The effects of purified mojave toxin on rat synaptic membrane (Ca +2+Mg +2)-ATPase and the dihydropyridine receptor

Abstract

Effects of purified Mojave toxin on rat synaptic membrane (Ca +2+Mg +2)-ATPase and dihydropyridine receptor were determined. The toxin was observed to stimulate specifically (Ca +2+Mg +2)-ATPase approximately two-fold with no effect on Mg +2 dependent ATPase activity. Examination of the effects of increasing amounts of purified Mojave toxin on binding of the calcium channel blocker, nitrendipine, indicated that the addition of 10 μ,g (4.5 × 10 −10 moles) of toxin resulted in greater than 90% inhibition of nitrendipine binding. Furthermore, binding studies revealed the toxin to have little affinity for the ligand indicating its interaction with calcium channel components. Since Mojave toxin has associated with it a phospholipase A 2 activity, we investigated the effects of 4-bromophenacylbromide, a known inhibitor of phospholipase A 2 activity in order to discern the possible effects of the purified toxin on synaptic membranes. At concentrations previously shown to be inhibitory of purified phospholipase A 2 from cobra venom, both ATPase activity and nitrendipine binding of synaptic membranes were significantly inhibited. Thus we cannot rule out the possibility that the endogenous phospholipase activity of the purified toxin is responsible for its effects on the rat brain synaptic functions studied here. Binding studies conducted in the presence of verapamil and diltiazem indicated that the toxin interacts with allosteric sites responsible for regulation of the binding of nitrendipine. Although we have not tested the effects of Mojave toxin on other ion channels and/or receptors, results presented here suggest the potential usefulness of this toxin as a molecular probe of the calcium channel complex.