Comparative effects of phospholipase A2 neurotoxins and enzymes on membrane potential and Na+/K+ ATPase activity in rat brain synaptosomes.
Beta-Bungarotoxin (beta-BuTX) and notexin are phospholipase A2 (PLA2) neurotoxins which cause an irreversible blockade of neurotransmitter release through specific and potent effects at the presynaptic nerve terminal; however, their mechanism of action is uncertain. We examined the effects of beta-BuTX and notexin on Na+/K+ ATPase activity using Sprague-Dawley rat brain synaptosomes in order to determine if alterations in activity might modulate neurotoxin-induced depolarization. Treatment of synaptosomes with 0.05 to 5 nM beta-BuTX, notexin, and Naja naja atra and Naja nigricollis PLA2 (PLA2 enzymes without selective presynaptic actions) caused a dose-dependent depolarization of synaptosomes with no differences being observed between the effects of the PLA2 neurotoxins and enzymes. N. nigricollis PLA2 (0.5 nM; 20 min) slightly stimulated Na+/K+ ATPase activity while beta-BuTX and notexin (0.5 nM: 10 and 20 min) were without effect. With 50 nM concentrations beta-BuTX and notexin stimulated Na+/K+ ATPase activity, while N. nigricollis and N. n. atra PLA2 inhibited activity. The effects on membrane potential and Na+/K+ ATPase were antagonized or blocked by EDTA (10 mM) and bovine serum albumin (1 mg/ml), suggesting that PLA2 enzymatic activity is essential for their effects on membrane potential and Na+/K+ ATPase activity. Following neurotoxin and enzyme pretreatment, we found a biphasic correlation between synaptosomal free fatty acid (FFA) levels and Na+/K+ ATPase activity, where Na+/K+ ATPase is stimulated by low levels of FFA (0.13 to 0.22 mumol/mg protein) and antagonized by FFA levels in excess of 0.34 mumol/mg protein. In contrast there was a linear correlation between the extent of FFA production and membrane depolarization. We propose that the presynaptic depolarizing actions of beta-BuTX and notexin are not mediated through modulation of Na+/K+ ATPase activity and that the changes observed in ATPase activity and possibly membrane potential are directly due to PLA2 enzymatic activity and the production of FFA.
Yates
S L
SL
Section of Pharmacology and Toxicology, University of Connecticut, School of Pharmacy Storrs 06269.
Rosenberg
P
P
D000818:Animals; D001921:Brain / Q000187:drug effects* / Q000378:metabolism; D002038:Bungarotoxins / Q000633:toxicity; D004305:Dose-Response Relationship, Drug; D004546:Elapid Venoms / Q000633:toxicity; D005230:Fatty Acids, Nonesterified / Q000096:biosynthesis; D008297:Male; D008564:Membrane Potentials / Q000187:drug effects*; D009498:Neurotoxins / Q000633:toxicity*; D010741:Phospholipases A / Q000187:drug effects*; D054467:Phospholipases A2; D051381:Rats; D011919:Rats, Inbred Strains; D000254:Sodium-Potassium-Exchanging ATPase / Q000187:drug effects / Q000378:metabolism*; D013574:Synaptosomes / Q000187:drug effects* / Q000378:metabolism; D013997:Time Factors
D002038:Bungarotoxins; D004546:Elapid Venoms; D005230:Fatty Acids, Nonesterified; D009498:Neurotoxins; C005082:notexin; D010741:Phospholipases A; D054467:Phospholipases A2; D000254:Sodium-Potassium-Exchanging ATPase