Hypoxia-induced preconditioning in adult stimulated cardiomyocytes is mediated by the opening and trafficking of sarcolemmal KATP channels.
The opening of sarcolemmal and mitochondrial ATP-sensitive K(+) (KATP) channels in the heart is believed to mediate ischemic preconditioning, a phenomenon whereby brief periods of ischemia/reperfusion protect the heart against myocardial infarction. Here, we have applied digital epifluorescent microscopy, immunoprecipitation and Western blotting, perforated patch clamp electrophysiology, and immunofluorescence/laser confocal microscopy to examine the involvement of KATP channels in cardioprotection afforded by preconditioning. We have shown that adult, stimulated-to-beat, guinea-pig cardiomyocytes survived in sustained hypoxia for approximately 17 min. An episode of 5-min-long hypoxia/5-min-long reoxygenation before sustained hypoxia dramatically increased the duration of cellular survival. Experiments with different antagonists of KATP channels, applied at different times during the experimental protocol, suggested that the opening of sarcolemmal KATP channels at the beginning of sustained hypoxia mediate preconditioning. This conclusion was supported by perforated patch clamp experiments that revealed activation of sarcolemmal KATP channels by preconditioning. Immunoprecipitation and Western blotting as well as immunofluorescence and laser confocal microscopy showed that the preconditioning is associated with the increase in KATP channel proteins in sarcolemma. Inhibition of trafficking of KATP channel subunits prevented preconditioning without affecting sensitivity of cardiomyocytes to hypoxia in the absence of preconditioning. We conclude that the preconditioning is mediated by the activation and trafficking of sarcolemmal KATP channels.
Budas
Grant R
GR
Maternal and Child Health Sciences, Tayside Institute of Child Health, Ninewells Hospital & Medical School, University of Dundee, Dundee, Scotland, UK.
Jovanovic
Sofija
S
Crawford
Russell M
RM
Jovanovic
Aleksandar
A
D000255:Adenosine Triphosphate / Q000378:metabolism*; D000818:Animals; D016923:Cell Death / Q000187:drug effects; D015687:Cell Hypoxia / Q000187:drug effects / Q000502:physiology*; D002470:Cell Survival / Q000187:drug effects; D006168:Guinea Pigs; D015640:Ion Channel Gating / Q000187:drug effects; D019157:Ischemic Preconditioning, Myocardial; D008297:Male; D032383:Myocytes, Cardiac / Q000166:cytology / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D018408:Patch-Clamp Techniques; D026902:Potassium Channel Blockers / Q000494:pharmacology; D015221:Potassium Channels / Q000378:metabolism*; D021381:Protein Transport / Q000187:drug effects; D015427:Reperfusion Injury / Q000473:pathology / Q000503:physiopathology; D012508:Sarcolemma / Q000378:metabolism*; D013997:Time Factors
D026902:Potassium Channel Blockers; D015221:Potassium Channels; D000255:Adenosine Triphosphate