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 s ecretion. Tamoxifen and ATP synergistically activate Cl- channels of pigmen ted ciliary epithelial (PE) cells facing the stroma, potentially reducing n et secretion. The actions of nucleotides alone on Cl- channel activity of b ovine PE cells were studied by electronic cell sorting, patch clamping, and luciferin/luciferase ATP assay. Cl- channels were activated by ATP. UTP, A DP, 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 acti vated Cl- channels without enhancing effects of 100 mM ATP. Buffering intra cellular Ca2+ activity with 1,2-bis(2-aminophenoxy) ethane-N,N,N',N'-tetraa cetic acid or blocking PGE(2) formation with indomethacin inhibited ATP-tri ggered channel activation. The Rp stereoisomer of 8-bromoadenosine 3',5'-cy clic 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 s tromal release and potentially reducing net aqueous humor formation and int raocular pressure.