PATHWAYS SIGNALING THE REGULATORY VOLUME DECREASE OF CULTURED NONPIGMENTED CILIARY EPITHELIAL-CELLS

Purpose. The authors identify the signaling pathways for the regulator y volume decrease (RVD) of nonpigmented ciliary epithelial (NPE) cells . The RVD is a regulatory response triggered by swelling and reflectin g KCl release by NPE cells. Methods. The cell Volumes of human nonpigm ented ciliary epithelial cells were measured in suspension by electron ic cell sorting. Measurements were conducted in test solutions of cons tant ionic strength, but osmolality was varied by sucrose. Results. Cy clic AMP (cAMP), forskolin, PGE(2), the PKC-inhibitor staurosporine, a nd increasing cytoplasmic Ca2+ activity with thapsigargin all enhanced the RVD. Leukotrienes A(4), D-4, E(4), and the protein phosphatase in hibitor okadaic acid had no detectable effect under the current experi mental conditions. The cyclooxygenase inhibitor indomethacin, the epox ygenase inhibitors ketoconazole and SKF 525A, and the PKC activator Di C(8) all downregulated the RVD. The addition of the cation ionophore, gramicidin, increased the RVD. In the presence of gramicidin, cAMP, PG E(2), and indomethacin did not affect the RVD, but ketoconazole, DiC(8 ), and the calcium-calmodulin blocker trifluoroperazine still inhibite d-and staurosporine still enhanced-the RVD. Many of these observations are strikingly different from results reported with other cells. Anis osmotic swelling did not increase intracellular cAMP concentration. Co nclusions. The pathways signaling the regulatory responses to swelling are unique for each cell type. The authors propose that hypotonic swe lling of NPE cells stimulates arachidonic acid turnover, triggering PG E(2)-mediated upregulation of K+ channels and epoxide-mediated upregul ation of Cl- channels. Swelling may also reduce endogenous PKC activit y, further upregulating Cl- channels. Calcium-calmodulin plays a permi ssive role in upregulating the Cl- channels.