects of P2Y1 blockade on calcium signals in mouse β-cells, but does not report reduced glucose-induced insulin secretion in mice. (LLM, conf=0.88)

  Experimental — accuracy not yet measured
• cites [737] Wuttke A Idevall-Hagren O Tengholm A P2Y(1) receptor-dependent diacylglycerol signaling microdomains in beta cells promote insulin secretion FASEB J 2013 27 1610 1620 23299857 sourceNot checked — no full text of the source

There is a positive correlation between CHRNB2 expression and GIIS.

“There is a positive correlation between the expression of CHRNB2 expression and GIIS (199).”

• cites [199] Ganic E Singh T Luan C Fadista J Johansson JK Cyphert HA Bennet H Storm P Prost G Ahlenius H Renstrom E MafA-Controlled Nicotinic Receptor Expression Is Essential for Insulin Secretion and Is Impaired in Patients with Type 2 Diabetes Cell Rep 2016 14 1991 2002 26904947 PMC5918632 sourceSupported

A common polymorphism in SLC30A8 (rs13266634) affects T2DM risk.

“A common polymorphism in SLC30A8 (rs13266634) affects T2DM risk (384, 627).”

• cites [384] Lemaire K Ravier MA Schraenen A Creemers JW Van de Plas R Granvik M Van Lommel L Waelkens E Chimienti F Rutter GA Gilon P Insulin crystallization depends on zinc transporter ZnT8 expression, but is not required for normal glucose homeostasis in mice Proc Natl Acad Sci U S A 2009 106 14872 14877 19706465 PMC2736467 sourcePartly supported

  The introduction reports an association between a SNP marker in SLC30A8 and genetic susceptibility to type 2 diabetes. Our copy does not identify that marker as rs13266634 or state that it is common, so it supports the risk association but not those specifics. (LLM, conf=0.85)

  Experimental — accuracy not yet measured
• cites [627] Sladek R Rocheleau G Rung J Dina C Shen L Serre D Boutin P Vincent D Belisle A Hadjadj S Balkau B A genome-wide association study identifies novel risk loci for type 2 diabetes Nature 2007 445 881 885 17293876 sourceNot checked — no full text of the source

GABA induces conversion of mouse α-cells to β-cells in vivo.

“Fascinatingly, GABA induces conversion of mouse α-cells to β-cells in vivo, and treatment of transplanted human islets with GABA results (via activation of GABAA receptors in α-cells) in an increased percentage of β-cells and a reduced percentage of α-cells, suggesting a similar conversion may occur in humans (55).”

• cites [55] Ben-Othman N Vieira A Courtney M Record F Gjernes E Avolio F Hadzic B Druelle N Napolitano T Navarro-Sanz S Silvano S Long-Term GABA Administration Induces Alpha Cell-Mediated Beta-like Cell Neogenesis Cell 2016 27916274 sourceSupported

Stimulation of insulin secretion is associated with ATP release.

“Although SLC17A9 expression in human β-cells is almost undetectable (68, 475), stimulation of insulin secretion is still associated with ATP release (347).”

• cites [347] Khan S Yan-Do R Duong E Wu X Bautista A Cheley S MacDonald PE Braun M Autocrine activation of P2Y1 receptors couples Ca (2+) influx to Ca (2+) release in human pancreatic beta cells Diabetologia 2014 57 2535 2545 25208758 sourceSupported

Pancreatic β-cells have a high intracellular Cl- concentration.

“Because pancreatic β-cells have a high intracellular Cl- concentration (89), an increased Cl- conductance will depolarise the β-cells towards the Cl- equilibrium potential (ECl), which is around -35 mV (89).”

• cites [89] Braun M Ramracheya R Bengtsson M Clark A Walker JN Johnson PR Rorsman P Gamma-aminobutyric acid (GABA) is an autocrine excitatory transmitter in human pancreatic beta-cells Diabetes 2010 59 1694 1701 20413510 PMC2889769 sourceSupported

SLC17A9 expression in human β-cells is almost undetectable.

“Although SLC17A9 expression in human β-cells is almost undetectable (68, 475), stimulation of insulin secretion is still associated with ATP release (347).”

• cites [68] Blodgett DM Nowosielska A Afik S Pechhold S Cura AJ Kennedy NJ Kim S Kucukural A Davis RJ Kent SC Greiner DL Novel Observations From Next-Generation RNA Sequencing of Highly Purified Human Adult and Fetal Islet Cell Subsets Diabetes 2015 64 3172 3181 25931473 PMC4542439 sourceCould not verify

  The Results section discusses gene-expression profiles in purified human β-cells, but the passages in our copy do not report SLC17A9 expression. Searches for “SLC17A9” and for low expression of a vesicular nucleotide transporter in β-cells found no passage that establishes the claim; no page numbers were provided. (LLM, conf=0.72)

  Experimental — accuracy not yet measured
• cites [475] Nica AC Ongen H Irminger JC Bosco D Berney T Antonarakis SE Halban PA Dermitzakis ET Cell-type, allelic, and genetic signatures in the human pancreatic beta cell transcriptome Genome Res 2013 23 1554 1562 23716500 PMC3759730 sourceCould not verify

  The Results describe RNA sequencing of purified human β-cells and comparisons of gene expression, but the passages in our copy do not report SLC17A9 expression. Searches for SLC17A9 and its transporter name, VNUT, found no passage establishing that its expression is almost undetectable. (LLM, conf=0.75)

  Experimental — accuracy not yet measured

The stimulatory effect of fasiglifam can be antagonized by the TRPC3 channel-specific blocker pyrazole-3.

“The stimulatory effect of the fasiglifam could be antagonized by the TRPC3 channel-specific blocker pyrazole-3 (743).”

• cites [743] Yamada H Yoshida M Ito K Dezaki K Yada T Ishikawa SE Kakei M Potentiation of Glucose-stimulated Insulin Secretion by the GPR40-PLC-TRPC Pathway in Pancreatic beta-Cells Sci Rep 2016 6 25912 27180622 PMC4867641 sourceSupported

The GPR40 agonist fasiglifam activates a small inward current in the presence of glucose and tolbutamide (~1 pA/pF at -70 mV).

“More recently, the GPR40 agonist fasiglifam was shown to activate a small inward current in the presence of glucose and tolbutamide (~1 pA/pF at -70 mV) (743).”

• cites [743] Yamada H Yoshida M Ito K Dezaki K Yada T Ishikawa SE Kakei M Potentiation of Glucose-stimulated Insulin Secretion by the GPR40-PLC-TRPC Pathway in Pancreatic beta-Cells Sci Rep 2016 6 25912 27180622 PMC4867641 sourcePartly supported

  The Results section (‘Fasiglifam, a GPR40 agonist…’; Fig. 1a,b) reports that fasiglifam increased an inward current in beta-cells held at −70 mV with 5.6 mM glucose and tolbutamide present. The passages in our copy do not give the claimed magnitude of approximately 1 pA/pF, so that numerical detail is unverified. (LLM, conf=0.86)

  Experimental — accuracy not yet measured

The rate of oxidation of 0.7 mM oleate and palmitate is comparable to that of 3 mM glucose.

“Thus, the rate of oxidation of 0.7 mM oleate and palmitate is comparable to that of 3 mM glucose (63).”

• cites [63] Berne C The metabolism of lipids in mouse pancreatic islets. The oxidation of fatty acids and ketone bodies Biochem J 1975 152 661 666 776173 PMC1172521 sourceSupported

Glutamate may be co-released with insulin from human β-cells.

“Similarly, Vglut2 (SLC17A6) is expressed in human β-cells ((68) and glutamate may therefore be co-released with insulin from β-cells (205).”

• cites [205] Gheni G Ogura M Iwasaki M Yokoi N Minami K Nakayama Y Harada K Hastoy B Wu X Takahashi H Kimura K Glutamate acts as a key signal linking glucose metabolism to incretin/cAMP action to amplify insulin secretion Cell Rep 2014 9 661 673 25373904 PMC4536302 sourcePartly supported

  The Results section on glutamate transport into insulin granules reports that β-cell insulin granules can accumulate glutamate, and the Discussion links granular glutamate to insulin-granule exocytosis. This supports the possibility behind the claim, but the paper does not show glutamate being co-released with insulin from human β-cells; the experiments described here use cell lines and animal islets. Page numbers were not available in our copy. (LLM, conf=0.82)

  Experimental — accuracy not yet measured

IAPP is co-secreted with insulin from β-cells in response to glucose stimulation at an approximate >1:20 stoichiometry.

“IAPP is co-secreted with insulin from β-cells in response to glucose stimulation at an approximate >1:20 stoichiometry (330).”

• cites [330] Kanatsuka A Makino H Ohsawa H Tokuyama Y Yamaguchi T Yoshida S Adachi M Secretion of Islet Amyloid Polypeptide in Response to Glucose Febs Letters 1989 259 199 201 2689229 sourcePartly supported

  The discussion (section labeled “Dilution”; page unavailable) says that IAPP was co-secreted with insulin from isolated rat pancreatic islets and that secretion was regulated by glucose. It does not state an IAPP-to-insulin stoichiometry of approximately >1:20; the Figure 3 caption describes the measurements but gives no ratio. (LLM, conf=0.84)

  Experimental — accuracy not yet measured

After 25-30 action potentials, the fall in cytoplasmic/submembrane ATP/ADP-ratio results in sufficient activation of KATP channels to terminate the burst and fully repolarize the β-cell to -70 mV.

“Eventually, after 25-30 action potentials, the fall in the cytoplasmic/submembrane ATP/ADP-ratio results in sufficient activation of KATP channels to terminate the burst and fully repolarize the β-cell to -70 mV (143, 675) (Figure 12A-B).”

• cites [143] Detimary P Gilon P Henquin JC Interplay between cytoplasmic Ca2+ and the ATP/ADP ratio: a feedback control mechanism in mouse pancreatic islets Biochem J 1998 333 Pt 2 269 274 9657965 PMC1219582 sourcePartly supported

  The ‘Functional significance of the changes in the ATP/ADP ratio induced by Ca2+’ section and its Figure 4 caption (page not provided) propose that a falling ATP/ADP ratio reopens KATP channels and arrests Ca2+ influx. The caption specifies partial membrane repolarization, not full repolarization to −70 mV; our copy also does not give a count of 25–30 action potentials or establish a submembrane ATP/ADP change. (LLM, conf=0.88)

  Experimental — accuracy not yet measured
• cites [675] Tarasov AI Griffiths EJ Rutter GA Regulation of ATP production by mitochondrial Ca(2+) Cell Calcium 2012 52 28 35 22502861 PMC3396849 sourcePartly supported

  In “The pancreatic islet β-cell” section (page unavailable), the paper says that increased cytosolic ATP/ADP blocks KATP channels and triggers electrical activity; it also describes Ca2+-driven ATP consumption. It does not say that a subsequent ATP/ADP fall activates KATP channels after 25–30 action potentials, terminates a burst, or repolarizes the cell to −70 mV. (LLM, conf=0.82)

  Experimental — accuracy not yet measured

Action potential firing in this subset of β-cells may trigger electrical activity in neighboring cells that lack Na+ channels by spread of depolarization via gap junctions.

“Action potential firing in this subset of β-cells may trigger electrical activity in neighboring cells that lack Na+ channels by spread of depolarization via gap junctions (554).”

• cites [554] Rocheleau JV Remedi MS Granada B Head WS Koster JC Nichols CG Piston DW Critical role of gap junction coupled KATP channel activity for regulated insulin secretion PLoS Biol 2006 4 e26 16402858 PMC1334237 sourcePartly supported

  The Introduction (page not specified) describes synchronization of β-cell electrical activity through gap junctions. The Results/Discussion, including Figure 5 (page not specified), shows that coupling affects neighboring cells’ membrane potential and glucose-induced depolarization. The paper does not identify a firing subset defined by Na+ channels or say that action potentials from that subset trigger activity in neighbors lacking those channels. (LLM, conf=0.85)

  Experimental — accuracy not yet measured

At non-stimulatory glucose concentrations (<3 mM), KATP channel activity maintains a negative membrane potential (-70 to -60 mV) in human β-cells.

“At non-stimulatory glucose concentrations (<3 mM) (89), KATP channel activity maintains a negative membrane potential (-70 to -60 mV) (Figures 6A and 13A).”

• cites [89] Braun M Ramracheya R Bengtsson M Clark A Walker JN Johnson PR Rorsman P Gamma-aminobutyric acid (GABA) is an autocrine excitatory transmitter in human pancreatic beta-cells Diabetes 2010 59 1694 1701 20413510 PMC2889769 sourceSupported

In vivo membrane potential recordings demonstrate transition from oscillatory electrical activity to continuous action potential firing at glucose concentrations as low as 10 mM.

“Although it is questionable whether such high concentrations are attained physiologically, it is notable that in vivo membrane potential recordings also demonstrate the transition from oscillatory electrical activity to continuous action potential firing with the latter occurring at glucose concentrations as low as 10 mM (587).”

• cites [587] Sanchez-Andres JV Gomis A Valdeolmillos M The electrical activity of mouse pancreatic beta-cells recorded in vivo shows glucose-dependent oscillations J Physiol 1995 486 Pt 1 223 228 7562637 PMC1156510 sourceSupported

Both human and mouse β-cells express Slc6a17/SLC6a17, which encodes a synaptic vesicle glycine transporter.

“Both human and mouse β-cells also express Slc6a17/SLC6a17 (3, 68, 146, 475) that encodes a synaptic vesicle glycine transporter (502).”

• cites [3] Adriaenssens AE Svendsen B Lam BY Yeo GS Holst JJ Reimann F Gribble FM Transcriptomic profiling of pancreatic alpha, beta and delta cell populations identifies delta cells as a principal target for ghrelin in mouse islets Diabetologia 2016 59 2156 2165 27390011 PMC5016554 sourceCould not verify

  The Methods (Islet isolation and FACS) and Results (Transcriptomic profiling) describe sequencing mouse β-cells, but the passages in our copy do not identify Slc6a17 among the expressed genes or report its expression in human β-cells. Searches for Slc6a17, SLC6A17, and the glycine-transporter description found no passage establishing the expression claim. The separate citation (502) is attached to the transporter-function statement, so that statement is not assessed against this source. (LLM, conf=0.70)

  Experimental — accuracy not yet measured
• cites [68] Blodgett DM Nowosielska A Afik S Pechhold S Cura AJ Kennedy NJ Kim S Kucukural A Davis RJ Kent SC Greiner DL Novel Observations From Next-Generation RNA Sequencing of Highly Purified Human Adult and Fetal Islet Cell Subsets Diabetes 2015 64 3172 3181 25931473 PMC4542439 sourceCould not verify

  The Results section on enriched β-cell transcripts lists several genes but does not mention SLC6A17. Searches of our copy for SLC6A17 and glycine transporter found no passage establishing its expression in human or mouse β-cells. The transporter description is attached to a different citation in the citing sentence. (LLM, conf=0.72)

  Experimental — accuracy not yet measured
• cites [146] DiGruccio MR Mawla AM Donaldson CJ Noguchi GM Vaughan J Cowing-Zitron C van der Meulen T Huising MO Comprehensive alpha, beta and delta cell transcriptomes reveal that ghrelin selectively activates delta cells and promotes somatostatin release from pancreatic islets Mol Metab 2016 5 449 458 27408771 PMC4921781 sourceCould not verify

  The Results section, “Generation and validation of alpha, beta and delta cell transcriptomes” (page unavailable), describes mouse β-cell transcriptomes but does not report Slc6a17 expression. Searches for Slc6a17/SLC6A17 and glycine transporter found no passage establishing its expression in either mouse or human β-cells. The transporter-function clause has a separate citation in the citing sentence and is not judged here. (LLM, conf=0.72)

  Experimental — accuracy not yet measured
• cites [475] Nica AC Ongen H Irminger JC Bosco D Berney T Antonarakis SE Halban PA Dermitzakis ET Cell-type, allelic, and genetic signatures in the human pancreatic beta cell transcriptome Genome Res 2013 23 1554 1562 23716500 PMC3759730 sourceCould not verify

  The Results describe transcriptomes of human β-cells and list selected expressed genes, but the passages found for SLC6A17/Slc6a17 do not mention that gene. The paper also does not establish its expression in mouse β-cells. I searched for the gene name and the synaptic-vesicle glycine-transporter description; neither yielded a passage supporting the attributed claim. (LLM, conf=0.72)

  Experimental — accuracy not yet measured
• cites [502] Parra LA Baust T El Mestikawy S Quiroz M Hoffman B Haflett JM Yao JK Torres GE The orphan transporter Rxt1/NTT4 (SLC6A17) functions as a synaptic vesicle amino acid transporter selective for proline, glycine, leucine, and alanine Mol Pharmacol 2008 74 1521 1532 18768736 sourceNot checked — no full text of the source

KATP channel activity in human β-cells, even at low (1 mM) glucose concentrations, is only ~10% of that in mouse β-cells.

“First, KATP channel activity in human β-cells, even at low (1 mM) glucose concentrations, is only ~10% of that in mouse β-cells (535).”

• cites [535] Ramracheya R Ward C Shigeto M Walker JN Amisten S Zhang Q Johnson PR Rorsman P Braun M Membrane potential-dependent inactivation of voltage-gated ion channels in alpha-cells inhibits glucagon secretion from human islets Diabetes 2010 59 2198 2208 20547976 PMC2927942 sourceCould not verify

  The Results section on background membrane conductance reports a measurement in human α-cells at 1 mmol/l glucose, not KATP-channel activity in β-cells. Searches for human-versus-mouse β-cell KATP activity and the claimed ~10% ratio found no passage in our copy addressing that comparison. (LLM, conf=0.76)

  Experimental — accuracy not yet measured

Expression of functional GABAA receptors is very low in human α-cells.

“However, it should be noted that the expression of functional GABAA receptors is very low in human α-cells (89), an observation confirmed at the mRNA level (68, 475).”

• cites [89] Braun M Ramracheya R Bengtsson M Clark A Walker JN Johnson PR Rorsman P Gamma-aminobutyric acid (GABA) is an autocrine excitatory transmitter in human pancreatic beta-cells Diabetes 2010 59 1694 1701 20413510 PMC2889769 sourceSupported
• cites [68] Blodgett DM Nowosielska A Afik S Pechhold S Cura AJ Kennedy NJ Kim S Kucukural A Davis RJ Kent SC Greiner DL Novel Observations From Next-Generation RNA Sequencing of Highly Purified Human Adult and Fetal Islet Cell Subsets Diabetes 2015 64 3172 3181 25931473 PMC4542439 sourceCould not verify

  The citation to this paper is attached to the claimed mRNA-level confirmation. The Results section on enriched α- and β-cell transcripts discusses genes such as GCG and ARX, but the passages found by searching for GABA, GABAA receptors, and receptor subunits do not report GABAA-receptor mRNA levels in human α-cells. (LLM, conf=0.70)

  Experimental — accuracy not yet measured
• cites [475] Nica AC Ongen H Irminger JC Bosco D Berney T Antonarakis SE Halban PA Dermitzakis ET Cell-type, allelic, and genetic signatures in the human pancreatic beta cell transcriptome Genome Res 2013 23 1554 1562 23716500 PMC3759730 sourceCould not verify

  The Results describe RNA expression differences between beta cells, islets, and nonbeta cells, but do not report GABAA-receptor transcript levels specifically in human α-cells. Searches for GABA, GABAA-receptor subunits, and gamma-aminobutyric acid found no passage confirming the claimed low expression at the mRNA level. (LLM, conf=0.72)

  Experimental — accuracy not yet measured

In 30% of β-cells, voltage-gated Na+ channels also contribute to action potential firing.

“In 30% of β-cells, voltage-gated Na+ channels also contribute to action potential firing (761).”

• cites [761] Zhang Q Chibalina MV Bengtsson M Groschner LN Ramracheya R Rorsman NJ Leiss V Nassar MA Welling A Gribble FM Reimann F Na+ current properties in islet alpha- and beta-cells reflect cell-specific Scn3a and Scn9a expression J Physiol 2014 592 4677 4696 25172946 PMC4253470 sourceSupported

Vglut2 (SLC17A6) is expressed in human β-cells.

“Similarly, Vglut2 (SLC17A6) is expressed in human β-cells ((68) and glutamate may therefore be co-released with insulin from β-cells (205).”

• cites [68] Blodgett DM Nowosielska A Afik S Pechhold S Cura AJ Kennedy NJ Kim S Kucukural A Davis RJ Kent SC Greiner DL Novel Observations From Next-Generation RNA Sequencing of Highly Purified Human Adult and Fetal Islet Cell Subsets Diabetes 2015 64 3172 3181 25931473 PMC4542439 sourceCould not verify

  The Results section on enriched transcripts in purified human β-cells lists several expressed genes but does not mention SLC17A6 or VGLUT2. Searches for both names and for vesicular glutamate transporter found no passage in our copy establishing its expression in human β-cells. (LLM, conf=0.72)

  Experimental — accuracy not yet measured

Initiation of electrical activity is associated with a transient drop in the cytoplasmic ATP/ADP ratio.

“They show that initiation of electrical activity is associated with a transient drop in the cytoplasmic ATP/ADP ratio (676, 677).”

• cites [676] Tarasov AI Semplici F Li D Rizzuto R Ravier MA Gilon P Rutter GA Frequency-dependent mitochondrial Ca(2+) accumulation regulates ATP synthesis in pancreatic beta cells Pflugers Arch 2013 465 543 554 23149488 PMC3631125 sourcePartly supported

  The Results section, “Cytosolic ATP/ADP increases are controlled by electrical bursting and [Ca2+]cyt oscillation frequency” (page unavailable), reports a transient drop when bursts were imposed at 4 min−1. The claim omits that frequency condition: imposed oscillations at 1 min−1 had no discernible effect on the cytosolic ATP/ADP ratio. (LLM, conf=0.85)

  Experimental — accuracy not yet measured
• cites [677] Tarasov AI Semplici F Ravier MA Bellomo EA Pullen TJ Gilon P Sekler I Rizzuto R Rutter GA The mitochondrial Ca2+ uniporter MCU is essential for glucose-induced ATP increases in pancreatic beta-cells PLoS One 2012 7 e39722 22829870 PMC3400633 sourceSupported

There is a negative correlation between VAMP2 expression in human islets and HbA1C in vivo.

“There is a negative correlation between VAMP2 expression in human islets and HbA1C in vivo (16).”

• cites [16] Andersson SA Olsson AH Esguerra JL Heimann E Ladenvall C Edlund A Salehi A Taneera J Degerman E Groop L Ling C Reduced insulin secretion correlates with decreased expression of exocytotic genes in pancreatic islets from patients with type 2 diabetes Mol Cell Endocrinol 2012 364 36 45 22939844 sourceSupported

Syntaxin-1 modulates the activity of KATP and voltage-gated delayed rectifying K+ channels in β-cells.

“Intriguingly, syntaxin-1 also modulates the activity of the KATP and voltage-gated delayed rectifying K+ channels in β-cells (387) and its function may therefore not be limited to the regulation of exocytosis.”

• cites [387] Leung YM Kwan EP Ng B Kang Y Gaisano HY SNAREing voltage-gated K+ and ATP-sensitive K+ channels: tuning beta-cell excitability with syntaxin-1A and other exocytotic proteins Endocr Rev 2007 28 653 663 17878408 sourceSupported

There is good agreement between GIIS in vitro and in vivo in humans.

“As we have previously reported (569), there is also a good agreement between GIIS in vitro and in vivo in humans.”

• cites [569] Rorsman P Braun M Zhang Q Regulation of calcium in pancreatic alpha- and beta-cells in health and disease Cell Calcium 2012 51 300 308 22177710 PMC3334273 sourceCould not verify

  The section ‘The consensus model for glucose-induced insulin secretion’ describes glucose-triggered electrical activity and calcium responses in human β-cells, but does not compare insulin secretion in vitro with secretion in vivo. Searches for an in vitro–in vivo comparison and for agreement in human glucose-induced insulin secretion found no passage addressing the claim; page numbers were unavailable in our copy. (LLM, conf=0.73)

  Experimental — accuracy not yet measured

Islets from T2DM donors have reduced expression of STX1A (30% less) compared to islets from non-diabetic individuals.

“Islets from T2DM donors have reduced expression of STX1A (30% less) compared to islets from non-diabetic individuals (16).”

• cites [16] Andersson SA Olsson AH Esguerra JL Heimann E Ladenvall C Edlund A Salehi A Taneera J Degerman E Groop L Ling C Reduced insulin secretion correlates with decreased expression of exocytotic genes in pancreatic islets from patients with type 2 diabetes Mol Cell Endocrinol 2012 364 36 45 22939844 sourcePartly supported

  Section 3.1 reports reduced STX1A gene expression in islets from T2D donors compared with non-diabetic donors, confirmed by qRT-PCR. The passages do not give a 30% reduction, so the stated magnitude is not supported by our copy. (LLM, conf=0.85)

  Experimental — accuracy not yet measured

SNAP25 expression is reduced in islets from T2DM donors.

“SNAP25 expression is reduced in islets from T2DM donors (16, 493).”

• cites [16] Andersson SA Olsson AH Esguerra JL Heimann E Ladenvall C Edlund A Salehi A Taneera J Degerman E Groop L Ling C Reduced insulin secretion correlates with decreased expression of exocytotic genes in pancreatic islets from patients with type 2 diabetes Mol Cell Endocrinol 2012 364 36 45 22939844 sourceSupported
• cites [493] Ostenson CG Gaisano H Sheu L Tibell A Bartfai T Impaired gene and protein expression of exocytotic soluble N-ethylmaleimide attachment protein receptor complex proteins in pancreatic islets of type 2 diabetic patients Diabetes 2006 55 435 440 16443778 sourceNot checked — no full text of the source

Ablation of munc13-1 selectively suppresses 2nd phase insulin secretion.

“The role of munc13-2 in the β-cell has not been studied, but ablation of munc13-1 selectively suppresses 2nd phase insulin secretion (333).”

• cites [333] Kang L He Z Xu P Fan J Betz A Brose N Xu T Munc13-1 is required for the sustained release of insulin from pancreatic beta cells Cell Metab 2006 3 463 468 16697276 sourceSupported

Deletion of RIM leads to a decrease in the number of vesicles available for release.

“In keeping with this idea, deletion of RIM leads to a decrease in the number of vesicles available for release (252).”

• cites [252] Han Y Kaeser PS Sudhof TC Schneggenburger R RIM determines Ca(2)+ channel density and vesicle docking at the presynaptic active zone Neuron 2011 69 304 316 21262468 PMC3259453 sourceSupported

In intact mouse islets, secretory capacity varies considerably between β-cells.

“Imaging of exocytosis in intact mouse islets indicates that in fact the secretory capacity varies considerably between β-cells (402).”

• cites [402] Low JT Mitchell JM Do OH Bax J Rawlings A Zavortink M Morgan G Parton RG Gaisano HY Thorn P Glucose principally regulates insulin secretion in mouse islets by controlling the numbers of granule fusion events per cell Diabetologia 2013 56 2629 2637 23995471 PMC3825531 sourcePartly supported

  The Results (section ‘Increasing glucose concentration drives increases in the number of responding cells and granule fusion events within a cell’; page unavailable) report that more cells show fusion events as glucose rises, supporting some heterogeneity in observed responses. But the authors caution that apparently non-responding cells may have fused granules outside the imaged area, and the Discussion emphasizes coordinated responses across cells. The imaging therefore does not establish that intrinsic secretory capacity varies considerably between β-cells. (LLM, conf=0.85)

  Experimental — accuracy not yet measured

Plasma insulin oscillations are less apparent in patients with T2DM.

“These oscillations are less apparent in patients with T2DM (285, 483).”

• cites [285] Hollingdal M Juhl CB Pincus SM Sturis J Veldhuis JD Polonsky KS Porksen N Schmitz O Failure of physiological plasma glucose excursions to entrain high-frequency pulsatile insulin secretion in type 2 diabetes Diabetes 2000 49 1334 1340 10923634 sourcePartly supported

  The Serum insulin and plasma glucose concentrations section reports much smaller insulin oscillations in participants with type 2 diabetes during glucose entrainment. But during saline administration, oscillatory peaks were comparable between groups. The unqualified claim extends the finding beyond the condition in which the difference was observed. (LLM, conf=0.88)

  Experimental — accuracy not yet measured
• cites [483] O'Rahilly S Turner RC Matthews DR Impaired pulsatile secretion of insulin in relatives of patients with non-insulin-dependent diabetes N Engl J Med 1988 318 1225 1230 3283553 sourceNot checked — no full text of the source

There is evidence that attachment of a large and bulky protein to proteins involved in exocytosis may influence the kinetics of secretion.

“There is some is evidence that this may influence the kinetics of secretion (49, 445).”

• cites [49] Barg S Olofsson CS Schriever-Abeln J Wendt A Gebre-Medhin S Renstrom E Rorsman P Delay between fusion pore opening and peptide release from large dense-core vesicles in neuroendocrine cells Neuron 2002 33 287 299 11804575 sourceCould not verify

  The Discussion (page unavailable) says that labeling vesicle cargo with IAPP-EGFP did not appear to interfere with vesicle trafficking or, presumably, exocytosis. The paper reports delayed peptide release, but the passages found do not address whether attaching a bulky protein to proteins involved in exocytosis changes secretion kinetics. Searched for bulky protein tags, GFP fusions, and effects on exocytosis kinetics. (LLM, conf=0.72)

  Experimental — accuracy not yet measured
• cites [445] Michael DJ Geng X Cawley NX Loh YP Rhodes CJ Drain P Chow RH Fluorescent cargo proteins in pancreatic beta-cells: design determines secretion kinetics at exocytosis Biophys J 2004 87 L03 05 15516519 PMC1304941 sourcePartly supported

  The Abstract reports that attaching different fluorescent proteins to secretory cargo, and changing the linker between them, altered secretion kinetics in pancreatic β-cells. It does not establish that the proteins' large or bulky size caused the effect, or test attachments to exocytosis proteins generally. (LLM, conf=0.85)

  Experimental — accuracy not yet measured

Insulin secretion has been detected in real time using the phosphoinositide biosensor GFP4-Grp1 in conjunction with TIRF microscopy, to measure an increase in plasma membrane PIP3.

“Insulin secretion has also been detected in real time using the phosphoinositide biosensor GFP4-Grp1 in conjunction with TIRF microscopy, to measure an increase in the plasma membrane level of phosphatidylinositol (3,4,5) triphosphate (PIP3) (164).”

• cites [164] Dyachok O Idevall-Hagren O Sagetorp J Tian G Wuttke A Arrieumerlou C Akusjarvi G Gylfe E Tengholm A Glucose-induced cyclic AMP oscillations regulate pulsatile insulin secretion Cell Metab 2008 8 26 37 18590690 sourceSupported

Modified carbon fibers have been used to detect the release of insulin itself.

“and modified carbon fibers have been used to detect the release of insulin itself (294).”

• cites [294] Huang L Shen H Atkinson MA Kennedy RT Detection of exocytosis at individual pancreatic beta cells by amperometry at a chemically modified microelectrode Proc Natl Acad Sci U S A 1995 92 9608 9612 7568182 PMC40851 sourceSupported

Mouse β-cells express Epac2 at approximately 10-fold higher levels than their human counterparts.

“Interestingly, mouse β-cells express Epac2 at ≈10-fold higher levels than their human counterparts (56).”

• cites [56] Benner C van der Meulen T Caceres E Tigyi K Donaldson CJ Huising MO The transcriptional landscape of mouse beta cells compared to human beta cells reveals notable species differences in long non-coding RNA and protein-coding gene expression BMC Genomics 2014 15 620 25051960 PMC4124169 sourceCould not verify

  The Results section, under “Comprehensive comparison of mouse and human beta cell transcriptomes” (page unavailable), reports genes with more than 10-fold higher expression in mouse β-cells, but does not identify Epac2 among them. Searches for Epac2 and Rapgef4 found no passage stating its mouse-to-human expression ratio. (LLM, conf=0.72)

  Experimental — accuracy not yet measured

MiR-7 is the 2nd most abundant miR in human β-cells.

“MiR-7 is the 2nd most abundant miR in human β-cells.”

• cites [371] Latreille M Hausser J Stutzer I Zhang Q Hastoy B Gargani S Kerr-Conte J Pattou F Zavolan M Esguerra JL Eliasson L MicroRNA-7a regulates pancreatic beta cell function J Clin Invest 2014 124 2722 2735 24789908 PMC4038573 sourcePartly supported

  The Results report high miR-7 expression in human pancreatic islets, but do not rank it against other miRNAs in human β-cells. The statement that miR-7a2 is the most abundant member of the miR-7 family concerns variants within that family, not miR-7’s rank among all miRNAs. The claimed second-place ranking is therefore unsupported. (LLM, conf=0.85)

  Experimental — accuracy not yet measured

Carbon fiber amperometry has been successfully applied to rodent β-cells in which the granules have been preloaded with serotonin.

“This technique has been successfully applied to rodent β-cells in which the granules have preloaded with serotonin (36, 74, 634, 663) and modified carbon fibers have been used to detect the release of insulin itself (294).”

• cites [36] Aspinwall CA Brooks SA Kennedy RT Lakey JR Effects of intravesicular H+ and extracellular H+ and Zn2+ on insulin secretion in pancreatic beta cells J Biol Chem 1997 272 31308 31314 9395458 sourceSupported
• cites [74] Bokvist K Holmqvist M Gromada J Rorsman P Compound exocytosis in voltage-clamped mouse pancreatic beta-cells revealed by carbon fibre amperometry Pflugers Arch 2000 439 634 645 10764224 sourceNot checked — no full text of the source
• cites [634] Smith PA Duchen MR Ashcroft FM A fluorimetric and amperometric study of calcium and secretion in isolated mouse pancreatic beta-cells Pflugers Arch 1995 430 808 818 7478937 sourceNot checked — no full text of the source

Overexpression of miR-96 inhibits insulin secretion.

“Overexpression of miR-96 also inhibits insulin secretion.”

• cites [401] Lovis P Gattesco S Regazzi R Regulation of the expression of components of the exocytotic machinery of insulin-secreting cells by microRNAs Biol Chem 2008 389 305 312 18177263 sourcePartly supported

  The Results (p. 2) report that miR-96 overexpression diminished the secretory response of MIN6B1 cells to glucose and cAMP-raising agents. The unqualified claim is broader than that finding: basal secretion was unaffected, and the experiment measured co-released hGH as a proxy for insulin secretion. (LLM, conf=0.86)

  Experimental — accuracy not yet measured

Mir-375 is involved in adaptive β-cell expansion in response to insulin resistance.

“Mir-375 is also involved in adaptive β-cell expansion in response to insulin resistance (518).”

• cites [518] Poy MN Hausser J Trajkovski M Braun M Collins S Rorsman P Zavolan M Stoffel M miR-375 maintains normal pancreatic alpha- and beta-cell mass Proc Natl Acad Sci U S A 2009 106 5813 5818 19289822 PMC2656556 sourceSupported

Capacitance measurements report the increase in cell surface area that occurs when the secretory granule fuses with the plasma membrane.

“Measurements of membrane capacitance report the increase in cell surface area that occurs when the secretory granule fuses with the plasma membrane (334, 505) (Figure 17A).”

• cites [334] Kanno T Ma X Barg S Eliasson L Galvanovskis J Gopel S Larsson M Renstrom E Rorsman P Large dense-core vesicle exocytosis in pancreatic beta-cells monitored by capacitance measurements Methods 2004 33 302 311 15183179 sourceNot checked — no full text of the source
• cites [505] Penner R Neher E The patch-clamp technique in the study of secretion Trends Neurosci 1989 12 159 163 2470174 sourceSupported

MiR-29 is increased in β-cells exposed to elevated glucose.

“MiR-29 is increased in β-cells exposed to elevated glucose and it was proposed that this mediates the reduction of glucose-stimulated insulin secretion caused by long-term hypoglycemia via reduced expression of syntaxin-1 (186, 218).”

• cites [186] Filios SR Shalev A beta-Cell MicroRNAs: Small but Powerful Diabetes 2015 64 3631 3644 26494215 PMC4613982 sourceSupported
• cites [218] Gomes PR Graciano MF Pantaleao LC Renno AL Rodrigues SC Velloso LA Latorraca MQ Carpinelli AR Anhe GF Bordin S Long-term disruption of maternal glucose homeostasis induced by prenatal glucocorticoid treatment correlates with miR-29 upregulation Am J Physiol Endocrinol Metab 2014 306 E109 120 24253049 sourceNot checked — no full text of the source

Only 30% of docking attempts are successful.

“Only 30% of docking attempts are successful.”

• cites [196] Gandasi NR Barg S Contact-induced clustering of syntaxin and munc18 docks secretory granules at the exocytosis site Nat Commun 2014 5 3914 24835618 sourceSupported

The interaction between Ca2+ channels and secretory granules is mediated by residues 753-893 of the Cav1.2 Ca2+ channel, corresponding to the intracellular loop between the 2nd and 3rd homologous domain of the α1-subunit.

“This interaction is mediated by residues 753-893 of the Cav1.2 Ca2+ channel, which correspond to the intracellular loop between the 2nd and the 3rd homologous domain of the α1-subunit (731).”

• cites [731] Wiser O Trus M Hernandez A Renstrom E Barg S Rorsman P Atlas D The voltage sensitive Lc-type Ca2+ channel is functionally coupled to the exocytotic machinery Proc Natl Acad Sci U S A 1999 96 248 253 9874804 PMC15125 sourceSupported

Based on the length of the synprint peptide, the secretory granules must reside within 10 nm from the inner mouth of the Ca2+ channel.

“Based on the length of the synprint peptide, it has been estimated that the secretory granules must reside within 10 nm from the inner mouth of the Ca2+ channel (48) (Figure 20A).”

• cites [48] Barg S Ma X Eliasson L Galvanovskis J Gopel SO Obermuller S Platzer J Renstrom E Trus M Atlas D Striessnig J Fast exocytosis with few Ca(2+) channels in insulin-secreting mouse pancreatic B cells Biophys J 2001 81 3308 3323 11720994 PMC1301788 sourceSupported

Using a FRET probe that reports the conformational state of the t-SNARE SNAP25, the SNARE complex exists in two different conformations: a tight (high-FRET) and a loose (low-FRET) conformation.

“Using a fluorescence resonance energy transfer (FRET) probe that can report the conformational state of the t-SNARE SNAP25, it was shown that the SNARE complex exists in two different conformations: a tight (high-FRET) and a loose (low-FRET) conformation (661).”

• cites [661] Takahashi N Hatakeyama H Okado H Noguchi J Ohno M Kasai H SNARE conformational changes that prepare vesicles for exocytosis Cell Metab 2010 12 19 29 20620992 sourcePartly supported

  The Results (page unavailable) describe a SNAP25 FRET probe and distinct high- and low-FRET populations. But the authors interpret these as SNAP25 assembly with syntaxin being complete or absent, not as tight and loose conformations of an existing SNARE complex. (LLM, conf=0.87)

  Experimental — accuracy not yet measured

Ca2+ channel clustering is absent following chronic exposure to palmitate or in glucose-intolerant high-fat-fed mice.

“Interestingly, Ca2+ channel clustering is absent following chronic exposure to palmitate (a diabetogenic condition) or in glucose-intolerant high-fat-fed mice (123).”

• cites [123] Collins SC Hoppa MB Walker JN Amisten S Abdulkader F Bengtsson M Fearnside J Ramracheya R Toye AA Zhang Q Clark A Progression of diet-induced diabetes in C57BL6J mice involves functional dissociation of Ca2(+) channels from secretory vesicles Diabetes 2010 59 1192 1201 20150285 PMC2857899 sourcePartly supported

  The Discussion (page unavailable) says Ca2+ entry becomes more diffuse in high-fat-fed β-cells, suggesting less efficient channel clustering; the Results report glucose intolerance in the high-fat-fed mice. The Discussion also says that findings after more than 72 hours of palmitate exposure were similar. It does not establish that clustering is absent in either condition, so the claim is stronger than the source. (LLM, conf=0.88)

  Experimental — accuracy not yet measured

Infusion of an excess of recombinant synprint peptide into the β-cell cytosol causes selective loss of rapid exocytosis.

“The importance of the interaction between the Ca2+ channels and secretory granules for rapid exocytosis is illustrated by the selective loss of rapid exocytosis following infusion of an excess of recombinant synprint peptide into the β-cell cytosol (731) (Figure 20B).”

• cites [731] Wiser O Trus M Hernandez A Renstrom E Barg S Rorsman P Atlas D The voltage sensitive Lc-type Ca2+ channel is functionally coupled to the exocytotic machinery Proc Natl Acad Sci U S A 1999 96 248 253 9874804 PMC15125 sourcePartly supported

  The Results (page not available) show that a recombinant Lc-channel II–III loop peptide, Lc753–893, was infused into β-cells and nearly abolished depolarization-evoked exocytosis. But the paper does not show selective loss of a rapid phase: it reports inhibition of depolarization-evoked exocytosis overall, while rapid secretion elicited by photoreleased Ca²⁺ remained intact. The Discussion (page not available) uses “synprint” for an N-channel peptide discussed in previous studies, rather than explicitly naming the infused Lc peptide synprint. (LLM, conf=0.86)

  Experimental — accuracy not yet measured

Exocytosis evoked by a train of depolarizations or photorelease of caged Ca2+, two conditions that can be assumed to produce a high [Ca2+]i throughout the β-cell, was unaffected by the synprint peptide.

“Exocytosis evoked by a train of depolarizations (48) or photorelease of caged Ca2+ (731), two conditions that can be assumed to produce a high [Ca2+]I throughout the β-cell, was unaffected by the synprint peptide.”

• cites [48] Barg S Ma X Eliasson L Galvanovskis J Gopel SO Obermuller S Platzer J Renstrom E Trus M Atlas D Striessnig J Fast exocytosis with few Ca(2+) channels in insulin-secreting mouse pancreatic B cells Biophys J 2001 81 3308 3323 11720994 PMC1301788 sourcePartly supported

  In “Assembly of a functional complex between Ca²⁺ channels and secretory granules” (page unavailable), the paper reports that later pulses in a depolarization train evoked responses comparable to controls despite the synprint peptide, but the first response was much reduced. The “Solutions” section (page unavailable) specifies the peptide in solution I and caged-Ca²⁺ photorelease in solution II; the copy does not report that photorelease-evoked exocytosis was unaffected by the peptide. (LLM, conf=0.88)

  Experimental — accuracy not yet measured
• cites [731] Wiser O Trus M Hernandez A Renstrom E Barg S Rorsman P Atlas D The voltage sensitive Lc-type Ca2+ channel is functionally coupled to the exocytotic machinery Proc Natl Acad Sci U S A 1999 96 248 253 9874804 PMC15125 sourcePartly supported

  The Discussion reports that exocytosis could still be elicited by photorelease of caged Ca2+ or a train of depolarizations in the presence of Lc753–893, with a global rather than localized calcium increase. But the paper identifies the tested peptide as Lc753–893; it uses “synprint” for an N-type channel peptide discussed in connection with previous studies, so the peptide identification is not supported. (LLM, conf=0.83)

  Experimental — accuracy not yet measured

There is also agreement that 2nd phase GIIS largely reflects the release of 'newcomer' granules rather than granules that are initially docked below the plasma membrane.

“There is also agreement that 2nd phase GIIS largely reflects the release of ‘newcomer’ granules (Figure 23A) rather than granules that are initially docked below the plasma membrane (610).”

• cites [610] Seino S Shibasaki T Minami K Dynamics of insulin secretion and the clinical implications for obesity and diabetes J Clin Invest 2011 121 2118 2125 21633180 PMC3104758 sourceSupported

There is general agreement that many docked granules are released in response to membrane depolarization produced by sulfonylureas or high [K+]o.

“There is general agreement that many docked granules are released in response to membrane depolarization produced by sulfonylureas or high [K+]o (288, 446, 487, 617).”

• cites [288] Hoppa MB Collins S Ramracheya R Hodson L Amisten S Zhang Q Johnson P Ashcroft FM Rorsman P Chronic palmitate exposure inhibits insulin secretion by dissociation of Ca(2+) channels from secretory granules Cell Metab 2009 10 455 465 19945403 PMC2814048 sourcePartly supported

  The Results section (“Inhibition of Rapid Exocytosis in Palmitate-Treated Cells”; page not provided) reports that 20%–25% of docked granules were released during a train of prolonged electrical depolarizations. The Discussion says tolbutamide triggers brief action potentials, but our copy does not show that sulfonylureas or high extracellular K+ produced the reported docked-granule release. (LLM, conf=0.84)

  Experimental — accuracy not yet measured
• cites [446] Michael DJ Ritzel RA Haataja L Chow RH Pancreatic beta-cells secrete insulin in fast- and slow-release forms Diabetes 2006 55 600 607 16505221 sourceNot checked — no full text of the source
• cites [487] Ohara-Imaizumi M Fujiwara T Nakamichi Y Okamura T Akimoto Y Kawai J Matsushima S Kawakami H Watanabe T Akagawa K Nagamatsu S Imaging analysis reveals mechanistic differences between first- and second-phase insulin exocytosis J Cell Biol 2007 177 695 705 17502420 PMC2064214 sourcePartly supported

  The Introduction (page unavailable) says high KCl causes membrane depolarization and can evoke first-phase insulin release. The Results (page unavailable) report that about 75% of first-phase granule fusion under glucose stimulation came from previously docked granules. Our copy does not address sulfonylureas or directly show that high KCl releases docked granules, so it supports the general mechanism but not both specified stimuli. (LLM, conf=0.84)

  Experimental — accuracy not yet measured

Granules belonging to the reserve pool have not yet acquired release competence and must undergo a series of ATP-, Ca2+- and temperature-dependent reactions ('mobilization' or 'priming') prior to exocytosis.

“Granules belonging to the reserve pool have not yet acquired release competence and must undergo a series of ATP-, Ca2+- and temperature-dependent reactions (‘mobilization’ or ‘priming’) prior to exocytosis (571).”

• cites [571] Rorsman P Renstrom E Insulin granule dynamics in pancreatic beta cells Diabetologia 2003 46 1029 1045 12879249 sourceSupported

The high-FRET state was associated with rapid exocytosis.

“The high-FRET state was associated with rapid exocytosis (visualised by Alexa 594 uptake into the granule lumen; see Figure 18D-E).”

• cites [661] Takahashi N Hatakeyama H Okado H Noguchi J Ohno M Kasai H SNARE conformational changes that prepare vesicles for exocytosis Cell Metab 2010 12 19 29 20620992 sourceSupported

Given that granule mobility is 1.5 μm/s, an increase in the intracellular ATP/ADP ratio increases exocytosis principally by priming of granules that are already situated close to the plasma membrane (within ≈0.5 μm).

“Given that granule mobility is 1.5 μm/s (49, 702), this suggests that an increase in the intracellular ATP/ADP ratio increases exocytosis principally by priming of granules that are already situated close to the plasma membrane (within ≈0.5 μm).”

• cites [49] Barg S Olofsson CS Schriever-Abeln J Wendt A Gebre-Medhin S Renstrom E Rorsman P Delay between fusion pore opening and peptide release from large dense-core vesicles in neuroendocrine cells Neuron 2002 33 287 299 11804575 sourceSupported
• cites [702] Varadi A Ainscow EK Allan VJ Rutter GA Involvement of conventional kinesin in glucose-stimulated secretory granule movements and exocytosis in clonal pancreatic beta-cells J Cell Sci 2002 115 4177 4189 12356920 sourceNot checked — no full text of the source

Insulin secretion elicited by the combination of high-[K+]o depolarization and glucose exhibits a sustained 2nd phase insulin secretion.

“Rather than rapidly declining to basal levels, insulin secretion elicited by the combination of high-[K+]o depolarization and glucose exhibited a sustained 2nd phase insulin secretion (268).”

• cites [268] Henquin JC Ishiyama N Nenquin M Ravier MA Jonas JC Signals and pools underlying biphasic insulin secretion Diabetes 2002 51 Suppl 1 S60 67 11815460 sourceSupported

There is evidence that Ca2+ channels are physically anchored to the secretory granules.

“There is evidence that Ca2+ channels are physically anchored to the secretory granules (Figure 20).”

• cites [731] Wiser O Trus M Hernandez A Renstrom E Barg S Rorsman P Atlas D The voltage sensitive Lc-type Ca2+ channel is functionally coupled to the exocytotic machinery Proc Natl Acad Sci U S A 1999 96 248 253 9874804 PMC15125 sourceSupported

Regulation of rapid insulin granule exocytosis by [Ca2+]i close to the Ca2+ channel would be consistent with the observed low Ca2+ sensitivity of exocytosis and the finding that 1st phase (but not 2nd phase) insulin secretion is resistant to EGTA but is inhibited by BAPTA.

“Regulation of rapid insulin granule exocytosis by [Ca2+]i close to the Ca2+ channel would be consistent with the observed low Ca2+ sensitivity of exocytosis (tens of micromolar) and the finding that 1st phase (but not 2nd phase) insulin secretion is resistant to introduction of the slow Ca2+ chelator EGTA but is inhibited by the fast chelator BAPTA (510) (illustrated schematically in Figure 20C).”

• cites [510] Pertusa JA Sanchez-Andres JV Martin F Soria B Effects of calcium buffering on glucose-induced insulin release in mouse pancreatic islets: an approximation to the calcium sensor J Physiol 1999 520 Pt 2 473 483 10523416 PMC2269605 sourcePartly supported

  Split into 2 atoms — 0 confirmed, 2 partially supported, 0 disputed, 0 misattributed, 0 unverifiable. Expand the claim to see per-atom verdicts.

  Experimental — accuracy not yet measured

Secretory granules in β-cells belong to at least two functionally distinct pools: the readily releasable pool (RRP) and the reserve pool (RP).

“Secretory granules in β-cells belong to (at least) two functionally distinct pools: the readily releasable pool (RRP) and the reserve pool (RP) (472).”

• cites [472] Neher E Vesicle pools and Ca2+ microdomains: new tools for understanding their roles in neurotransmitter release Neuron 1998 20 389 399 9539117 sourcePartly supported

  In “Dynamics of Vesicle Pools” (indexed as Abstract; page unavailable), Neher describes functionally distinct vesicle pools and an immediately releasable store supplied by a larger store. The copy also discusses readily releasable secretory granules in neuroendocrine cells, but it does not establish that β-cell granules specifically belong to an RRP and a reserve pool (RP). (LLM, conf=0.84)

  Experimental — accuracy not yet measured

In β-cells, the SNAREs are unassembled in the resting state and assemble only shortly before exocytosis.

“Thus, in β-cells, the SNAREs are unassembled in the resting state and assemble only shortly before exocytosis (665).”

• cites [665] Takahashi N Sawada W Noguchi J Watanabe S Ucar H Hayashi-Takagi A Yagishita S Ohno M Tokumaru H Kasai H Two-photon fluorescence lifetime imaging of primed SNARE complexes in presynaptic terminals and beta cells Nat Commun 2015 6 8531 26439845 PMC4600761 sourceSupported

Similar values for granule density have been obtained by live-cell imaging using fluorescently tagged granules: 0.8 granule/μm2 in mouse β-cells and 0.7 granules/μm2 in human β-cells.

“Similar values for granule density have been obtained by live-cell imaging using fluorescently tagged granules: 0.8 granule/μm2 in mouse β-cells (288)and 0.7 granules/μm2 in human β-cells (447).”

• cites [288] Hoppa MB Collins S Ramracheya R Hodson L Amisten S Zhang Q Johnson P Ashcroft FM Rorsman P Chronic palmitate exposure inhibits insulin secretion by dissociation of Ca(2+) channels from secretory granules Cell Metab 2009 10 455 465 19945403 PMC2814048 sourcePartly supported

  In Results, under “Inhibition of Rapid Exocytosis in Palmitate-Treated Cells” (page unavailable), the paper reports granule density from fluorescent imaging in β-cells, but gives approximately 0.7 granules/μm², not the cited 0.8 granules/μm². The human-cell value has a separate citation in the citing sentence and is not attributed to this paper. (LLM, conf=0.88)

  Experimental — accuracy not yet measured
• cites [447] Michael DJ Xiong W Geng X Drain P Chow RH Human insulin vesicle dynamics during pulsatile secretion Diabetes 2007 56 1277 1288 17317765 sourceNot checked — no full text of the source

In a subsequent study, FRET increased - indicating assembly of the SNARE complex - 2-10 s before exocytosis.

“It was found that FRET increased - indicating assembly of the SNARE complex - 2-10 s before exocytosis.”

• cites [665] Takahashi N Sawada W Noguchi J Watanabe S Ucar H Hayashi-Takagi A Yagishita S Ohno M Tokumaru H Kasai H Two-photon fluorescence lifetime imaging of primed SNARE complexes in presynaptic terminals and beta cells Nat Commun 2015 6 8531 26439845 PMC4600761 sourceSupported

Clustering of Ca2+ channels has dramatic effects on the likelihood of a granule being exposed to elevated Ca2+ for long enough (10 ms) to evoke exocytosis: from 0.5% for individual Ca2+ channels to ~20% for a triplet of Ca2+ channels.

“Clustering of Ca2+ channels has, due to spatiotemporal [Ca2+]i domain overlap (Figure 22B-C), dramatic effects on the likelihood of a granule being exposed to elevated Ca2+ for long enough (10 ms) to evoke exocytosis: from 0.5% for individual Ca2+ channels to ~20% for a triplet of Ca2+ channels (48).”

• cites [48] Barg S Ma X Eliasson L Galvanovskis J Gopel SO Obermuller S Platzer J Renstrom E Trus M Atlas D Striessnig J Fast exocytosis with few Ca(2+) channels in insulin-secreting mouse pancreatic B cells Biophys J 2001 81 3308 3323 11720994 PMC1301788 sourceSupported

No depletion of docked granules occurred when islets were depolarized in the presence of glucose.

“Importantly, no depletion of docked granules occurred when islets were depolarized in the presence of glucose.”

• cites [268] Henquin JC Ishiyama N Nenquin M Ravier MA Jonas JC Signals and pools underlying biphasic insulin secretion Diabetes 2002 51 Suppl 1 S60 67 11815460 sourcePartly supported

  In “Does a change in the [Ca²⁺]i pattern affect the secretory pattern?” (page unavailable), the paper reports that glucose maintained insulin responses during repeated depolarization and suggests that glucose refills a limited releasable granule pool. It does not report measurements of docked granules or establish that no depletion occurred; the responses initially decreased slightly. (LLM, conf=0.82)

  Experimental — accuracy not yet measured

In INS1-cells, granule docking involves the clustering of syntaxin-1 and munc18 from a pool of freely diffusible molecules.

“In these cells, granule docking involves the clustering of the syntaxin-1 and munc18 from a pool of freely diffusible molecules.”

• cites [196] Gandasi NR Barg S Contact-induced clustering of syntaxin and munc18 docks secretory granules at the exocytosis site Nat Commun 2014 5 3914 24835618 sourceSupported

When glucose is added to islets stimulated by glibenclamide or high [K+]o after insulin secretion has almost declined to basal levels, a secondary slowly developing acceleration of insulin secretion is observed.

“Indeed, when glucose is added to islets stimulated by glibenclamide or high [K+]o after insulin secretion has almost declined to basal levels, a secondary slowly developing acceleration of insulin secretion is observed (Figures 14B and 23C).”

• cites [268] Henquin JC Ishiyama N Nenquin M Ravier MA Jonas JC Signals and pools underlying biphasic insulin secretion Diabetes 2002 51 Suppl 1 S60 67 11815460 sourcePartly supported

  The ‘Perturbations of biphasic insulin secretion’ section says that adding glucose to islets stimulated by high K+ or a sulfonylurea produces a delayed, progressive increase in insulin secretion. A separate passage in ‘Does a change in the [Ca2+]i pattern affect the secretory pattern?’ says high-K+-induced secretion returns almost to basal values. The copy does not specifically name glibenclamide or establish that glucose was added after secretion had nearly returned to basal levels. (LLM, conf=0.82)

  Experimental — accuracy not yet measured

Additional proteins required for exocytosis, including munc13 and SNAP25, are recruited to the incipient release site in INS1-cells.

“Additional proteins required for exocytosis (including munc13 and SNAP25) are then recruited to the incipient release site (196).”

• cites [196] Gandasi NR Barg S Contact-induced clustering of syntaxin and munc18 docks secretory granules at the exocytosis site Nat Commun 2014 5 3914 24835618 sourceSupported

Enhanced RRP increased 1st phase insulin secretion in response to a subsequent challenge with glucose, tolbutamide or high [K+]o.

“This enhanced 1st phase insulin secretion in response to a subsequent challenge with glucose, tolbutamide or high [K+]o (274), as expected if the size of the RRP determines 1st phase insulin secretion.”

• cites [274] Henquin JC Nenquin M Activators of PKA and Epac distinctly influence insulin secretion and cytosolic Ca2+ in female mouse islets stimulated by glucose and tolbutamide Endocrinology 2014 155 3274 3287 24977470 PMC4255079 sourcePartly supported

  The glucose- and tolbutamide-pretreatment Results sections report increased first-phase insulin secretion with Epac activation; the metabolic-amplification section also reports increased secretion during KCl stimulation. The paper does not establish that an enlarged readily releasable pool (RRP) caused these effects, and the cited KCl result is not identified as an increase specifically in first-phase secretion. Page numbers were not available in our copy. (LLM, conf=0.84)

  Experimental — accuracy not yet measured

• AssumptionRests on an assumption: The RRP amounts to 100-200 granules.

Following depletion of the RRP, refilling of the RRP follows an exponential time course with a time constant of ~30 s.

“Capacitance measurements have revealed that following depletion of the RRP - as evidenced by the cessation of exocytosis despite continued stimulation - refilling of the RRP follows an exponential time course with a time constant of ~30 s (241).”

• cites [241] Gromada J Hoy M Renstrom E Bokvist K Eliasson L Gopel S Rorsman P CaM kinase II-dependent mobilization of secretory granules underlies acetylcholine-induced stimulation of exocytosis in mouse pancreatic B-cells J Physiol 1999 518 Pt 3 745 759 10420011 PMC2269462 sourceSupported

• AssumptionRests on an assumption: The RRP amounts to 100-200 granules.

In both mouse and human β-cells, the modulation of exocytosis by glucose involves the reversible attachment of SUMO-1 to synaptotagmin-7.

“In both mouse and human β-cells there is evidence that this modulation involves the reversible attachment of small ubiquitin-like modifier protein 1 (SUMO-1) to synaptotagmin-7.”

• cites [136] Dai XQ Plummer G Casimir M Kang Y Hajmrle C Gaisano HY Manning Fox JE MacDonald PE SUMOylation regulates insulin exocytosis downstream of secretory granule docking in rodents and humans Diabetes 2011 60 838 847 21266332 PMC3046844 sourcePartly supported

  The Results section, “SUMO1 interacts dynamically with synaptotagmin VII” (page not provided), reports a SUMOylation-dependent interaction in rodent cells and human islets that is transiently disrupted by glucose. But the Discussion (page not provided) explicitly says further work is needed to determine whether SUMO1 is attached directly to synaptotagmin VII or to a binding partner. The evidence supports a reversible association, not the specified attachment. (LLM, conf=0.89)

  Experimental — accuracy not yet measured

Granule mobilization is accelerated by Ca2+ via activation of CAMKII.

“Granule mobilization is accelerated by cAMP (via activation of PKA) (545) and by Ca2+ (via activation of CAMKII) (241).”

• cites [241] Gromada J Hoy M Renstrom E Bokvist K Eliasson L Gopel S Rorsman P CaM kinase II-dependent mobilization of secretory granules underlies acetylcholine-induced stimulation of exocytosis in mouse pancreatic B-cells J Physiol 1999 518 Pt 3 745 759 10420011 PMC2269462 sourceSupported

• AssumptionRests on an assumption: The RRP amounts to 100-200 granules.

GLP-1 stimulates exocytosis via Epac2.

“For example, GLP-1 stimulates exocytosis via Epac2 (170) and by cAMP-dependent activation of PKA, which phosphorylates synaptotagmin-7 at serine-103 (735).”

• cites [170] Eliasson L Ma X Renstrom E Barg S Berggren PO Galvanovskis J Gromada J Jing X Lundquist I Salehi A Sewing S SUR1 regulates PKA-independent cAMP-induced granule priming in mouse pancreatic B-cells J Gen Physiol 2003 121 181 197 12601083 PMC2217330 sourceSupported

The final approach of granules towards the plasma membrane is influenced by the submembrane actin web.

“The final approach towards the plasma membrane is influenced by the submembrane actin web (721).”

• cites [721] Wang Z Thurmond DC Mechanisms of biphasic insulin-granule exocytosis - roles of the cytoskeleton, small GTPases and SNARE proteins J Cell Sci 2009 122 893 903 19295123 PMC2720925 sourceSupported

The amplifying effect of glucose via desumoylation of synaptotagmin-7 can occur without physical translocation of secretory granules.

“Notably, the amplifying effect of glucose (via desumoylation of synaptotagmin-7) can occur without physical translocation of secretory granules (136).”

• cites [136] Dai XQ Plummer G Casimir M Kang Y Hajmrle C Gaisano HY Manning Fox JE MacDonald PE SUMOylation regulates insulin exocytosis downstream of secretory granule docking in rodents and humans Diabetes 2011 60 838 847 21266332 PMC3046844 sourcePartly supported

  The Results (‘SUMOylation increases granule docking and SNARE complex formation’; page unavailable) say SUMO1 does not inhibit secretion by reducing granule recruitment or docking, placing its effect downstream of docking. The Discussion reports that glucose transiently disrupts the SUMO1/synaptotagmin VII interaction and that SENP1 mimics glucose’s enhancement of exocytosis, but says the underlying mechanism remains to be elucidated. Thus the paper supports an effect downstream of granule translocation, not the definite claim that glucose acts via desumoylation of synaptotagmin-7. (LLM, conf=0.84)

  Experimental — accuracy not yet measured

Glucose amplifies Ca2+-induced insulin secretion by a direct effect on the secretory machinery.

“Glucose has been shown to amplify Ca2+-induced insulin secretion by a direct effect on the secretory machinery (203, 264, 595).”

• cites [203] Gembal M Detimary P Gilon P Gao ZY Henquin JC Mechanisms by which glucose can control insulin release independently from its action on adenosine triphosphate-sensitive K+ channels in mouse B cells J Clin Invest 1993 91 871 880 8383702 PMC288039 sourcePartly supported

  The Discussion (page not provided) reports that glucose increased insulin release without a further rise in cytoplasmic Ca2+ and concludes that it modulates Ca2+’s effectiveness on the secretory machinery. It does not establish a direct effect on that machinery; the authors say the mechanism remains uncertain and may involve the beta cell’s energy state. (LLM, conf=0.84)

  Experimental — accuracy not yet measured
• cites [264] Henquin JC Triggering and amplifying pathways of regulation of insulin secretion by glucose Diabetes 2000 49 1751 1760 11078440 sourceNot checked — no full text of the source
• cites [595] Sato Y Aizawa T Komatsu M Okada N Yamada T Dual functional role of membrane depolarization/Ca2+ influx in rat pancreatic B-cell Diabetes 1992 41 438 443 1318855 sourceNot checked — no full text of the source

There is evidence that somatostatin's inhibitory effect on exocytosis is mediated by activation of the protein phosphatase calcineurin.

“There is some evidence that somatostatin’s inhibitory effect on exocytosis is mediated by activation of the protein phosphatase calcineurin (543), possibly by dephosphorylation of the same serine residue in Syt7 that is phosphorylated in the presence of GLP-1 (735).”

• cites [543] Renstrom E Ding WG Bokvist K Rorsman P Neurotransmitter-induced inhibition of exocytosis in insulin-secreting beta cells by activation of calcineurin Neuron 1996 17 513 522 8816714 sourceSupported

Glucose increases exocytosis by NADPH-dependent activation of the deSUMOylating enzyme SENP-1.

“Glucose increases exocytosis by NADPH-dependent activation of the deSUMOylating enzyme SENP-1 (136).”

• cites [136] Dai XQ Plummer G Casimir M Kang Y Hajmrle C Gaisano HY Manning Fox JE MacDonald PE SUMOylation regulates insulin exocytosis downstream of secretory granule docking in rodents and humans Diabetes 2011 60 838 847 21266332 PMC3046844 sourcePartly supported

  The Results section reports that glucose enhances β-cell exocytosis and that SENP1 knockdown prevents this enhancement. The Discussion says SENP1 mimics glucose’s effect, but explicitly states that the underlying mechanism remains to be elucidated. Our copy does not establish NADPH-dependent activation of SENP1. (LLM, conf=0.90)

  Experimental — accuracy not yet measured

Glucose-induced remodeling of the submembrane actin network is required for the full insulin secretory response.

“Glucose-induced remodeling of the submembrane actin network is required for the full insulin secretory response.”

• cites [24] Asahara S Shibutani Y Teruyama K Inoue HY Kawada Y Etoh H Matsuda T Kimura-Koyanagi M Hashimoto N Sakahara M Fujimoto W Ras-related C3 botulinum toxin substrate 1 (RAC1) regulates glucose-stimulated insulin secretion via modulation of F-actin Diabetologia 2013 56 1088 1097 23412604 PMC3622740 sourceSupported
• cites [720] Wang Z Oh E Thurmond DC Glucose-stimulated Cdc42 signaling is essential for the second phase of insulin secretion J Biol Chem 2007 282 9536 9546 17289663 PMC2396332 sourceSupported

Accumulating data point to an important role for NADPH in the amplifying effect of glucose.

“Instead, accumulating data point to an important role for NADPH (307).”

• cites [307] Ivarsson R Quintens R Dejonghe S Tsukamoto K in 't Veld P Renstrom E Schuit FC Redox control of exocytosis: regulatory role of NADPH, thioredoxin, and glutaredoxin Diabetes 2005 54 2132 2142 15983215 sourceSupported

Genetic knockdown of PAK1 and Rac1 leads to impaired 2nd phase insulin secretion.

“Genetic knockdown of PAK1 and Rac1 leads to impaired 2nd phase insulin secretion (24, 720).”

• cites [24] Asahara S Shibutani Y Teruyama K Inoue HY Kawada Y Etoh H Matsuda T Kimura-Koyanagi M Hashimoto N Sakahara M Fujimoto W Ras-related C3 botulinum toxin substrate 1 (RAC1) regulates glucose-stimulated insulin secretion via modulation of F-actin Diabetologia 2013 56 1088 1097 23412604 PMC3622740 sourcePartly supported

  The Results report reduced glucose-stimulated insulin secretion after RAC1 knockdown in INS-1 cells, and the Discussion states that only second-phase secretion was significantly reduced in beta-cell-specific Rac1-knockout mice. The paper mentions PAK1 as a RAC1 effector but does not report PAK1 knockdown or show that it impairs second-phase secretion. Page numbers are unavailable in our copy. (LLM, conf=0.88)

  Experimental — accuracy not yet measured
• cites [720] Wang Z Oh E Thurmond DC Glucose-stimulated Cdc42 signaling is essential for the second phase of insulin secretion J Biol Chem 2007 282 9536 9546 17289663 PMC2396332 sourcePartly supported

  The Results section ('Pak1 Functions in Mediating the Cdc42-dependent Activation of Rac1'; page not provided) reports that siRNA depletion of Pak1 inhibited glucose-induced insulin secretion by more than 50%. The paper links Pak1 and Rac1 to the second-phase pathway, but it does not report that Rac1 knockdown impaired secretion or that Pak1 knockdown selectively impaired the second phase; its direct selective second-phase knockdown result concerns Cdc42. (LLM, conf=0.88)

  Experimental — accuracy not yet measured

When β-cells were exposed to forskolin or to ACh, the RRP increased 3- to 4-fold relative to that seen under control conditions.

“When β-cells were exposed to forskolin (to increase cAMP and activate both PKA and Epac2) or to ACh (to mobilize intracellular Ca2+ and activate CAMKII), the RRP increased 3- to 4-fold relative that seen under control conditions (241, 545).”

• cites [241] Gromada J Hoy M Renstrom E Bokvist K Eliasson L Gopel S Rorsman P CaM kinase II-dependent mobilization of secretory granules underlies acetylcholine-induced stimulation of exocytosis in mouse pancreatic B-cells J Physiol 1999 518 Pt 3 745 759 10420011 PMC2269462 sourceSupported
• cites [545] Renstrom E Eliasson L Rorsman P Protein kinase A-dependent and -independent stimulation of exocytosis by cAMP in mouse pancreatic B-cells J Physiol 1997 502 Pt 1 105 118 9234200 PMC1159575 sourcePartly supported

  The paper’s numbered findings report that forskolin produced a sixfold increase in the maximal exocytotic response, not a three- to fourfold increase in readily releasable pool (RRP) size. Its conclusion discusses increased release probability and faster RRP refilling; the passages in our copy do not report an ACh treatment or the claimed RRP-size comparison. (LLM, conf=0.83)

  Experimental — accuracy not yet measured

• AssumptionRests on an assumption: The RRP amounts to 100-200 granules.

Inhibitory agonists like somatostatin and adrenaline act on late steps of exocytosis to reduce insulin release, for example by depriming of granules.

“Inhibitory agonists (like somatostatin and adrenaline) also act on late steps of exocytosis to reduce insulin release, for example by depriming of granules (240, 543).”

• cites [240] Gromada J Hoy M Buschard K Salehi A Rorsman P Somatostatin inhibits exocytosis in rat pancreatic alpha-cells by G(i2)-dependent activation of calcineurin and depriming of secretory granules J Physiol 2001 535 519 532 11533141 PMC2278803 sourcePartly supported

  The Discussion (page unavailable) reports that somatostatin inhibits exocytosis by depriming granules, but the experiments concern glucagon-releasing rat α-cells, not insulin-releasing cells. The Abstract (page unavailable) says adrenaline stimulates glucagon release; this paper does not establish adrenaline-induced depriming or reduced insulin release. (LLM, conf=0.89)

  Experimental — accuracy not yet measured
• cites [543] Renstrom E Ding WG Bokvist K Rorsman P Neurotransmitter-induced inhibition of exocytosis in insulin-secreting beta cells by activation of calcineurin Neuron 1996 17 513 522 8816714 sourcePartly supported

  The Abstract and Discussion report that somatostatin and adrenalin inhibit insulin secretion by acting directly on exocytosis at a step distal to the rise in cytoplasmic Ca2+. They identify calcineurin activation as the mechanism, but do not establish depriming of granules as an example of that action. (LLM, conf=0.87)

  Experimental — accuracy not yet measured

Activation of Cdc42 leads to activation of Rac1, an effect that is mediated by p21-activated kinase 1 (PAK1).

“Activation of Cdc42 leads to activation of ras-related C3 botulinum toxin substrate 1 (Rac1), an effect that is mediated by p21-activated kinase 1 (PAK1) (720).”

• cites [720] Wang Z Oh E Thurmond DC Glucose-stimulated Cdc42 signaling is essential for the second phase of insulin secretion J Biol Chem 2007 282 9536 9546 17289663 PMC2396332 sourcePartly supported

  The section “Pak1 Functions in Mediating the Cdc42-dependent Activation of Rac1” reports that Pak1 depletion abolished glucose-induced Rac1 activation in MIN6 cells, supporting a Cdc42–Pak1–Rac1 pathway. The claim omits the pancreatic β-cell and glucose-stimulation context; the paper also notes that the order of activation can differ in other cells. (LLM, conf=0.86)

  Experimental — accuracy not yet measured

Insulin granule exocytosis is targeted towards the capillaries.

“There is evidence that insulin granule exocytosis is targeted towards the capillaries and that the ‘scaffolding’ proteins RIM2 and piccolo (but not the SNARE proteins) localise to these regions (403).”

• cites [403] Low JT Zavortink M Mitchell JM Gan WJ Do OH Schwiening CJ Gaisano HY Thorn P Insulin secretion from beta cells in intact mouse islets is targeted towards the vasculature Diabetologia 2014 57 1655 1663 24795086 PMC4079948 sourceSupported

GLP-1 stimulates exocytosis by cAMP-dependent activation of PKA, which phosphorylates synaptotagmin-7 at serine-103.

“For example, GLP-1 stimulates exocytosis via Epac2 (170) and by cAMP-dependent activation of PKA, which phosphorylates synaptotagmin-7 at serine-103 (735).”

• cites [735] Wu B Wei S Petersen N Ali Y Wang X Bacaj T Rorsman P Hong W Sudhof TC Han W Synaptotagmin-7 phosphorylation mediates GLP-1-dependent potentiation of insulin secretion from beta-cells Proc Natl Acad Sci U S A 2015 112 9996 10001 26216970 PMC4538675 sourceSupported

Many potentiators of insulin secretion exert a direct effect on late steps of exocytosis in addition to stimulating β-cell electrical activity.

“Many potentiators of insulin secretion exert a direct effect on late steps of exocytosis in addition to stimulating β-cell electrical activity.”

• cites [170] Eliasson L Ma X Renstrom E Barg S Berggren PO Galvanovskis J Gromada J Jing X Lundquist I Salehi A Sewing S SUR1 regulates PKA-independent cAMP-induced granule priming in mouse pancreatic B-cells J Gen Physiol 2003 121 181 197 12601083 PMC2217330 sourceSupported
• cites [735] Wu B Wei S Petersen N Ali Y Wang X Bacaj T Rorsman P Hong W Sudhof TC Han W Synaptotagmin-7 phosphorylation mediates GLP-1-dependent potentiation of insulin secretion from beta-cells Proc Natl Acad Sci U S A 2015 112 9996 10001 26216970 PMC4538675 sourcePartly supported

  The Significance section reports that GLP-1 directly enhances Ca²⁺-triggered insulin exocytosis, supporting the claim for one potentiator. The Discussion also describes GLP-1 increasing calcium through Epac2, but it does not establish that many potentiators both act on late exocytotic steps and stimulate β-cell electrical activity. Page numbers are unavailable in our copy. (LLM, conf=0.86)

  Experimental — accuracy not yet measured

Granule mobilization is accelerated by cAMP via activation of PKA.

“Granule mobilization is accelerated by cAMP (via activation of PKA) (545) and by Ca2+ (via activation of CAMKII) (241).”

• cites [545] Renstrom E Eliasson L Rorsman P Protein kinase A-dependent and -independent stimulation of exocytosis by cAMP in mouse pancreatic B-cells J Physiol 1997 502 Pt 1 105 118 9234200 PMC1159575 sourceSupported

• AssumptionRests on an assumption: The RRP amounts to 100-200 granules.

In human β-cells, electrical activity is rapidly initiated but only slowly reversible on return to low glucose, taking as long as 30 min following exposure to 22 mM glucose.

“A distinctive feature of electrical activity in human β-cells is that, while it is rapidly initiated, it is only slowly reversible on return to low glucose - taking as long as 30 min following exposure to 22 mM glucose (Figure 6A).”

Rat β-cells generate oscillatory electrical activity very similar to that of mouse β-cells.

“It was initially reported that rat β-cells do not show oscillatory electrical activity (20) but more recent data indicate that in fact they generate oscillatory electrical activity very similar to that of mouse β-cells (424).”

• cites [424] Manning Fox JE Gyulkhandanyan AV Satin LS Wheeler MB Oscillatory membrane potential response to glucose in islet beta-cells: a comparison of islet-cell electrical activity in mouse and rat Endocrinology 2006 147 4655 4663 16857746 sourceNot checked — no full text of the source

Abcc8/ABCC8 is expressed at 5-20 fold higher levels than Kcnj11/KCNJ11.

“It is notable that Abcc8/ABCC8 is expressed at 5-20 fold higher levels than Kcnj11/KCNJ11 (see Appendix Figure 6).”

The MgATP sensitivity of the KATP channel is much lower in the whole-cell configuration, with half-maximal inhibition at 0.4-0.8 mM rather than 0.01 mM as in excised patches.

“Resolution of this puzzle is provided by the fact that the MgATP sensitivity of the channel is much lower when measured in the whole-cell configuration: inhibition is half-maximal at 0.4-0.8 mM rather than 0.01 mM as in excised patches (641).”

• cites [641] Speier S Yang SB Sroka K Rose T Rupnik M KATP-channels in beta-cells in tissue slices are directly modulated by millimolar ATP Mol Cell Endocrinol 2005 230 51 58 15664451 sourceNot checked — no full text of the source

With the proviso that more than 90% of most sulfonylureas are bound to plasma proteins in vivo, there is a good correlation between drug binding to β-cell membranes, KATP channel inhibition and the free concentration of the drug in the plasma of T2DM patients.

“It is important to remember that, in vivo, more than 90% of most sulfonylureas are bound to plasma proteins. With this proviso, there is a good correlation between drug binding to β-cell membranes, KATP channel inhibition and the free concentration of the drug in the plasma of T2DM patients (773).”

• cites [773] Zunkler BJ Lenzen S Manner K Panten U Trube G Concentration-dependent effects of tolbutamide, meglitinide, glipizide, glibenclamide and diazoxide on ATP-regulated K+ currents in pancreatic B-cells Naunyn Schmiedebergs Arch Pharmacol 1988 337 225 230 2452991 sourceNot checked — no full text of the source

Mouse and human β-cells both express high levels of CAT-1 and CAT-2.

“Mouse and human β-cells both express high levels of cationic amino acid transporters CAT-1 (Slc7a1/SLC7A1) and CAT-2 (Slc7a2/SLC7A2) (Appendix Figure 11A,D).”

• cites Appendix Figure 11A,DNot matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

Mouse β-cells express very high levels of Slc38a4 (SNAT4), a Na+-dependent transporter of neutral amino acids.

“Mouse β-cells also express very high levels of Slc38a4 (SNAT4), another Na+-dependent transporter of neutral amino acids (Appendix Figure 11C).”

• cites Appendix Figure 11CNot matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

Trpc3/TRPC3 is expressed at very low levels, if at all, in both mouse and human β-cells.

“However, Trpc3/TRPC3 is expressed at very low levels (if at all) in both mouse and human β-cells (Appendix Figure 2B).”

• cites Appendix Figure 2BNot matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

Both mouse and human β-cells express SNAP-25 at high levels.

“Both mouse and human β-cells express SNAP-25 at high levels (Snap25/SNAP25) (Appendix Figure 17A).”

When glucose is elevated to 20 mM in the continued presence of glibenclamide, insulin secretion increases to a level equivalent to steady-state 2nd phase GIIS.

“When glucose is subsequently elevated to 20 mM in the continued presence of glibenclamide, insulin secretion increases to a level equivalent to steady-state 2nd phase GIIS.”

Given a mouse pancreas of 1000 islets with on average ~80 β-cells, the release rates correspond to 0.25 and 0.62 granules/β-cell/min.

“Given that a mouse pancreas consists of 1000 islets that contain on average ~80 β-cells (see Section II.A), these release rates correspond to 0.25 and 0.62 granules/β-cell/min.”

Solving the differential equation suggests that 20,000 granules/min and 50,000 granules/min are released at 8.9 and 16.7 mM glucose, respectively.

“Solving the above equation suggests 20,000 granules/min and 50,000 granules/min are released at 8.9 and 16.7 mM glucose, respectively.”

An increase in glucose from 1 to 20 mM elicits a biphasic stimulation of insulin secretion after a delay of a few minutes.

“Figure 14A shows insulin secretion elicited by an increase in glucose from 1 to 20 mM. After a delay of a few minutes, glucose elicits a biphasic stimulation of insulin secretion.”

Of the Ca2+-dependent synaptotagmins, only Syt5 and Syt7 are expressed at significant levels in mouse and human pancreatic β-cells.

“Of the Ca2+-dependent synaptotagmins (Syt1, 2, 3, 5, 6, 7, 9 and 10) only Syt5 and 7 are expressed at significant levels in mouse and human pancreatic β-cells.”

Noc2 is highly expressed in both human and mouse β-cells.

“By contrast, Noc2 (Rph3al/RPH3AL; ‘rabphilin3a-like’) is highly expressed in both human and mouse β-cells (Appendix Figure 15F).”

The β-cell matches insulin output to glucose via metabolically induced changes in electrical activity, culminating in an increase in cytoplasmic Ca2+ and initiation of Ca2+-dependent exocytosis.

“It does so via metabolically induced changes in electrical activity, which culminate in an increase in the cytoplasmic Ca2+ concentration and initiation of Ca2+-dependent exocytosis of insulin-containing secretory granules.”

In human islets from non-diabetic donors, only a fraction of glucose entering glycolysis proceeds into the Krebs cycle, unlike mouse β-cells.

“Surprisingly, it seems that (unlike mouse β-cells) only a fraction of glucose entering glycolysis proceeds into the Krebs cycle (150).”

• cites [150] Doliba NM Qin W Najafi H Liu C Buettger CW Sotiris J Collins HW Li C Stanley CA Wilson DF Grimsby J Glucokinase activation repairs defective bioenergetics of islets of Langerhans isolated from type 2 diabetics Am J Physiol Endocrinol Metab 2012 302 E87 E102 21952036 PMC3328091 sourceNot checked — no full text of the source

Increasing glucose concentrations produce a progressive increase in burst duration at the expense of repolarized intervals until firing becomes continuous, usually above ~20 mM glucose.

“Increasing glucose concentrations produce a progressive increase in burst duration at the expense of the repolarized intervals until firing finally becomes continuous, usually above ~20 mM glucose (154, 273).”

• cites [154] Drews G Krippeit-Drews P Dufer M Electrophysiology of islet cells Adv Exp Med Biol 2010 654 115 163 20217497 sourceNot checked — no full text of the source
• cites [273] Henquin JC Meissner HP Significance of ionic fluxes and changes in membrane potential for stimulus-secretion coupling in pancreatic B-cells Experientia 1984 40 1043 1052 6386515 sourceNot checked — no full text of the source

Glucose induces a triphasic response in [Ca2+]i consisting of an initial small fall, followed by a sharp peak and then a sustained plateau with small oscillations.

“Glucose induces a triphasic response in [Ca2+]i consisting of an initial small fall in [Ca2+]i, followed by sharp peak and then a sustained plateau on which small oscillations in [Ca2+]i are superimposed.”