COMPARATIVE EFFECTS OF INORGANIC-PHOSPHATE AND OXALATE ON UPTAKE AND RELEASE OF CA2-RETICULUM IN SAPONIN SKINNED RAT CARDIAC TRABECULAE( BYTHE SARCOPLASMIC)

1. Ventricular trabeculae from the right ventricle of rat heart were s uspended in a 6 mu l bath and 'skinned' with saponin (50 mg ml(-1)). P reparations were perfused with solutions mimicking the intracellular m ilieu and the [Ca2+] within the bath was monitored continuously using fura-2. 2. Application of 20 mM caffeine released Ca2+ from the sarcop lasmic reticulum (SR), resulting in a transient increase in the fura-2 fluorescence ratio. Caffeine-induced Ca2+ transients were smaller in the presence of 30 or 60 mM inorganic phosphate (P-i). This depressive effect of P-i on SR function was reversed by 10 mM creatine phosphate (CP). Caffeine-induced Ca2+ transients were also reduced in the prese nce of 10 mM oxalate, although this effect was not reversed by CP. 3. When perfusion was stopped in the presence of 30 or 60 mM P-i, the [Ca 2+] within the bath remained constant. However, when the flow was stop ped in the presence of 60 mM P-i and 10 mM CP, a prolonged decrease in [Ca2+] occurred, consistent with precipitation of calcium phosphate w ithin the SR. A similar decrease in [Ca2+] was observed when perfusion was stopped in the presence of 2 or 20 mM oxalate, in the absence or presence of CP. 4. The SR was Ca2+ depleted by withdrawal of ATP and e xposure to 20 mM caffeine. Perfusion was then stopped and ATP reapplie d, resulting in a maintained decrease in [Ca2+] within the bath, due t o SR Ca2+ uptake. Net Ca2+ uptake was markedly reduced in the presence of 30 mM P-i. In contrast, 20 mM oxalate increased Ca2+ uptake and th e [Ca2+] within the bath continued to fall over 2-3 min.5. Introductio n of P-i released Ca2+ from the SR. Ryanodine (100 mu M) abolished caf feine-induced Ca2+ release while P-i-induced Ca2+ release was unaffect ed. P-i-induced Ca2+ release was reduced in the constant presence of 2 0 mM caffeine or 10 mM CP and was abolished completely by disruption o f the SR membrane with Triton X-100. P-i-induced Ca2+ release occurred after abolition of SR Ca2+ uptake by ATP withdrawal. 6. These results suggest that the P-i-induced decrease in releasable Ca2+ does not res ult from precipitation of calcium phosphate within the SR lumen. P-i i nhibits net SR Ca2+ uptake, but this appears to result from activation of a ryanodine-insensitive Ca2+ efflux pathway rather then inhibition of Ca2+ uptake. Possible mechanisms are considered, including reversa l of the SR Ca2+ pump.