Vol. 287, Issue 3, 889-896, December 1998

Pardaxin, a New Pharmacological Tool to Stimulate the Arachidonic Acid Cascade in PC12 Cells¹

Saleh Abu-Raya, Eugenia Bloch-Shilderman, Esther Shohami, Victoria Trembovler, Yechiel Shai², Joseph Weidenfeld³, Saul Yedgar⁴, Yehuda Gutman and Philip Lazarovici

Department of Pharmacology and Experimental Therapeutics, School of Pharmacy (S.A.R., E.B.S., V.T., E.S., Y.G., P.L.), Faculty of Medicine, The Hebrew University, Jerusalem, Israel

The effect of Pardaxin, a neurotoxin that induces neurotransmitter release from neurons, on the arachidonic acid (AA) cascade was studied in PC12 cells. Both native and the synthetic Pardaxin selectively stimulated phospholipase A₂ (PLA₂) activity (measured by [³H]AA release) in the presence as well as in the absence of extracellular calcium. Pardaxin-stimulated PLA₂ activity was also evident in the increased formation of lysophosphatidylcholine. Pardaxin analogs, lacking the -helical structure that is essential for insertion into the plasma membrane, were ineffective in stimulating the AA cascade in PC12 cells. Pardaxin stimulation of PLA₂ was markedly inhibited by the nonselective PLA₂ inhibitors bromophenacyl bromide and mepacrine, by methyl arachidonyl fluorophosphonate, a dual inhibitor of calcium-dependent cytosolic PLA₂ and the calcium-independent PLA₂ and by bromoenol lactone[(E)-6-(bromoethylene)tetrahydro-3-(1-naphthalenyl-2H-pyran-2-one], a highly specific inhibitor of calcium-independent PLA₂. After Pardaxin treatment, there was increased release of AA metabolites produced by the cyclooxygenase pathway as expressed in an 8-fold increase of PGE₂ release. The release of other eicosanoids, such as 6-keto-PGF₁ and thromboxane B₂, was also augmented. Pardaxin-induced PGE₂ release was observed in calcium-free medium and in the absence of any increase in cytosolic calcium. Dexamethasone partially inhibited Pardaxin-induced PGE₂ release. This effect was reversed by the type II corticosteroid receptor antagonist RU-38486. Our results indicate that Pardaxin stimulates release of AA and eicosanoids, independently of calcium, and suggest that calcium-independent PLA₂ plays an important role in Pardaxin stimulation of the AA cascade.