Inhibitory effect of EDTA · Ca2+ on the hydrolysis of synaptosomal phospholipids by phospholipase A2 toxins and enzymes

Abstract Phospholipases A 2 (PLA 2 ) are Ca 2+ -dependent enzymes that are inhibited by EDTA; this inhibition would be expected to be reversed by restoring the Ca 2+ concentration. By examining the hydrolysis of synaptosomal phospholipids by PLA 2 enzymes, Naja naja atra and Naja nigricollis , and by toxins with PLA 2 activity, β-bungarotoxin (β-BuTX) and notexin, we demonstrated a novel inhibitory action of EDTA manifested in the presence of excess Ca 2+ . We postulate the formation of an EDTA·Ca 2+ complex which inhibits PLA 2 activity in connection-dependent manner. Synaptosomes in which phospholipids are hydrolyzed by PLA 2 , have membranal damage expressed by increased acetylcholine (ACh) release and decreased osmotic activity. Addition of EDTA·Ca 2+ , which inhibits phospholipid hydrolysis, also reversed the PLA 2 effect on ACh release, but not its effect on osmotic activity. The inhibition of PLA 2 was observed on membranal phospholipids as well as on an artificial substrate of phospholipid-Triton mixed micelles. Moreover, we found that another enzyme, lactate dehydrogenase, was also inhibited. Our results indicate a non-specific inhibition exerted on the enzyme rather than on the substrate.