COMPARATIVE-STUDIES ON PROSTANOID RECEPTORS IN HUMAN NONPIGMENTED CILIARY EPITHELIAL AND MOUSE FIBROBLAST CELL-LINES

To examine the expression of functional prostanoid receptors in the hu man non-pigmented ciliary epithelial (ODMC1-2) and mouse fibroblast ce ll lines (NIH 3T3) we have measured the generation of the second messe ngers, cyclic AMP, inositol phosphates and the mobilization of intrace llular calcium ([Ca2+](i)) following stimulation by prostaglandin rece ptor agonists. The amount of cyclic AMP generated was measured by a pr otein binding method. Radiolabeled inositol phosphates were separated using ion exchange columns and quantified by counting the radioactivit y. For intracellular calcium measurements, Fura 2-AM loaded cells were stimulated by PG receptor agonists and the calcium activated fluoresc ence was measured in a spectrofluorometer. In the ODMC1-2 cell line, t he formation of cyclic AMP was stimulated by prostaglandin E(2), butap rost and 11-deoxy-prostaglandin E(1). The stimulation of cyclic AMP pr oduction by prostaglandin E(2) was partially inhibited by the EP(4) re ceptor antagonist AH23848. Prostaglandin E(2) and 11-deoxy-prostagland ir E(1) stimulated the formation of cyclic AMP in NIH 3T3 cells. In OD MC1-2 cells, total inositol phosphate turnover was not increased by 17 -phenyl-trinor-prostaglandin F-2 alpha, 17-phenyl-trinor-prostaglandin E(2) or 11-deoxy-prostaglandin E(1). In contrast, all these agonists, with the exception of 11-deoxy-prostaglandin E(1), significantly incr eased total inositol phosphates in NIH 3T3 cells. In the NIH 3T3 cell line, only prostaglandin F-2 alpha and 17-phenyl-trinor-prostaglandin F-2 alpha increased [Ca2+](i) in a dose-dependent manner; in ODMC1-2 c ells, neither these agonists nor 17-phenyl-trinor-prostaglandin E(2) i ncreased [Ca2+](i). The present studies suggest that in ODMC1-2 cells, EP(2) and EP(4) receptors but neither EP(1) nor FP receptors are expr essed; these studies also imply, NIH 3T3 cells express EP(2) and FP re ceptors, while EP(1) receptors appear to be absent in this cell line.