Droese 2006 J Biol Chem: Difference between revisions
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{{Publication | {{Publication | ||
|title=DrΓΆse S, Brandt U, Hanley PJ (2006) K+-independent actions of diazoxide question the role of inner membrane | |title=DrΓΆse S, Brandt U, Hanley PJ (2006) K<sup>+</sup>-independent actions of diazoxide question the role of inner membrane K<sub>ATP</sub> channels in mitochondrial cytoprotective signaling. J Biol Chem 281:23733β9. | ||
|authors=Droese S, Brandt U, Hanley PJ Β | |info=[http://www.ncbi.nlm.nih.gov/pubmed/16709571 PMID: 16709571 Open Access] | ||
|authors=Droese S, Brandt U, Hanley PJ | |||
|year=2006 | |year=2006 | ||
|journal= | |journal=J Biol Chem | ||
|abstract=Activation by diazoxide and inhibition by 5-hydroxydecanoate are the hallmarks of mitochondrial ATP-sensitive K+ ( | |abstract=Activation by diazoxide and inhibition by 5-hydroxydecanoate are the hallmarks of mitochondrial ATP-sensitive K<sup>+</sup>(K<sub>ATP</sub>) channels. Opening of these channels is thought to trigger cytoprotection (preconditioning) through the generation of reactive oxygen species. However, we found that diazoxide-induced oxidation of the widely used reactive oxygen species indicator 2β²,7β²-dichlorodihydrofluorescein in isolated liver and heart mitochondria was observed in the absence of ATP or K<sup>+</sup> and therefore independent of K<sub>ATP</sub> channels. The response was blocked by stigmatellin, implying a role for the cytochrome ''bc''<sub>1</sub> complex (Complex III). Diazoxide, though, did not increase hydrogen peroxide (H<sub>2</sub>O<sub>2</sub>) production (quantitatively measured with Amplex Red) in intact mitochondria, submitochondrial particles, or purified cytochrome ''bc''<sub>1</sub> complex. We confirmed that diazoxide inhibited succinate oxidation, but it also weakly stimulated State 4 respiration even in K<sup>+</sup>-free buffer, excluding a role for K<sub>ATP</sub> channels. Furthermore, we have shown previously that 5-hydroxydecanoate is partially metabolized, and we hypothesized that fatty acid metabolism may explain the ability of this putative mitochondrial K<sub>ATP</sub> channel blocker to inhibit diazoxide-induced flavoprotein fluorescence, commonly used as an assay of K<sub>ATP</sub> channel activity. Indeed, consistent with our hypothesis, we found that decanoate inhibited diazoxide-induced flavoprotein oxidation. Taken together, our data question the βmitochondrial K<sub>ATP</sub> channelβ hypothesis of preconditioning. Diazoxide did not evoke superoxide (which dismutates to H<sub>2</sub>O<sub>2</sub>) from the respiratory chain by a direct mechanism, and the stimulatory effects of this compound on mitochondrial respiration and 2β²,7β²-dichlorodihydrofluorescein oxidation were not due to the opening of K<sub>ATP</sub> channels. | ||
| | |mipnetlab=NL Nijmegen Brandt U, DE Frankfurt Droese S | ||
|discipline=Mitochondrial Physiology, Pharmacology; Biotechnology | |||
}} | }} | ||
{{Labeling | {{Labeling | ||
| | |area=Respiration, Pharmacology;toxicology | ||
| | |organism=Rat | ||
| | |tissues=Heart, Liver | ||
| | |preparations=Isolated mitochondria | ||
|instruments=Oxygraph-2k | |injuries=Oxidative stress;RONS | ||
|instruments=Oxygraph-2k | |||
}} | }} |
Latest revision as of 17:04, 19 February 2018
DrΓΆse S, Brandt U, Hanley PJ (2006) K+-independent actions of diazoxide question the role of inner membrane KATP channels in mitochondrial cytoprotective signaling. J Biol Chem 281:23733β9. |
Droese S, Brandt U, Hanley PJ (2006) J Biol Chem
Abstract: Activation by diazoxide and inhibition by 5-hydroxydecanoate are the hallmarks of mitochondrial ATP-sensitive K+(KATP) channels. Opening of these channels is thought to trigger cytoprotection (preconditioning) through the generation of reactive oxygen species. However, we found that diazoxide-induced oxidation of the widely used reactive oxygen species indicator 2β²,7β²-dichlorodihydrofluorescein in isolated liver and heart mitochondria was observed in the absence of ATP or K+ and therefore independent of KATP channels. The response was blocked by stigmatellin, implying a role for the cytochrome bc1 complex (Complex III). Diazoxide, though, did not increase hydrogen peroxide (H2O2) production (quantitatively measured with Amplex Red) in intact mitochondria, submitochondrial particles, or purified cytochrome bc1 complex. We confirmed that diazoxide inhibited succinate oxidation, but it also weakly stimulated State 4 respiration even in K+-free buffer, excluding a role for KATP channels. Furthermore, we have shown previously that 5-hydroxydecanoate is partially metabolized, and we hypothesized that fatty acid metabolism may explain the ability of this putative mitochondrial KATP channel blocker to inhibit diazoxide-induced flavoprotein fluorescence, commonly used as an assay of KATP channel activity. Indeed, consistent with our hypothesis, we found that decanoate inhibited diazoxide-induced flavoprotein oxidation. Taken together, our data question the βmitochondrial KATP channelβ hypothesis of preconditioning. Diazoxide did not evoke superoxide (which dismutates to H2O2) from the respiratory chain by a direct mechanism, and the stimulatory effects of this compound on mitochondrial respiration and 2β²,7β²-dichlorodihydrofluorescein oxidation were not due to the opening of KATP channels.
β’ O2k-Network Lab: NL Nijmegen Brandt U, DE Frankfurt Droese S
Labels: MiParea: Respiration, Pharmacology;toxicology
Stress:Oxidative stress;RONS Organism: Rat Tissue;cell: Heart, Liver Preparation: Isolated mitochondria
HRR: Oxygraph-2k