Prostaglandins mediate the stimulatory effects of endothelin-1 on cyclic adenosine monophosphate accumulation in ciliary smooth muscle isolated from bovine, cat, and other mammalian species.
OBJECTIVE
To examine the effects and mechanisms of endothelin-1 (ET-1) on cyclic adenosine monophosphate (cAMP) accumulation, inositol 1,4,5-trisphosphate (IP3) production, and contraction in ciliary muscle (CM) isolated from bovine, cat and other mammalian species.


METHODS
Ciliary muscle was incubated in the absence and presence of ET-1 for 5 minutes. Indomethacin (Indo, 2.5 microM) and IBMX (0.1 mM) were added 10 minutes before the addition of the peptide. Cyclic AMP accumulation and prostaglandin E2 (PGE2) release were measured by radioimmunoassay, IP3 production was measured by ion-exchange chromatography, and changes in tension were recorded isometrically.


RESULTS
First, ET-1 (0.1 microM) increased PGE2 release by 58% to 105% and cAMP accumulation by 98% to 393% in CMs isolated from bovine, cat, dog and human, and these effects were blocked completely by Indo (2.5 microM). Unlike any other species, in bovine CM, ET-1 increased IP3 production (EC50 = 17 nM) and contraction (EC50 = 13 nM), and these effects were not inhibited by Indo. Second, kinetic studies revealed that in bovine and cat CMs, ET-1 stimulated cAMP accumulation and PGE2 release in a time- and dose-dependent manner, and these effects were inhibited by Indo in a time- and dose-dependent manner, and PGE2 increased cAMP accumulation in a dose-dependent manner (EC50 = 0.175 microM). The stimulatory effect of ET-1 on cAMP accumulation is mediated through the ETA receptor subtype, because in contrast to ET-1, which is an ETA receptor agonist, ET-3 and Sarafotoxin-S6c, two ETB receptor agonists, had little effect on cAMP accumulation. In addition, BQ 610, an ETA receptor subtype antagonist, inhibited ET-1-induced cAMP accumulation in a dose-dependent manner (IC50s for bovine and cat were 11 and 19.5 nM, respectively). Quinacrine, a phospholipase A2 inhibitor, inhibited ET-1-induced cAMP accumulation in a dose-dependent manner (IC50s for bovine and cat were 22 and 19 microM, respectively). PGE2, but not ET-1, stimulated adenylyl cyclase activity in membranes isolated from bovine and cat CMs.


CONCLUSIONS
In CMs isolated from bovine, cat, dog and human, ET-1-induced cAMP accumulation is mediated through the release of PGs. ET-1 binds to the ETA receptor subtype to activate phospholipase A2 and to release arachidonic acid for PG synthesis. PGs, such as PGE2, may interact with the EP receptor to stimulate adenylyl cyclase. Although ET-1-induced PG release could function to modulate, through cAMP, the responses to muscarinic receptor stimulation, the precise role of these effects in intraocular pressure lowering and accommodation remains to be delineated.
Abdel-Latif
A A
AA
Department of Biochemistry and Molecular Biology, Medical College of Georgia, Augusta 30912-2100, USA.
Yousufzai
S Y
SY
el-Mowafy
A M
AM
Ye
Z
Z
D015056:1-Methyl-3-isobutylxanthine / Q000494:pharmacology; D000262:Adenylyl Cyclases / Q000378:metabolism; D000818:Animals; D016718:Arachidonic Acid / Q000378:metabolism; D002415:Cats; D002417:Cattle; D002924:Ciliary Body / Q000187:drug effects / Q000378:metabolism*; D000242:Cyclic AMP / Q000096:biosynthesis*; D016861:Cyclooxygenase Inhibitors / Q000494:pharmacology; D015232:Dinoprostone / Q000378:metabolism / Q000494:pharmacology; D004285:Dogs; D004305:Dose-Response Relationship, Drug; D065128:Endothelin Receptor Antagonists; D016232:Endothelins / Q000494:pharmacology*; D006801:Humans; D007213:Indomethacin / Q000494:pharmacology; D015544:Inositol 1,4,5-Trisphosphate / Q000096:biosynthesis; D007537:Isometric Contraction / Q000187:drug effects; D009637:Masoprocol / Q000494:pharmacology; D009130:Muscle, Smooth / Q000187:drug effects / Q000378:metabolism*; D010726:Phosphodiesterase Inhibitors / Q000494:pharmacology; D010741:Phospholipases A / Q000378:metabolism; D054467:Phospholipases A2; D011453:Prostaglandins / Q000378:metabolism*; D017466:Receptors, Endothelin / Q000378:metabolism; D013997:Time Factors
D016861:Cyclooxygenase Inhibitors; D065128:Endothelin Receptor Antagonists; D016232:Endothelins; D010726:Phosphodiesterase Inhibitors; D011453:Prostaglandins; D017466:Receptors, Endothelin; D016718:Arachidonic Acid; D009637:Masoprocol; D015544:Inositol 1,4,5-Trisphosphate; D000242:Cyclic AMP; D010741:Phospholipases A; D054467:Phospholipases A2; D000262:Adenylyl Cyclases; D015232:Dinoprostone; D015056:1-Methyl-3-isobutylxanthine; D007213:Indomethacin