Toshihiko Murayama, Haruko Oda, Asako Watanabe and Yasuyuki Nomura
Department of Pharmacology, Faculty of Pharmaceutical Sciences, Hokkaido University, Sapporo 060, Japan
Abstract: Phospholipase A2 has recently been proposed as the effector enzyme involved in the receptor-mediated release of arachidonic acid (AA). Released AA and its metabolites have been demonstrated to play an important role in the regulation of cell functions. [3H]AA release from prelabeled PC12 cells was stimulated by a Ca ionophore such as ionomycin or A23187. Although ATP and its effective analogs, adenosine 5'-O-(3-thiotrisphosphate) (ATPgammaS), 2-methylthio ATP and 3'-O-(4-benzoyl)benzoyl ATP, did not stimulate [3H]AA release on their own, they did enhance Ca ionophore-stimulated [3H]AA release. The effect of ATP analogs was dose-dependent. ADP, UTP, GTP, ITP, alphabeta-methylene ATP, betagamma-methylene ATP and 8-bromo ATP showed no effect or very limited effect. The effect of ATPalphaS was antagonized by suramin, a putative P2Y receptor antagonist. The effective ATP analogs also increased [Ca2+]i (cytosolic free Ca2+ concentration) via Ca2+ influx. However, the addition of 50 mM KCl or 10 microM bradykinin, which are well-known to increase [Ca2+]i by different pathways, did not stimulate [3H]AA release, either with or without the Ca ionophore. The addition of phorbol 12-myristate 13-acetate, an activator of protein kinase C, showed no effect on [3H]AA release, either with or without the Ca ionophore. These data suggest that 1)ATP increased Ca ionophore-stimulated AA release via a P2Y-like ATP receptor, and that 2)the elevation of [Ca2+]i by ATP does not quantitatively explain the ATP-stimulated AA release in PC12 cells.
Keywords: ATP, Arachidonic acid, Intracellular calcium, PC12 cell