Regulation of Ca signaling by acute hypoxia and acidosis in rat neonatal cardiomyocytes.

Unlabelled: Ischemic heart disease is an arrhythmogenic condition, accompanied by hypoxia, acidosis, and impaired Ca signaling. Here we report on effects of acute hypoxia and acidification in rat neonatal cardiomyocytes cultures.

Results: Two populations of neonatal cardiomyocyte were identified based on inactivation kinetics of L-type I: rapidly-inactivating I (τ~20ms) myocytes (prevalent in 3-4-day cultures), and slow-inactivating I (τ≥40ms) myocytes (dominant in 7-day cultures). Acute hypoxia (pO<5mmHg for 50-100s) suppressed I reversibly in both cell-types to different extent and with different kinetics. This disparity disappeared when Ba was the channel charge carrier, or when the intracellular Ca buffering capacity was increased by dialysis of high concentrations of EGTA and BAPTA, suggesting critical role for calcium-dependent inactivation. Suppressive effect of acute acidosis on I (~40%, pH6.7), on the other hand, was not cell-type dependent. Isoproterenol enhanced I in both cell-types, but protected only against suppressive effects of acidosis and not hypoxia. Hypoxia and acidosis suppressed global Ca transients by ~20%, but suppression was larger, ~35%, at the RyR2 microdomains, using GCaMP6-FKBP targeted probe. Hypoxia and acidosis also suppressed mitochondrial Ca uptake by 40% and 10%, respectively, using mitochondrial targeted Ca biosensor (mito-GCaMP6).

Conclusion: Our studies suggest that acute hypoxia suppresses I in rapidly inactivating cell population by a mechanism involving Ca-dependent inactivation, while compromised mitochondrial Ca uptake seems also to contribute to I suppression in slowly inactivating cell population. Proximity of cellular Ca pools to sarcolemmal Ca channels may contribute to the variability of inactivation kinetics of I in the two cell populations, while acidosis suppression of I appears mediated by proton-induced block of the calcium channel.