CHARACTERIZATION OF THE CALCIUM OVERLOAD IN CULTURED NEONATAL RAT CARDIOMYOCYTES UNDER METABOLIC INHIBITION

We studied the calcium compartmentation of the Ca overload induced by 50 min of metabolic inhibition (MI) in cultured neonatal rat cardiomyo cytes. MI was achieved by application of 1 mM iodoacetic acid (IAA) an d 10 mM 2-deoxyglucose (2-DOG) with omission of glucose in the perfusa te. MI per se abolished spontaneous beating, but 10 mM caffeine pulses at 15, 30 and 45 min were able to induce contractile responses. The c ontractile responses were abolished in the presence of Thg. After 20 m in of MI, cells began to accumulate Ca. After 50 min of MI, the Ca exc hanged via Na/Ca exchange (Na/Ca exchange-dependent Ca content) was in creased by 3.21 +/- 0.15 mmol/kg dry weight (n = 5). The net increase remaining exchangeable via Na/Ca exchange represents 80% of the global net Ca-45 uptake after 50 min in MI which was 3.98 mmol/kg dry weight , obtained in the ''on-line'' experiments. The release of this compone nt of Ca is completed within 150 s via Na/Ca exchange. There also was a 50% increase in the Ca content of the so-called ''slow'' compartment , 0.44 +/- 0.08 mmol/kg dry weight (n = 10) after 50 min under MI. The t(1/2) of this compartment was 24.8 +/- 3.8 min typical of a mitochon drial origin in these cells. Thus 92% of the net Ca increase after 50 min MI remains exchangeable-80% of this is completed within 150 s via Na/Ca exchange and 20% more slowly, t(1/2) about 25 min. Cell ATP cont ent, 27.35 +/- 2.07 nmoles/mg protein, was decreased after 10 min of M I to 2.25 +/- 0.29 nmoles/mg protein (n = 10) and remained close to th is level after 20 min of MI (2.11 +/- 0.17 nmoles/mg protein; n = 6). It is to be noted that our results indicate no compromise of the Na/Ca exchange system after 50 min of MI although the global ATP is deplete d in these cells. (C) 1996 Academic Press Limited