ATP-dependent and ATP-independent calcium buffers determined in isolated squid axoplasm by axoplasmic dilution using...

Ca2+ buffering in axoplasm extruded from the squid giant axon was studied by axoplasmic dilution and Ca2+-selective electrodes. ATP-dependent Ca2+ buffering was lost when metabolism and endogenous ATP production were inhibited by axoplasmic dilution. Under these conditions ('low ATP' axoplasmic suspensions) Ca2+ activity (aCa) increased spontaneously. This increase was prevented by ATP addition. Addition of a Ca2+ load to axoplasmic suspensions following ATP addition ('normal ATP' axoplasmic suspensions) did not increase aCa. When a Ca2+ load was added to low ATP axoplasmic suspensions the value of aCa was only 5% of that expected from the Ca2+ load in a non-Ca2+ buffered system. Addition of the Ca2+ ionophore A23187 to Ca2+-loaded normal ATP axoplasmic suspensions increased aCa to a value 5% of that expected in a non-Ca2+ buffered system. Addition of A23187 or KCN to low ATP axoplasmic suspensions did not increase aCa above that produced by axoplasmic dilution. These data indicate the presence of ATP-dependent and ATP-independent Ca2+ buffering mechanisms in the squid axoplasm and that the ATP-dependent Ca2+ buffering was mediated by membrane-limited organelles presumably the mitochondria and endoplasmic reticulum.