NOVEL PARACRINE SIGNALING MECHANISM IN THE OCULAR CILIARY EPITHELIUM

The ciliary body contains an epithelial bilayer consisting of an outer pigmented cell layer (PE) and an inner nonpigmented cell layer (NPE) responsible for aqueous humor secretion. Secretion may be mediated in part by cytosolic Ca2+ concentration ([Ca2+](i)), but whether or how t he two Layers could coordinate their Ca2+ signals to regulate secretio n is unclear. To investigate interactions between PE and NPE, we exami ned [Ca2+](i) signaling in isolated intact ciliary epithelial bilayers using confocal microscopy, Phenylephrine selectively increased [Ca2+] (i) in PE and acetylcholine increased [Ca2+](i) in NPE, but epinephrin e increased [Ca2+](i) in both layers, This increase spread from PE to NPE, and [Ca2+](i) signaling across the bilayer remained coordinated d uring [Ca2+](i) oscillations. All epinephrine-induced [Ca2+](i) signal ing was blocked by the alpha(1)-adrenergic antagonist prazosin, wherea s signaling in the NPE but not PE was blocked by the beta-adrenergic a ntagonist propranolol, the gap junction blockers octanol and 18 alpha- glycyrrhetinic acid, or the A kinase inhibitor R-p diastereomer of ade nosine 3',5'-cyclic monophosphothioate. The beta-adrenergic agonist is oproterenol failed to increase Ca2+ by itself, but isoproterenol plus phenylephrineinduced [Ca2+](i) signals across the bilayer similar to t hose induced by epinephrine, Finally, isoproterenol increased cell-to- cell spread of lucifer yellow via gap junctions, whereas cell-to-cell spread of [Ca2+](i) signals could be induced by photorelease of caged inositol 1,4,5-trisphosphate, Thus, calcium signals are coordinated in the epithelial bilayer so that adrenergic stimulation can increase [C a2+](i) in NPE, but only if NPE are primed by activation of endogenous adenylyl cyclase, whereupon they receive stimulation from adjacent PE via gap junctions, This novel interplay between endocrine and paracri ne pathways may coordinate [Ca2+](i) signaling across the ciliary epit helial bilayer.