Journal
CIRCULATION RESEARCH
Volume 95, Issue 3, Pages 325-332Publisher
LIPPINCOTT WILLIAMS & WILKINS
DOI: 10.1161/01.RES.0000137727.34938.35
Keywords
ischemic preconditioning; cardiomyocytes; ischemia; potassium channels; gene transfer
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Volume regulatory Cl- channels are key regulators of ischemic preconditioning (IPC). Because Cl- efflux must be balanced by an efflux of cations to maintain cell membrane electroneutrality during volume regulation, we hypothesize that I-K1 channels may play a role in IPC. We subjected cultured cardiomyocytes to 60-minute simulated ischemia ( SI) followed by 60-minute of simulated reperfusion (SR) and assessed percent cell death using trypan blue staining. Ischemic preconditioning (10-minute SI/10-minute SR) significantly (P < 0.0001) reduced the percent cell death in nontransfected cardiomyocytes [IPCCM 18.0 ± 2.1% versus control (C-CM) 48.3 ± 1.0%]. IPC protection was not altered by overexpression of the reporter gene ( enhanced green fluorescent protein, EGFP). However, overexpression of dominant-negative Kir2.1 or Kir2.2 genes using adenoviruses (AdEGFPKir2.1DN or AdEGFPKir2.2DN) encoding the reporter gene EGFP prevented IPC protection [both IPCCM + AdEGFPKir2.1DN 45.8 ± 2.3% (mean ± SEM) and IPCCM + AdEGFPKir2.2DN 47.9 ± 1.4% versus IPCCM; P < 0.0001] in cultured cardiomyocytes (n =8 hearts). Transfection of cardiomyocytes with AdEGFPKir2.1DN or AdEGFPKir2.2DN did not affect cell death in control (nonpreconditioned) cardiomyocytes (both C-CM +AdEGFPKir2.1DN 45.8 +/- 0.7% and C-CM + dEGFPKir2.2DN 46.2 +/- 1.3% versus C-CM; not statistically significant). Similar effects were observed in both cultured (n = 5 hearts) and freshly isolated (n = 4 hearts) ventricular cardiomyocytes after I-K1 blockade with 20 mumol/L BaCl2 plus 1 mumol/L nifedipine (to prevent Ba2+ uptake). Nifedipine alone neither protected against ischemic injury nor blocked IPC protection. Our findings establish that I-K1 channels play an important role in IPC protection.
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