Abstract
A variety of direct and indirect techniques have revealed the existence of ATP-sensitive potassium (KATP) channels in the inner membranes of mitochondria. The molecular identity of these mitochondrial KATP (mitoKATP) channels remains unclear. We used a pharmacological approach to distinguish mitoKATP channels from classical, molecularly defined cardiac sarcolemmal KATP (surfaceKATP) channels encoded by the sulfonylurea receptor SUR2A and the pore-forming subunit Kir6.2. SUR2A and Kir6.2 were expressed in human embryonic kidney (HEK)293 cells, and their activities were measured by patch-clamp recordings of membrane current. SurfaceKATP channels are activated potently by 100 μM pinacidil but only weakly by 100 μM diazoxide; in addition, they are blocked by 10 μM glibenclamide, but are insensitive to 500 μM 5-hydroxydecanoate. This pharmacology, which was confirmed with patch-clamp recordings in intact rabbit ventricular myocytes, contrasts with that of mitoKATP channels as indexed by flavoprotein oxidation. MitoKATP channels in myocytes are activated equally by 100 μM diazoxide and 100 μM pinacidil. In contrast to its lack of effect on surfaceKATP channels, 5-hydroxydecanoate is an effective blocker of mitoKATPchannels. Glibenclamide’s effects on mitoKATP channels are difficult to assess, because it independently activates flavoprotein fluorescence, consistent with a previously described primary uncoupling effect. Confocal imaging of the subcellular distribution of expressed fluorescent Kir6.2 in HEK cells and in myocytes revealed no targeting of mitochondrial membranes. The differences in drug sensitivity and subcellular localization indicate that mitoKATP channels are distinct from surface KATP channels at a molecular level.
Footnotes
- Received August 18, 1998.
- Accepted March 7, 1999.
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Send reprint requests to: Eduardo Marbán, M.D., Ph.D., 844 Ross Bldg., The Johns Hopkins University School of Medicine, Baltimore, MD 21205. E-mail: marban{at}jhmi.edu
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↵1 These authors contributed equally to the work.
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↵2 Current address: Osiris Therapeutics, Inc., 2001 Aliceanna St., Baltimore, MD 21231.
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↵3 Current address: Maryland Research Laboratories, Otsuka America Pharmaceutical Inc., Rockville, MD 20850
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This work was supported by National Insitutes of Health Grants R01 HL52768 and R01 HL44065 (to E.M.), and T32 HL07227 (to H.H.), an American Heart Association-Maryland Fellowship (to H.H.), and a Banyu Fellowship in Lipid Metabolism and Atherosclerosis (to T.S.).
- The American Society for Pharmacology and Experimental Therapeutics
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