2-Deoxyglucose drives plasticity via an adaptive ER stress-ATF4 pathway and elicits stroke recovery and Alzheimer’s resilience

Intermittent fasting (IF) is a diet with salutary effects on cognitive aging, Alzheimer's disease (AD), and stroke.

“Intermittent fasting (IF) is a diet with salutary effects on cognitive aging, Alzheimer’s disease (AD), and stroke.”

• Also stated in: § Introduction · plain

  “Intermittent fasting (IF) is a dietary intervention widely implicated in improving brain health, cognitive aging, AD, and stroke.”

• cites [4] 4. Johnson, J.B. ∙ Summer, W. ∙ Cutler, R.G. ... Alternate day calorie restriction improves clinical findings and reduces markers of oxidative stress and inflammation in overweight adults with moderate asthma Free Radic. Biol. Med. 2007; 42:665-674 Crossref Scopus (476) PubMed Google Scholar sourcePartly supported

  The Introduction (page unavailable) says intermittent calorie restriction can protect multiple organ systems against disease in rodents, supporting a general health-benefit claim. It does not say that intermittent fasting benefits cognitive aging, Alzheimer’s disease, or stroke; the study itself examined adults with asthma. (LLM, conf=0.78)

  Experimental — accuracy not yet measured
• cites [5] 5. Courchesne-Loyer, A. ∙ Croteau, E. ∙ Castellano, C.A. ... Inverse relationship between brain glucose and ketone metabolism in adults during short-term moderate dietary ketosis: A dual tracer quantitative positron emission tomography study J. Cereb. Blood Flow Metab. 2017; 37:2485-2493 Crossref Scopus (117) PubMed Google Scholar sourceCould not verify

  The Introduction discusses a ketogenic diet and compares its ketone levels with those reached during fasting; it does not discuss intermittent fasting as an intervention. The Discussion proposes a possible metabolic benefit of ketogenic intervention in Alzheimer’s disease, while the Conclusion says effects in aging and Alzheimer’s disease remain to be assessed. Neither passage establishes benefits of intermittent fasting for cognitive aging, Alzheimer’s disease, or stroke (section pages not provided). (LLM, conf=0.76)

  Experimental — accuracy not yet measured
• cites [6] 6. Li, L. ∙ Wang, Z. ∙ Zuo, Z. Chronic intermittent fasting improves cognitive functions and brain structures in mice PLoS One. 2013; 8:e66069 Crossref Scopus (109) PubMed Google Scholar sourcePartly supported

  The Results and Discussion report improved learning and memory and reduced cerebral-cortex oxidative stress in mice after intermittent fasting; the Discussion links oxidative stress to brain aging. The paper does not show that intermittent fasting benefits Alzheimer’s disease or stroke, and its findings in mice do not establish effects on cognitive aging more broadly. No page numbers are available in our copy. (LLM, conf=0.87)

  Experimental — accuracy not yet measured

Known ATF4 target genes Trib3 and Chac1 were induced by 2-DG.

“We verified that known ATF4 target genes, Tribbles 3 (Trib3), a modulator of AKT, and Chac1, a cyclohydrolase that degrades glutathione,51”

• Also stated in: § Results · plain

  “were induced by 2-DG (Figures 5K and 5L).”

• cites [51] 51. Lange, P.S. ∙ Chavez, J.C. ∙ Pinto, J.T. ... ATF4 is an oxidative stress-inducible, prodeath transcription factor in neurons in vitro and in vivo J. Exp. Med. 2008; 205:1227-1242 Crossref Scopus (221) PubMed Google Scholar sourcePartly supported

  The Results section (page unavailable) reports that TRB3 mRNA is induced by HCA in an ATF4-dependent manner, supporting TRB3 as an ATF4 target. The passages found do not identify Chac1 as an ATF4 target or report induction of either gene by 2-DG; the source studies HCA-induced oxidative stress instead. (LLM, conf=0.86)

  Experimental — accuracy not yet measured
• cites [52] 52. Karuppagounder, S.S. ∙ Alim, I. ∙ Khim, S.J. ... Therapeutic targeting of oxygen-sensing prolyl hydroxylases abrogates ATF4-dependent neuronal death and improves outcomes after brain hemorrhage in several rodent models Sci. Transl. Med. 2016; 8:328ra29 Crossref Scopus (114) PubMed Google Scholar sourcePartly supported

  The Results section identifies Trib3 as an ATF4-regulated gene and a negative regulator of Akt. Our copy does not establish Chac1 as an ATF4 target or report induction of either gene by 2-DG; the experiments described here use other treatments, including HCA. (LLM, conf=0.88)

  Experimental — accuracy not yet measured

The dosage of 2-DG used in our study (10 mg/kg) was one-two magnitudes less than in prior studies where ketosis was induced.

“Accordingly, to achieve therapeutic benefit, the dosage of 2-DG used in our study (10 mg/kg) was one-two magnitudes less than in prior studies where ketosis was induced.3”

• cites [3] 3. Duan, W. ∙ Mattson, M.P. Dietary restriction and 2-deoxyglucose administration improve behavioral outcome and reduce degeneration of dopaminergic neurons in models of Parkinson’s disease J. Neurosci. Res. 1999; 57:195-206 Crossref Scopus (415) PubMed Google Scholar sourceNot checked — no full text of the source
• cites [25] 25. Yao, J. ∙ Chen, S. ∙ Mao, Z. ... 2-Deoxy-D-glucose treatment induces ketogenesis, sustains mitochondrial function, and reduces pathology in female mouse model of Alzheimer's disease PLoS One. 2011; 6:e21788 Crossref Scopus (137) PubMed Google Scholar sourcePartly supported

  The Results section reports that a 0.04% 2-DG diet for seven weeks increased serum ketone bodies in female 3xTgAD mice. The paper does not give that dose in mg/kg or establish that the citing study’s 10 mg/kg dose was one to two orders of magnitude lower. No page number is available in our copy. (LLM, conf=0.84)

  Experimental — accuracy not yet measured
• cites [32] 32. Yu, Z.F. ∙ Mattson, M.P. Dietary restriction and 2-deoxyglucose administration reduce focal ischemic brain damage and improve behavioral outcome: evidence for a preconditioning mechanism J. Neurosci. Res. 1999; 57:830-839 Crossref Scopus (322) PubMed Google Scholar sourceNot checked — no full text of the source

GSK2606414 significantly inhibited transcription of the ATF4 target gene Trib3 in primary neurons.

“GSK2606414 significantly inhibited 2-DG-induced eIF2α phosphorylation (Figures 6G and 6H) and transcription of the ATF4 target gene, Trib3, in primary neurons51”

• cites [51] 51. Lange, P.S. ∙ Chavez, J.C. ∙ Pinto, J.T. ... ATF4 is an oxidative stress-inducible, prodeath transcription factor in neurons in vitro and in vivo J. Exp. Med. 2008; 205:1227-1242 Crossref Scopus (221) PubMed Google Scholar sourcePartly supported

  The Results (page not provided) measure TRB3 mRNA in cortical neurons and examine an ATF4-binding site in its promoter, supporting the identification of Trib3/TRB3 as an ATF4 target gene. The paper does not report treatment with GSK2606414 or show that it inhibited Trib3 transcription; that experimental result is not supported by this citation. (LLM, conf=0.88)

  Experimental — accuracy not yet measured

2-DG induces transcription of Bdnf in the brain without ketosis.

“2-DG induced transcription of the pro-plasticity factor, Bdnf, in the brain without ketosis.”

• Also stated in: § 2 ∙ Rajiv R. Ratan · plain

  “2-deoxyglucose (2-DG), a glucose-restriction mimetic, drives Bdnf transcription”

GSK2606414 significantly inhibited 2-DG-induced eIF2α phosphorylation.

“GSK2606414 significantly inhibited 2-DG-induced eIF2α phosphorylation (Figures 6G and 6H) and transcription of the ATF4 target gene, Trib3, in primary neurons51”

ATF4 can drive Bdnf transcription.

“We found that it can also drive Bdnf transcription.”

Lowering extracellular glucose increases Bdnf message levels in a concentration-dependent manner.

“Like 2-DG, lowering extracellular glucose increased Bdnf message levels in a concentration-dependent manner (Figures S4A and S4G).”

2-DG normalized spatial long-term memory in 5xFAD mice to a level approximating WT mice treated with saline.

“2-DG normalized this capacity in 5xFAD mice to a level approximating that of WT mice treated with saline (Figures 3F–3H).”

RNA-seq data showed a significant increase as a function of 2-DG concentration in the expression of +3 kb eRNA.

“The RNA-seq data showed a significant increase as a function of 2-DG concentration in the expression of +3 kb eRNA (Figure 7B).”

ATF4 was sufficient to drive expression of ER stress-associated genes Chac1 and Trib3 in primary neurons.

“ATF4 was sufficient to drive expression of ER stress-associated genes, Chac1 and Trib3, but also Bdnf in primary neurons (Figures 6O and S3D–S3E).”

Genes in the oligosaccharide-transferase complex were significantly regulated by 2-DG.

“Cell enrichment compartment analysis also revealed genes in the oligosaccharide-transferase complex as significantly regulated by 2-DG.”

ATF4ΔRK completely blocked 2-DG-driven eRNA expression.

“ATF4ΔRK (ATF4 DNA binding site mutation) completely blocked 2-DG-driven eRNA expression (Figure 7E).”

2-DG improves functional recovery in an ischemic stroke model.

“functional recovery in an ischemic stroke model”

• Also stated in: § Results · plain

  “2-DG treatment beginning at 24 h post-stroke led to significant behavioral recovery in all tests (Figures 2B–2D).”

Reducing AMPKα1 phosphorylation did not abrogate the 2-DG-stimulated increase in Bdnf message.

“Reducing AMPKα1 phosphorylation did not abrogate the 2-DG-stimulated increase in Bdnf message (Figure 4H).”

Other genes implicated in plasticity, learning, and memory were upregulated, including Creb1, CaM kinase IID and IIG, Ryr2, Xpo4, Pten, Xbp1, and Atf4.

“Other genes implicated in plasticity, learning, and memory were upregulated, including Creb1, CaM kinase IID and IIG, ryanodine receptor 2 (Ryr2), Xpo4, Pten, Xbp1, and Atf4.”

Cellular component enrichment analysis showed significant enrichment of genes related to the ER.

“Moreover, cellular component enrichment analysis showed significant enrichment of genes related to the ER (Figures 5G and S2B)”

Conditional deletion of Ppp1r15b in the hippocampus led to constitutive phosphorylation of eIF2α.

“Conditional deletion of Ppp1r15b in the hippocampus led to a constitutive phosphorylation of eIF2α and downstream activation of ATF4 signaling as confirmed by significant upregulation of canonical ATF4 target genes Trib3, Chac1, and Ddit3 (Figures S3A–S3C).”

2-DG treatment increases Bdnf mRNA in cortical neurons in a concentration-dependent manner.

“The treatment of cortical neurons (2 days in vitro [2-DIV]) with 2-DG resulted in a concentration-dependent increase in the message levels of Bdnf (Figure 1A).”

• Also stated in: § Results · plain

  “2-DG treatment for 6 h showed a concentration-dependent increase in Bdnf message (Figure 1E).”

ActD completely blocked the 2-DG-mediated induction of LTP.

“Similarly, ActD completely blocked the 2-DG-mediated induction of LTP (Figure 5C).”

The +3 kb intron region with a single ATF4 ChIP-seq peak corresponds precisely with the +3 kb region previously identified as a region capable of regulating Bdnf gene expression as an enhancer.

“This +3 kb intron region with a single ATF4 ChIP-seq peak corresponds precisely with the +3 kb region previously identified as a region capable of regulating Bdnf gene expression as an enhancer.59”

• cites [59]Not matched

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

GSK2606414 completely blocked low extracellular glucose's ability to drive Bdnf gene expression.

“GSK2606414 completely blocked both the expression of Trib3 and Chac1 in response to low extracellular glucose and its ability to drive Bdnf gene expression (Figures S4D–S4F and S4J–S4L).”

2-DG did not induce an increase in ATF4 binding at the promoter region upstream of the TSS.

“2-DG did not induce an increase in ATF4 binding at the promoter region upstream of the TSS (transcription start site).”

Tunicamycin increased expression of the ER stress chaperone Bip.

“Tunicamycin induced an expected increase in the phosphorylation of eIF2α (Figures 6C and 6D) and an increased expression of the ER stress chaperone Bip (Figure 6E).”

2-DG does not increase BHB levels in cultured neurons.

“2-DG did not increase BHB levels (Figure S1A)”

2-DG-induced changes in Bdnf were not mediated via homeostatic activation of AMPK in response to decreases in ATP.

“Additionally, 2-DG-induced changes in Bdnf were not mediated via homeostatic activation of AMPK in response to decreases in ATP (Figure 4H) but rather via adaptive transcription in response to diminished N-glycosylation and ER stress (Figures 5 and 6).”

2-DG treatment enhanced binding of ATF4 to the regulatory regions of Jdp2, Creb5, Cdc42bpa, Xbp1, Ppp3ca, Cebpγ, Atf3, and Jun.

“Specifically, 2-DG treatment enhanced binding of ATF4 to the regulatory regions of Jdp2, Creb5, Cdc42bpa, Xbp1, Ppp3ca, Cebpγ, Atf3, and Jun.”

Improvement in sensorimotor behaviors was observed 1-2 weeks following 2-DG administration.

“In all cases, improvement in sensorimotor behaviors was observed 1–2 weeks following the administration of 2-DG”

Increasing concentrations of glucose or mannose decreased 2-DG-induced Bdnf message in a concentration-dependent manner.

“Indeed, increasing concentrations of glucose or mannose decreased 2-DG-induced Bdnf message in a concentration-dependent manner, consistent with the hypothesis that 2-DG-diminished N-linked glycosylation drives the 2-DG-induced Bdnf message (Figures 6A and 6B).”

Activation of eIF2α phosphorylation led to a significant increase in LTP.

“Activation of eIF2α phosphorylation led to a significant increase in Bdnf gene expression in the hippocampus (Figure 6L) with a significant increase in LTP (Figure 6M).”

ATF4 was sufficient to drive Bdnf expression in primary neurons.

“ATF4 was sufficient to drive expression of ER stress-associated genes, Chac1 and Trib3, but also Bdnf in primary neurons (Figures 6O and S3D–S3E).”

2-DG treatment beginning 24 h post-stroke leads to significant behavioral recovery in all tests.

“2-DG treatment beginning at 24 h post-stroke led to significant behavioral recovery in all tests (Figures 2B–2D).”

Primary neurons treated with increasing concentrations of 2-DG (1-15 mM) exhibited a weak correlation between AMPKα1 activation and Bdnf expression.

“Primary neurons treated with increasing concentrations of 2-DG (1–15 mM) exhibited a weak correlation between the level of AMPKα1 activation and Bdnf expression (Figures 4B–4E).”

ATF4 inhibition blocked 2-DG's ability to drive Bdnf message levels.

“ATF4 inhibition blocked 2-DG’s ability to drive Bdnf message levels (Figure 6N).”

2-DG increased ATF4 reporter translation in the brain, as monitored by in vivo bioluminescence imaging.

“We found an increase in ATF4 reporter translation in the brain, as monitored by in vivo bioluminescence imaging (Figures 7J and 7K).”

Activation of eIF2α phosphorylation led to a significant increase in Bdnf gene expression in the hippocampus.

“Activation of eIF2α phosphorylation led to a significant increase in Bdnf gene expression in the hippocampus (Figure 6L) with a significant increase in LTP (Figure 6M).”

The dose of 2-DG (10 mg/kg) that induced Bdnf in the brain was two orders of magnitude less than that used in prior studies.

“The dose of 2-DG (10 mg/kg) that induced Bdnf in the brain was two orders of magnitude less than that used in prior studies.”

Tunicamycin induced an increase in phosphorylation of eIF2α.

“Tunicamycin induced an expected increase in the phosphorylation of eIF2α (Figures 6C and 6D) and an increased expression of the ER stress chaperone Bip (Figure 6E).”

These genes were also upregulated in RNA-seq analysis.

“Of note, these genes were also upregulated in our RNA-seq analysis, supporting the notion that ATF4 drives the upregulation of a cassette of genes associated with plasticity in response to 2-DG (Figure 7I).”

4 weeks of 2-DG infusion normalized LTP in 5xFAD mice to a level like WT mice infused with saline.

“Similarly, 4 weeks of 2-DG infusion normalized LTP to a level like that of WT mice infused with saline (Figure 3C).”

GSK2606414 completely blocked expression of Trib3 and Chac1 in response to low extracellular glucose.

“GSK2606414 completely blocked both the expression of Trib3 and Chac1 in response to low extracellular glucose and its ability to drive Bdnf gene expression (Figures S4D–S4F and S4J–S4L).”

2-DG treatment for 24 h significantly increases mature BDNF protein.

“Further, 2-DG treatment for 24 h significantly increased mature BDNF protein (Figures 1B and 1C).”

Reducing glucose in the media drove expression of Trib3 and Chac1.

“Reducing glucose in the media also drove the expression of Trib3 and Chac1 (Figures S4B, S4C, S4H, and S4I).”

Increases in ATF4 reporter activity were confirmed in cortical lysates from these mice.

“Since IVIS imaging is a two-dimensional image of three-dimensional tissue, we confirmed that increases in ATF4 reporter activity in cortical lysates from these mice (Figure S7).”

LTP is significantly increased in response to acute 2-DG treatment for 6 h in hippocampal slices.

“LTP was significantly increased in response to acute treatment with 2-DG for 6 h (Figure 1J).”

GSEA showed maximal upregulation of gene signatures related to ER stress in response to 2-DG treatment.

“Gene set enrichment analysis (GSEA) showed maximal upregulation of the gene signatures related to ER stress in response to 2-DG treatment (Figures 5F and S2A).”

The ATF4 binding enhancer is evolutionarily conserved among mammals.

“This ATF4 binding enhancer is evolutionarily conserved among mammals, further suggesting its importance in Bdnf gene regulation (Figure S5).”

Tunicamycin led to a significant increase in expression of Bdnf.

“The same concentration of tunicamycin led to a significant increase in the expression of Bdnf (Figure 6F), indicating that inhibition of N-linked glycosylation appears necessary and sufficient to induce Bdnf message.”

GSK2606414 blocked 2-DG's ability to induce expression of Bdnf.

“In parallel, the same concentration of GSK2606414 blocked 2-DG’s ability to induce expression of Bdnf (Figure 6K).”

The ATF4 motif was identified as the most enriched motif that overlaps within the ChIP-seq peak of the +3 kb intron region.

“The ATF4 motif was identified as the most enriched motif that overlaps within the ChIP-seq peak of the +3 kb intron region (Figure 7H).”

2-DG enhances memory in an AD model (5xFAD).

“2-DG enhanced memory in an AD model (5xFAD)”

• Also stated in: § Results · plain

  “2-DG normalized this capacity in 5xFAD mice to a level approximating that of WT mice treated with saline (Figures 3F–3H).”

2-DG has significant therapeutic effects on learning and memory deficits in an AD mouse model.

“We were able to demonstrate (1) significant therapeutic effects on learning and memory deficits in an AD mouse model and (2) enhanced sensorimotor recovery following ischemic stroke (Figures 2 and 3).”

Ingenuity pathway analysis showed a significant upregulation of a broad cassette of ATF4 target genes in response to 2-DG treatment.

“Ingenuity pathway analysis for upstream transcriptional regulators showed a significant upregulation of a broad cassette of ATF4 target genes in response to 2-DG treatment (Fisher’s exact test, p < 0.05, Figure S2C).”

Conditional deletion of Ppp1r15b in the hippocampus led to significant upregulation of canonical ATF4 target genes Trib3, Chac1, and Ddit3.

“Conditional deletion of Ppp1r15b in the hippocampus led to a constitutive phosphorylation of eIF2α and downstream activation of ATF4 signaling as confirmed by significant upregulation of canonical ATF4 target genes Trib3, Chac1, and Ddit3 (Figures S3A–S3C).”

2-DG (1 mg/kg/day) resulted in a trend toward normalization of short-term memory in 5xFAD mice.

“2-DG (1 mg/kg/day) resulted in a trend toward the normalization of short-term memory in 5xFAD mice (Figures 3D and 3E).”

Inhibiting transcription with ActD completely blocked the ability of 2-DG to increase Bdnf message in cortical cultures and hippocampal slices.

“Inhibiting transcription with ActD completely blocked the ability of 2-DG to increase Bdnf message in cortical cultures and hippocampal slices (Figures 5A and 5B).”

2-DG led to increases in reporter expression in primary neurons expressing a Bdnf promoter I firefly luciferase construct containing a +3 kb forward or reverse enhancer.

“2-DG led to increases in reporter expression in primary neurons expressing a Bdnf promoter I firefly luciferase construct containing a +3 kb forward or reverse enhancer.”

2-DG treatment enhances Bdnf gene expression in synaptically active human neurons.

“Thus, like synaptically inactive mouse neurons, 2-DG treatment enhances Bdnf gene expression in synaptically active human neurons.”

All genes related to ER stress showed a concentration-dependent increase in response to 1 and 10 mM concentrations of 2-DG.

“All genes related to ER stress showed a concentration-dependent increase in response to 1 and 10 mM concentrations of 2-DG (Figure 5H).”

The ATF4 binding site mutant completely blocked the 2-DG-driven induction of Trib3 promoter activity.

“As predicted, the ATF4 binding site mutant completely blocked the 2-DG-driven induction of Trib3 promoter activity (Figure S3F).”

Mutation of the ATF4 binding site in the +3 kb enhancer region blocked 2-DG-induced increases in reporter expression.

“Mutation of the ATF4 binding site in the +3 kb enhancer region (corresponding to the site of enrichment of ATF4 binding by ChIP) blocked 2-DG-induced increases in reporter expression (Figure 7G).”

De novo motif analysis returned enriched sequences of TFs such as CHOP, CEBP:AP1, CEBPα, DDIT3:CEBPα, CEBPβ, JUN, and JUN(Var.2) within the peak region.

“De novo motif analysis returned enriched sequences of TFs such as CHOP, CEBP:AP1, CEBPα, DDIT3:CEBPα, CEBPβ, JUN, and JUN(Var.2) within the peak region (Figure S6).”

2-DG drove Bdnf gene expression by increasing its transcription.

“2-DG drove Bdnf gene expression by increasing its transcription (Figure 5).”

No difference in infarct size between vehicle and 2-DG-treated mice 3 days after stroke onset.

“Consistently, we found no difference in infarct size between vehicle and 2-DG-treated mice 3 days after stroke onset (Figures 2E and 2F).”

Release of 2-DG through Alzet mini-osmotic pump for 4 weeks did not increase BHB levels in the brain.

“The release of 2-DG through the Alzet mini-osmotic pump for 4 weeks did not increase BHB levels in the brain (Figures S1F and S1G)”

2-DG-driven eRNA expression was reduced in hippocampal slices with conditional deletion of neuronal ATF4.

“2-DG-driven eRNA expression was also reduced in hippocampal slices with conditional deletion of neuronal ATF4, indicating that neuronal ATF4 drives 2-DG-induced eRNA expression (Figures S3G and S3H).”

Quantitative real-time PCR indicated a significant increase in antisense eRNA expression in response to 2-DG treatment in primary neurons.

“A similar result was obtained with quantitative real-time PCR, which indicated a significant increase in antisense eRNA expression in response to 2-DG treatment in primary neurons (Figure 7C).”

2-DG enhances sensorimotor recovery following ischemic stroke.

“We were able to demonstrate (1) significant therapeutic effects on learning and memory deficits in an AD mouse model and (2) enhanced sensorimotor recovery following ischemic stroke (Figures 2 and 3).”

2-DG can drive transcription of Bdnf and other genes associated with plasticity or regeneration independent of ketones.

“We found 2-DG could drive the transcription of Bdnf and other genes associated with plasticity or regeneration (e.g., Atf3, Cdc42bpa, Ppp3cc, Cebpγ) independent of ketones (Figures 1A, 3B, and 5E).”

Bdnf gene expression is significantly higher in brain cortices of Glut3+/− mice than in Glut3+/+ mice.

“Bdnf gene expression in brain cortices of (Glut3+/−) mice was significantly higher than in Glut3+/+ mice”

2-DG enhances the paradoxical translation of ATF4 in the brain.

“We used in vivo bioluminescence imaging of an ATF4 5′ UTR-luciferase construct to confirm that 2-DG enhances the paradoxical translation of ATF4 in the brain (Figures 7J and 7K).68”

• cites [68]Not matched

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

2-DG did not increase levels of urinary ketones.

“It also did not increase levels of urinary ketones”

2-DG showed no nootropic increase in WT mice in either Y-maze or Morris water maze task.

“2-DG showed no nootropic increase in WT mice in either the Y-maze or Morris water maze task (Figures 3D–3H).”

2-DG (10 mM) increased the AMP-ATP ratio in cortical neurons.

“As predicted, 2-DG (10 mM) increased the AMP-ATP ratio, as measured by laser capture mass spectrometry (Figure 4A) in cortical neurons.”

ATF4 was selectively enriched near a 3 kb intron region downstream of the Bdnf exon I TSS.

“Instead, ATF4 was selectively enriched (indicated by a single ChIP-seq peak in the University of California Santa Cruz (UCSC) genome browser in the 2-DG treated i.p. fraction) near a 3 kb intron region downstream of the Bdnf exon I TSS (Figure 7A).”

Bdnf mRNA levels significantly increased in hippocampal slices from 5xFAD mice infused with 2-DG compared with saline alone.

“Bdnf mRNA levels significantly increased in hippocampal slices from 5xFAD mice infused with 2-DG compared with saline alone (Figure 3B).”

Changes in Bdnf expression were found at much lower 2-DG doses than previously evaluated.

“By contrast, we found changes in Bdnf expression at much lower 2-DG doses than previously evaluated.64”

• cites [64]Not matched

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

Doses of 2-DG extrapolated from our animal studies are approximately 60 times lower than those used in cancer patients.

“doses of 2-DG extrapolated from our animal studies are approximately 60 times lower than those used in cancer patients.”

2-deoxyglucose (2-DG) is a glucose analog that can be used to mimic glucose restriction.

“2-deoxyglucose (2-DG), a glucose analog, can be used to mimic glucose restriction.”

• Also stated in: § Introduction · plain

  “which inhibits the utilization of glucose by blocking its transport and metabolism.”

• cites [20] 20. Tower, D.B. The effects of 2-deoxy-D-glucose on metabolism of slices of cerebral cortex incubated in vitro J. Neurochem. 1958; 3:185-205 Crossref Scopus (108) PubMed Google Scholar sourceNot checked — no full text of the source
• cites [21] 21. Bachelard, H.S. Specificity and kinetic properties of monosaccharide uptake into guinea pig cerebral cortex in vitro J. Neurochem. 1971; 18:213-222 Crossref Scopus (103) PubMed Google Scholar sourceNot checked — no full text of the source
• cites [22] 22. Horton, R.W. ∙ Meldrum, B.S. ∙ Bachelard, H.S. Enzymic and cerebral metabolic effects of 2-deoxy-D-glucose J. Neurochem. 1973; 21:507-520 Crossref Scopus (144) PubMed Google Scholar sourceNot checked — no full text of the source

ATF4 drives the upregulation of a cassette of genes associated with plasticity in response to 2-DG.

“Of note, these genes were also upregulated in our RNA-seq analysis, supporting the notion that ATF4 drives the upregulation of a cassette of genes associated with plasticity in response to 2-DG (Figure 7I).”

This concentration of ActD inhibits global transcription in cortical neurons.

“This concentration of ActD inhibits global transcription in cortical neurons.”

• cites [48] 48. Ratan, R.R. ∙ Murphy, T.H. ∙ Baraban, J.M. Macromolecular synthesis inhibitors prevent oxidative stress-induced apoptosis in embryonic cortical neurons by shunting cysteine from protein synthesis to glutathione J. Neurosci. 1994; 14:4385-4392 Crossref PubMed Google ScholarNot matched

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

ER stress induced by 2-DG can, in the absence of ketosis, lead to transcription of genes involved in plasticity and cognitive resilience as well as proteostasis.

“They further demonstrate that ER stress induced by 2-DG can, in the absence of ketosis, lead to the transcription of genes involved in plasticity and cognitive resilience as well as proteostasis.”

2-DG reduces N-linked glycosylation.

“Our results suggest that 2-DG reduces N-linked glycosylation, induces sublethal ER stress, and drives Bdnf expression and LTP via hippocampal ATF4 in vivo (Figure 6R).”

2-DG does not increase cell death after 24 h exposure.

“Live-dead staining showed no increased cell death after 24 h of 2-DG exposure (Figure 1D).”

Effects of low glucose are independent of ketone bodies.

“suggesting that the effects of low glucose are independent of ketone bodies.”

The ED50 of drugs injected through i.c.v. route is usually around 10-fold less than ED50 of drugs injected through i.p. route.

“Since the ED50 (median effective dose value) of drugs injected through an intracerebroventricular (i.c.v.) route is usually around 10-fold less than the ED50 of drugs injected through an i.p. route to achieve a similar effect on the brain”

• cites [44] 44. Serralta, A. ∙ Barcia, J.A. ∙ Ortiz, P. ... Effect of intracerebroventricular continuous infusion of valproic acid versus single i.p. and i.c.v. injections in the amygdala kindling epilepsy model Epilepsy Res. 2006; 70:15-26 Crossref Scopus (34) PubMed Google Scholar sourceNot checked — no full text of the source

ER is a crucial organelle affected by 2-DG treatment.

“indicating that ER is a crucial organelle affected by 2-DG treatment.”

2-DG increases Bdnf transcription via reduced N-linked glycosylation, consequent ER stress, and activity of ATF4 at an enhancer of the Bdnf gene.

“2-DG increased Bdnf transcription via reduced N-linked glycosylation, consequent ER stress, and activity of ATF4 at an enhancer of the Bdnf gene”

• Also stated in: § 2 ∙ Rajiv R. Ratan · plain

  “2-DG inhibits N-glycosylation to induce ER stress, ATF4, and Bdnf transcription”

2-DG improves AD-related learning and memory defects by normalizing BDNF levels.

“These data suggest that 2-DG improves AD-related learning and memory defects by normalizing BDNF levels (Figure 3I).”

2-DG does not induce depolarization to induce Bdnf.

“suggesting that 2-DG does not induce depolarization to induce Bdnf (Figure S1B).”

N-linked glycosylation is a sensitive indicator of 2-DG modifications in glucose metabolism.

“N-linked glycosylation is a sensitive indicator of 2-DG modifications in glucose metabolism.”

Neuronal ATF4 drives 2-DG-induced eRNA expression.

“2-DG-driven eRNA expression was also reduced in hippocampal slices with conditional deletion of neuronal ATF4, indicating that neuronal ATF4 drives 2-DG-induced eRNA expression (Figures S3G and S3H).”

Our results reinforce the positive role that ATF4 can play in mediating learning and memory, particularly in a disease-related context.

“Our results reinforce the positive role that ATF4 can play in mediating learning and memory, particularly in a disease-related context.”

2-DG regulates the expression of Bdnf and other "plasticity-associated" genes.

“These data establish that 2-DG regulates the expression of Bdnf and other “plasticity-associated” genes.”

Specific ATF4 binding to the +3 kb enhancer region is necessary for increases in Bdnf gene expression induced by 2-DG.

“Thus, specific ATF4 binding to the +3 kb enhancer region is necessary for increases in Bdnf gene expression-induced 2-DG.”

This response culminates in ATF4 binding to its response element in the +3 kb intronic enhancer region of Bdnf.

“This response culminates in ATF4 binding to its response element in the +3 kb intronic enhancer region of Bdnf (Figure 7L).”

The effect of 2-DG on recovery is delayed rather than immediate neuroprotection.

“suggesting a delayed effect on recovery rather than an immediate effect on neuroprotection (Figure 2G).”

Haploinsufficiency of Glut3 in neurons is sufficient to drive changes in glucose that result in Bdnf upregulation.

“supporting the notion that haploinsufficiency of Glut3 in neurons is sufficient to drive changes in glucose that result in Bdnf upregulation (Figures 1G–1I).”

N-linked glycosylation acts as an adaptive sensor to reduced glucose availability.

“These findings demonstrate an unrecognized role for N-linked glycosylation as an adaptive sensor to reduced glucose availability.”

• Also stated in: § 2 ∙ Rajiv R. Ratan · plain

  “N-glycosylation senses low glucose to drive adaptation to stroke and AD”

ATF4 activation is sufficient to drive BDNF-associated +3 kb enhancer activity in the hippocampus.

“Moreover, upregulation of eRNA expression (Figure 7D) suggests that ATF4 activation is sufficient to drive BDNF-associated +3 kb enhancer activity in the hippocampus.”

BDNF is significantly downregulated in post-mortem AD brains.

“Since BDNF is significantly downregulated in post-mortem AD brains”

• cites [38] 38. Hock, C. ∙ Heese, K. ∙ Hulette, C. ... Region-specific neurotrophin imbalances in Alzheimer disease: decreased levels of brain-derived neurotrophic factor and increased levels of nerve growth factor in hippocampus and cortical areas Arch. Neurol. 2000; 57:846-851 Crossref Scopus (453) PubMed Google Scholar sourceNot checked — no full text of the source

2-DG-led increases in Bdnf gene expression and LTP are dependent on transcription that shows a dominant ER stress-associated gene signature.

“These data suggest that 2-DG-led increases in Bdnf gene expression and LTP are dependent on transcription that shows a dominant ER stress-associated gene signature (Figure 5M).”

An increase in Bdnf message may not result from 2-DG-induced AMPK activation.

“Thus, an increase in Bdnf message may not result from 2-DG-induced AMPK activation (Figure 4I).”

2-DG induces sublethal ER stress.

“Our results suggest that 2-DG reduces N-linked glycosylation, induces sublethal ER stress, and drives Bdnf expression and LTP via hippocampal ATF4 in vivo (Figure 6R).”

Effects of 2-DG at doses sufficient to induce Bdnf in the brain and improve functional recovery do not require systemic ketogenesis.

“suggesting that the effects of 2-DG at doses sufficient to induce Bdnf in the brain and improve functional recovery do not require systemic ketogenesis (Figures S1D and S1E).”

2-DG-diminished N-linked glycosylation drives the 2-DG-induced Bdnf message.

“Indeed, increasing concentrations of glucose or mannose decreased 2-DG-induced Bdnf message in a concentration-dependent manner, consistent with the hypothesis that 2-DG-diminished N-linked glycosylation drives the 2-DG-induced Bdnf message (Figures 6A and 6B).”

2-DG increases LTP, a neurophysiological correlate of learning and memory, in hippocampal slices.

“Thus, 2-DG increases Bdnf gene expression in vitro and in vivo and increases LTP, a neurophysiological correlate of learning and memory, in hippocampal slices.”

2-DG injection daily for 3-4 weeks did not cause changes in physiological parameters that could affect stroke outcomes independent of BDNF.

“2-DG injection daily for 3–4 weeks did not cause changes in physiological parameters that could affect stroke outcomes independent of BDNF (Table S1).”

2-DG-induced changes in Bdnf were mediated via adaptive transcription in response to diminished N-glycosylation and ER stress.

“Additionally, 2-DG-induced changes in Bdnf were not mediated via homeostatic activation of AMPK in response to decreases in ATP (Figure 4H) but rather via adaptive transcription in response to diminished N-glycosylation and ER stress (Figures 5 and 6).”

Low glucose increases BDNF via ketone-independent pathways in cultured neurons.

“suggesting that low glucose increased BDNF via ketone-independent pathways in cultured neurons.”

Conditional deletion of Ppp1R15 leads to constitutive phosphorylation of eIF2α.

“Conditional deletion of this phosphatase leads to constitutive phosphorylation of eIF2α.57”

• cites [57]Not matched

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

The breadth of genes positively affected by 2-DG likely explains its ability to significantly improve cognitive dysfunction in an AD model and functional recovery following ischemic stroke.

“The breadth of genes positively affected by 2-DG likely explains its ability to significantly improve cognitive dysfunction in an AD model and functional recovery following ischemic stroke.”

Together with pp1, Ppp1R15 specifically catalyzes the dephosphorylation of eIF2α.

“Together with pp1, Ppp1R15 specifically catalyzes the dephosphorylation of eIF2α.”

• cites [57]Not matched

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

Safe doses of 2-DG as an anticancer agent have been assessed through a phase I/II clinical trial.

“Moreover, safe doses of 2-DG as an anticancer agent have been assessed through a phase I/II clinical trial.75”

• cites [75]Not matched

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

• cites [76]Not matched

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

2-DG competes with mannose in adding critical N-linked sugars to proteins.

“and to compete with mannose in adding critical N-linked sugars to proteins.65”

• cites [65]Not matched

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