Activation of phospholipase C and guanylyl cyclase by endothelins in human trabecular meshwork cells.
OBJECTIVE
To characterize effects of endothelins on activities of phospholipase C (PLC) and nucleotide cyclases in human trabecular meshwork (TM) cells.


METHODS
Cultured simian virus 40-transformed human TM (HTM-3) or non-transformed (HTM-16) cells were used. Changes in the PLC activity were determined by assaying the production of [3H] inositol phosphates. Accumulation of cyclic GMP or cyclic AMP in cell lysate was measured by radioimmunoassay.


RESULTS
Endothelin-1 (ET-1; 1 microM) stimulated PLC in HTM-16 cells, but Sarafotoxin S6c (SRTX), an ET(B) receptor subtype-selective agonist (1 microM), did not. Similar results were obtained in HTM-3 cells: ET-1, but not ET-3 or SRTX, activated PLC in a dose-dependent manner, with a calculated EC50 of 646 pM. The peptide also stimulated the accumulation of cGMP in a concentration-dependent manner with an EC50 of 37.2 pM. ET-3 or SRTX was not effective except at much higher concentrations. Both the PLC and guanylyl cyclase stimulation induced by ET-1 (10 nM) were completely inhibited by pretreating the cells with BQ-123 (<10 microM), an ET(A) receptor selective antagonist, but not by BQ-788 (10 microM), an ET(B) receptor subtype-specific antagonist. Neither ET-1 nor ET-3 stimulated adenylyl cyclase activity in HTM-3 cells at concentration as high as 1 microM.


CONCLUSIONS
ET-1 activates PLC and guanylyl cyclase in TM cells. Potency profiles of ET receptor agonists and antagonists suggest that the ET(A) receptor subtype is involved in both actions of ET-1. The effects of the ET peptides in TM cells are interesting and could be part of the mechanism of their IOP-lowering effect.
Haque
M S
MS
Alcon Laboratories Inc., Fort Worth, TX 76134-2099, USA.
Pang
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Magnino
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DeSantis
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D002461:Cell Line, Transformed; D002478:Cells, Cultured; D000242:Cyclic AMP / Q000378:metabolism; D006152:Cyclic GMP / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D065128:Endothelin Receptor Antagonists; D019332:Endothelin-1 / Q000494:pharmacology*; D004789:Enzyme Activation / Q000187:drug effects; D006162:Guanylate Cyclase / Q000378:metabolism*; D006801:Humans; D007295:Inositol Phosphates / Q000378:metabolism; D010456:Peptides, Cyclic / Q000494:pharmacology; D011863:Radioimmunoassay; D017466:Receptors, Endothelin / Q000378:metabolism; D014129:Trabecular Meshwork / Q000187:drug effects* / Q000201:enzymology; D010738:Type C Phospholipases / Q000378:metabolism*; D014757:Viper Venoms / Q000494:pharmacology
D065128:Endothelin Receptor Antagonists; D019332:Endothelin-1; D007295:Inositol Phosphates; D010456:Peptides, Cyclic; D017466:Receptors, Endothelin; D014757:Viper Venoms; C057354:sarafotoxins s6; D000242:Cyclic AMP; D010738:Type C Phospholipases; D006162:Guanylate Cyclase; D006152:Cyclic GMP; C072247:cyclo(Trp-Asp-Pro-Val-Leu)