ENDOTHELIN-INDUCED CHANGES OF 2ND-MESSENGERS IN CULTURED HUMAN CILIARY MUSCLE-CELLS

Purpose. To characterize the pharmacology of endothelin-induced change s in phospholipase C (PLC) activity, intracellular calcium concentrati on ([Ca2+](i)), and cyclic adenosine monophosphate (cAMP) in cultured human ciliary muscle (HCM) cells. Methods. Changes in PLC activity of HCM cells were determined by production of [H-3] inositol phosphates. [Ca2+](i) was determined by single-cell dynamic fluorescence ratio ima ging. Radio-immunoassays were used to determine cAMP and prostaglandin E(2) (PGE(2)) concentrations. Results. Endothelin-l (ET-1) stimulated PLC (mean EC(50) = 335 pM) and activated calcium mobilization in HCM cells. These effects were mediated by the endothelin ET(A) receptor su btype because an ET(B) receptor-selective agonist, sarafotoxin S6c, wa s ineffective. Additionally, effects of ET-1 were inhibited by pretrea tment with a selective ET(A) antagonist, BQ610 (mean pK(i) = 9.96 for PLC). ET-1 also stimulated the production of PGE(2) (mean EC(50) = 12. 0 nM) and cAMP (mean EC(50) = 5.2 nM) by these cells. PGE(2) appeared to mediate the stimulatory effect of ET-I on adenylyl cyclase because blockade of ET-l-induced PGE(2) production by 10 mu M indomethacin als o completely blocked the ET-l-activated cAMP production. Conclusions. ET-1 stimulated PLC and increased [Ca2+](i) in HCM cells by the ET(A) receptor subtype. ET-1 also increased cAMP production, an effect likel y mediated by the enhanced production of prostaglandins.