ENDOTHELIN-1 STIMULATES THE RELEASE OF ARACHIDONIC-ACID AND PROSTAGLANDINS IN CULTURED HUMAN CILIARY MUSCLE-CELLS - ACTIVATION OF PHOSPHOLIPASE A(2)

In the present study we have examined the effects and mechanisms of en dothelin-1 (ET-1) on arachidonic acid (AA) release and prostaglandin ( PG) synthesis in human ciliary muscle (HCM) cells. ET-1 stimulated AA release in a time (t(1/2) = 1.5 min) and concentration-dependent (EC50 = 5 nM) manner, which is primarily mediated through the ETA receptor subtype. The AA liberated by ET-1 appears to derive mainly from the ph osphoinositides and phosphatidylcholine, Our data show that phospholip ase A, (PLA,), but not phospholipase C (PLC), plays an important role in ET-1-induced AA release. This conclusion is supported by the follow ing findings: (1) ET-1-evoked AA release was inhibited by the PLA, inh ibitors dexamethasone, mepacrine and manoalide in a concentration-depe ndent manner. Conversion of AA into PGE(2) was inhibited by the cycloo xygenase inhibitors in the following order: Indomethacin > naproxen > ibuprofen > NS-398 > aspirin. (2) The phorbol ester, PDBu, an activato r of protein kinase C, potentiated ET-1 induced AA release by 39%, but inhibited that of inositol phosphates formation by 62%. (3) Pretreatm ent of the labeled cells with isoproterenol lowered ET-1-induced inosi tol phosphates production, but had no effect on AA release. (4) U71322 , a PLC inhibitor, inhibited ET-l-induced inositol phosphates producti on, but had no effect on that of AA release. (5) Pretreatment of the c ells with pertussis toxin (0.1 mu g ml(-1)) attenuated the stimulatory effects of ET-1 on AA release and PGE, formation. These data demonstr ate that ET-1 is a potent agonist for AA release and PG synthesis in H CM cells, and that PLA(2), but not PLC, plays an important role in ET- 1-induced AA release and PG synthesis. In ciliary muscle, AA and its m etabolites play important roles in intracellular signalling, modulatio n of physiological processes, and regulation of intraocular pressure. (C) 1997 Academic Press Limited.