Characterization of human cardiac mitochondrial ATP-sensitive potassium channel and its regulation by phorbol ester in vitro.
Activation of the mitochondrial ATP-sensitive K+ channel (mitoKATP) and its regulation by PKC are critical events in preconditioning induced by ischemia or pharmaceutical agents in animals and humans. The properties of the human cardiac mitoKATP channel are unknown. Furthermore, there is no evidence that cytosolic PKC can directly regulate the mitoKATP channel located in the inner mitochondrial membrane (IMM) due to the physical barrier of the outer mitochondrial membrane. In the present study, we characterized the human cardiac mitoKATP channel and its potential regulation by PKC associated with the IMM. IMM fractions isolated from human left ventricles were fused into lipid bilayers in symmetrical potassium glutamate (150 mM). The conductance of native mitoKATP channels was usually below 80 pS ( approximately 70%), which was reduced by ATP and 5-hydroxydecanoic acid (5-HD) in a dose- and time-dependent manner. The native mitoKATP channel is activated by diazoxide and inhibited by ATP and 5-HD. The PKC activator phorbol 12-myristate 13-acetate (2 microM) increased the cumulative open probability of the mitoKATP channel previously inhibited by ATP (P < 0.05), but its inactive analog 4alpha-phorbol 12,13-didecanoate had no effect. Western blot analysis detected an inward rectifying K+ channel (Kir6.2) immunoreactive protein at 56 kDa and PKC-delta in the IMM. These data provide the first characterization of the human cardiac mitoKATP channel and its regulation by PKC(s) in IMM. This local PKC control mechanism may represent an alternative pathway to that proposed previously for cytosolic PKC during ischemic/pharmacological preconditioning.
Jiang
Ming Tao
MT
Department of Anesthesiology, Medical College of Wisconsin, 8701 Watertown Plank Rd., Milwaukee, WI 53226, USA. mtjiang@mcw.edu
Ljubkovic
Marko
M
Nakae
Yuri
Y
Shi
Yang
Y
Kwok
Wai-Meng
WM
Stowe
David F
DF
Bosnjak
Zeljko J
ZJ
D000255:Adenosine Triphosphate / Q000378:metabolism*; D002478:Cells, Cultured; D006801:Humans; D015640:Ion Channel Gating / Q000187:drug effects / Q000502:physiology*; D008051:Lipid Bilayers / Q000378:metabolism; D008929:Mitochondria, Heart / Q000378:metabolism*; D024101:Mitochondrial Proteins / Q000378:metabolism*; D017202:Myocardial Ischemia / Q000378:metabolism; D010703:Phorbol Esters / Q000494:pharmacology*; D015221:Potassium Channels / Q000378:metabolism*; D011493:Protein Kinase C / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology
D008051:Lipid Bilayers; D024101:Mitochondrial Proteins; D010703:Phorbol Esters; D015221:Potassium Channels; C117780:mitochondrial K(ATP) channel; D000255:Adenosine Triphosphate; D011493:Protein Kinase C