PGE2, Ca2+, and cAMP mediate ATP activation of Cl- channels in pigmented ciliary epithelial cells

Purines regulate intraocular pressure. Adenosine activates Cl- channels of nonpigmented ciliary epithelial cells facing the aqueous humor, enhancing secretion. Tamoxifen and ATP synergistically activate Cl- channels of pigmented ciliary epithelial (PE) cells facing the stroma, potentially reducing net secretion. The actions of nucleotides alone on Cl- channel activity of bovine PE cells were studied by electronic cell sorting, patch clamping, and luciferin/luciferase ATP assay. Cl- channels were activated by ATP. UTP, ADP, and UDP, but not by 2-methylthio-ATP, all at 100 muM. UTP triggered ATP release. The second messengers Ca2+, prostaglandin (PG) E-2, and cAMP activated Cl- channels without enhancing effects of 100 mM ATP. Buffering intracellular Ca2+ activity with 1,2-bis(2-aminophenoxy) ethane-N,N,N',N'-tetraacetic acid or blocking PGE(2) formation with indomethacin inhibited ATP-triggered channel activation. The Rp stereoisomer of 8-bromoadenosine 3',5'-cyclic monophosphothioate inhibited protein kinase A activity but mimicked 8-bromoadenosine 3',5'-cyclic monophosphate. We conclude that nucleotides can act at >1 P2Y receptor to trigger a sequential cascade involving Ca2+, PGE(2), and cAMP. cAMP acts directly on Cl- channels of PE cells, increasing stromal release and potentially reducing net aqueous humor formation and intraocular pressure.