EFFECT OF ALPHA-LATROTOXIN ON ACETYLCHOLINE-RELEASE AND INTRACELLULAR CA-2+ CONCENTRATION IN SYNAPTOSOMES - NA+-DEPENDENT AND NA+-INDEPENDENT COMPONENTS

We studied the effect of alpha-latrotoxin (alphaLTX) on [C-14]acetylcholine ([C-14]ACh) release, intracellular Ca2+ concentration ([Ca2+]i), plasma membrane potential, and high-affinity choline uptake of synaptosomes isolated from guinea pig cortex. AlphaLTX (10(-10)-10(-8) M) caused an elevation of the [Ca2+]i as detected by Fura 2 fluorescence and evoked [C-14]ACh efflux. Two components in the action of the toxin were distinguished: one that required the presence of Na+ in the external medium and another that did not. Displacement of Na+ by sucrose or N-methylglucamine in the medium considerably decreased the elevation of [Ca2+]i and [C-14]ACh release by alphaLTX. The Na+-dependent component of the alphaLTX action was obvious in the inhibition of the high-affinity choline uptake of synaptosomes. Some of the toxin action on both [Ca2+]i and [C-14]ACh release remained in the absence of Na+. Both the Na+-dependent and the Na+-independent components of the alphaLTX-evoked [C-14]ACh release partly required the presence of either Mg2+ or Ca2+. The nonneurotransmitter [C-14]choline was released along with [C-14]ACh, but this release did not depend on the presence of either Na+ or Ca2+, indicating nonspecific leakage through the plasma membrane. We conclude that there are two factors in the release of ACh from synaptosomes caused by the toxin: (1) cation-dependent ACh release, which is related to (a) Na+-dependent divalent cation entry and (b) Na+-independent divalent cation entry, and (2) nonspecific Na+- and divalent cation-independent leakage.