Lack of manifestations of diazoxide/5‐hydroxydecanoate‐sensitive KATP channel in rat brain nonsynaptosomal mitochondria
Abstract
Pharmacological modulation of the mitochondrial ATP‐sensitive K+ channel (mitoK ATP) sensitive to diazoxide and 5‐hydroxydecanoate (5‐HD) represents an attractive strategy to protect cells against ischaemia/reperfusion‐ and stroke‐related injury. To re‐evaluate a functional role for the mitoK ATP in brain, we used Percoll‐gradient‐purified brain nonsynaptosomal mitochondria in a light absorbance assay, in radioisotope measurements of matrix volume, and in measurements of respiration, membrane potential (ΔΨ) and depolarization‐induced K+ efflux. The changes in mitochondrial morphology were evaluated by transmission electron microscopy (TEM). Polyclonal antibodies raised against certain fragments of known sulphonylurea receptor subunits, SUR1 and SUR2, and against different epitopes of K+ inward rectifier subunits Kir 6.1 and Kir 6.2 of the ATP‐sensitive K+ channel of the plasma membrane (cellK ATP), were employed to detect similar subunits in brain mitochondria. A variety of plausible blockers (ATP, 5‐hydroxydecanoate, glibenclamide, tetraphenylphosphonium cation) and openers (diazoxide, pinacidil, chromakalim, minoxidil, testosterone) of the putative mitoK ATP were applied to show the role of the channel in regulating matrix volume, respiration, and ΔΨ and K+ fluxes across the inner mitochondrial membrane. None of the pharmacological agents applied to brain mitochondria in the various assays pinpointed processes that could be unequivocally associated with mitoK ATP activity. In addition, immunoblotting analysis did not provide explicit evidence for the presence of the mitoK ATP, similar to the cellK ATP, in brain mitochondria. On the other hand, the depolarization‐evoked release of K+ suppressed by ATP could be re‐activated by carboxyatractyloside, an inhibitor of the adenine nucleotide translocase (ANT). Moreover, bongkrekic acid, another inhibitor of the ANT, inhibited K+ efflux similarly to ATP. These observations implicate the ANT in ATP‐sensitive K+ transport in brain mitochondria.
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KATP channel subunits could not be identified in highly purified mitochondria, challenging the existence of mitoKATP.
“Especially, the fact that the K ATP channel subunits could not be identified so far in highly purified mitochondria challenges its existence [145,146].”
From Differential effects of complex II on mitochondrial ROS production and their relation to cardioprotective pre- and postconditioning (2013) · cites this paper as [145]
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O'Sullivan, J. C., Fu, D., Alam, H. B., & McCabe, J. T. (2008) · Journal of Surgical Research · doi:10.1016/j.jss.2007.12.750 · Cited by 8 · see citing works
Immunolocalization of mitoKATP subunits in human osteoblast-like cells
Butler, J. (2010) · Frontiers in Bioscience-Elite · doi:10.2741/e134 · Cited by 2 · see citing works
Levin, S. G., Shamsutdinova, A. A., & Godukhin, O. V. (2013) · Bulletin of Experimental Biology and Medicine · doi:10.1007/s10517-013-1972-0 · Cited by 1 · see citing works
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Electrophysiological Techniques for Mitochondrial Channels
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- Authors
- Brustovetsky, T., Shalbuyeva, N., & Brustovetsky, N.
- Affiliations
- Indiana University School of Medicine (US)
- Indiana University Indianapolis (US)
- Venue
- The Journal of Physiology
- Publisher
- Wiley
- Published
- 2005-09-28
- Volume
- 568
- Issue
- 1
- Pages
- 47-59
- ISSN
- 0022-3751 (ISSN-L)
- 1469-7793
- OpenAlex
- W1597339748
- Cited by
- 46
- References
- 59
- Citations by year
- 2022: 1 · 2021: 1 · 2019: 2 · 2017: 1 · 2016: 1 · 2015: 2 · 2014: 1 · 2013: 4 · 2012: 5
- FWCI
- 2.64
- Type
- journal article
- Language
- en
- Open access
- closed
- Locations
- The Journal of Physiology (publishedVersion)
- PubMed (publishedVersion)
- Funders
- National Institute of Neurological Disorders and Stroke (R01NS050131)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Mitochondrial Function and Pathology
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Ion channel regulation and function
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › ATP Synthase and ATPases Research
- Keywords
- mitochondrial ATP-sensitive potassium channel, diazoxide, 5-hydroxydecanoate, brain mitochondria, sulfonylurea receptor, Kir6.1, Kir6.2, mitochondrial membrane potential, adenine nucleotide translocase, carboxyatractyloside, bongkrekic acid, ATP, mitochondrial respiration, ischemia-reperfusion injury, stroke
- MeSH terms
- Kcnj11 Channel, Mitochondrial ADP, ATP Translocases, Adenosine Triphosphate, Animals, Atractyloside, Brain, Culture Media, Decanoic Acids, Diazoxide, Hydroxy Acids, Male, Mitochondria, Oxygen Consumption, Potassium, Potassium Chloride, Receptors, Drug, Potassium Channels, Rats, Sprague-Dawley, ATP-Binding Cassette Transporters, Potassium Channels, Inwardly Rectifying, Potassium Channel Blockers, Rats, KATP Channels, Sulfonylurea Receptors, In Vitro Techniques
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