Endothelin stimulation of PLC and PLA2 activity in cultured human ciliary muscle cells

K. White, P. Magnino, Iok-Hou Pang, Thomas Yorio

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Abstract

Purpose: To determine if the effects of endothelin (ET) on PLC and PLA2 activity in cultured human ciliary muscle (HCM) cells are mediated through independent ETA receptor-coupled events. Methods: PLC activity was measured as a changes in [Ca2+]i determined by single cell dynamic fluorescence ratio imaging in cells preloaded with fura-2/AM. PLA2 activity was determined by measuring PGE2 production by RIA. Results: In the HCM cells, endothelin-1 (ET-1) and endothelin-2 (ET-2) increased [Ca2+]i. Endothelin-3 (ET-3) had no effect on [Ca2+]i. BQ610, an ETA receptor subtype-selective antagonist, blocked the ET-1 and ET-2 increase in [Ca2+]i. IRL-1620, an ETB receptor subtype-selective agonist, had no effect on [Ca2+]i and IRL-1038, an ETB receptor subtype-selective antagonist, did not block Ca2+ mobilization. U73122, a phospholipase C (PLC) inhibitor, abolished the ET-1 stimulated calcium mobilization, whereas the phospholipase A2 inhibitor, AACOCF3(at 1μM) had no effect. Manoalide, but not AACOCF3, decreased ET-1 mediated PGE2 production and decreased [Ca2+]i. Conclusions: ET-1 and ET-2 stimulate [Ca2+]i in HCM cells via an ETA receptor subtype. The increase in [Ca2+]i mediated by ET-1 appears to be mediated through the phospholipase C signalling pathway, whereas the effects of ET-1 on PGE2 production appears to be the result of an ETA receptor coupled to PLA2.

Original languageEnglish
JournalInvestigative Ophthalmology and Visual Science
Volume37
Issue number3
StatePublished - 15 Feb 1996

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