EFFECTS OF ANOXIA ON INTRACELLULAR FREE CA2+ IN ISOLATED CARDIOMYOCYTES FROM TURTLES

One of the most important negative consequences of hypoxic stress in t he mammalian myocardium is a breakdown in intracellular calcium homeos tasis. This study examines the effects of anoxic stress on Intracellul ar calcium regulation in isolated ventricular myocytes from a hypoxia tolerant vertebrate, the western painted turtle (Chrysemys picta belli i). Isolated calcium tolerant cardiomyocytes from turtle hearts were m ounted on a glass cover slip that formed the bottom of a sealed, Plexi glas perfusion chamber. Free [Ca2+](i) (determined by FURA2 fluorescen ce) in isolated turtle cardiomyocytes averaged 31.7 +/- 3.2 nM after 3 0 min of normoxic perfusion (20 degrees C, pH(e) = 7.77). This value i s on the low end of the published range for mammalian cardiomyocytes. Perfusion with anoxic Ringer equilibrated with 3% CO2, resulted in a s ignificant increase in free [Ca2+](i) to 941 +/- 494.6 nM after 60 min . Increasing the CO2 in the perfusion solution to 5% or 6% blunted thi s rise (peak levels after 60 min of anoxia were 420.5 +/- 176.0 nM and 393.8 +/- 132.8 nM, respectively). A further increase to 8% CO2 incre ased the maximal value for free [Ca2+](i) to 610.9 +/- 297.5 nM. In ei ght cells from the 5% CO2 protocol in which [Ca2+](i) was monitored du ring recovery, reperfusion with normoxic Ringer rapidly lowered intrac ellular calcium to 92.8 +/- 9.7 nM within 15 min. Anoxia at relatively high extracellular (and hence intracellular) pH results in an increas e in free [Ca2+](i) comparable in magnitude and time course to that se en in some mammalian cardiomyocyte preparations. Perfusion of anoxic m yocytes with Ringer equilibrated with either 5% or 6% CO2 blunted this increase in intracellular calcium, possibly an example of the pH para dox effect. A more severe combination of respiratory acidosis and anox ia (8% CO2) removed this protective effect. Copyright (C) 1997 Elsevie r Science Inc.