title: Human choroidal melanocyte signal transduction responses to various pharmacological agents: focus on endothelin receptors.
abstract: OBJECTIVE
  The receptor-coupled signal transduction systems present in isolated human choroidal melanocytes (HCOMs) were investigated.
  
  
  METHODS
  [(3)H]-inositol phosphates ([(3)H]-IPs) generated in the cells were measured by ion-exchange chromatography. cAMP generated in the cells was quantified using an enzyme immunoassay.
  
  
  RESULTS
  Initially, HCOM cells were challenged with a relatively high concentration (e.g., 1 µM-1 mM) of a variety of pharmacological agents in order to determine which functional receptors were present in these cells. Full concentration-response pharmacological studies were subsequently conducted on endothelin receptors. While a number of prostaglandins (PGs) (e.g., PGD(2), PGE(2), PGF(2α), cloprostenol, latanoprost acid, U-46619), histamine, carbachol, bombesin, and arginine-vasopressin were essentially inactive at stimulating the phosphoinositide (PI) hydrolysis response, endothelin-1 (ET-1) potently and efficaciously generated [(3)H]-IPs. Concentration-response studies yielded the following potency (EC(50)) and efficacy (E(max) relative to ET-1) data: ET-1 EC(50) = 3.4 ± 1.4 nM, E(max) = 100%, n = 3; BQ-3020 (ET(B) receptor-selective agonist) EC(50) = 13 ± 4 nM, E(max) = 73 ± 2%, n = 3). The effects of ET-1 on [(3)H]-IPs production were blocked by the ET(B) receptor-selective antagonist, BQ-788 (IC(50) = 10 ± 5 nM, n = 3), while the ET(A) receptor-selective antagonist (BQ-610) was essentially inactive. In the adenylyl cyclase (AC) assay, while isoproterenol (10 µM), ET-1 (1 µM) and PGE(2) (10 µM) stimulated cAMP production, numerous other PGs (e.g., PGD(2), PGF(2α), PGI(2), latanoprost, latanoprost acid, U-46619 and BW245C [all at > 10 µM]) were inactive.
  
  
  CONCLUSIONS
  It is concluded that HCOMs express functionally active ET(B) receptors that mediate the production of [(3)H]-IPs. Additionally, HCOMs generate cAMP in response to ET-1, PGE(2), and isoproterenol. These data may have relevance to the melanogenic activity of HCOM cells.
authors.affiliation: Pharmaceutical Research, Alcon Research, Ltd., Fort Worth, Texas, USA.
authors.forename: Najam A
authors.initials: NA
authors.lastname: Sharif
authors.forename: Julie Y
authors.initials: JY
authors.lastname: Crider
mesh_terms: D000369:Aged, 80 and over; D002478:Cells, Cultured; D002829:Choroid / Q000166:cytology*; D002852:Chromatography, Ion Exchange; D000242:Cyclic AMP / Q000378:metabolism; D065131:Endothelin B Receptor Antagonists; D019332:Endothelin-1 / Q000494:pharmacology; D016232:Endothelins / Q000494:pharmacology; D005260:Female; D006801:Humans; D007124:Immunoenzyme Techniques; D007295:Inositol Phosphates / Q000378:metabolism; D008544:Melanocytes / Q000187:drug effects* / Q000378:metabolism; D010446:Peptide Fragments / Q000494:pharmacology; D011453:Prostaglandins / Q000494:pharmacology; D044023:Receptor, Endothelin B / Q000819:agonists / Q000378:metabolism*; D015398:Signal Transduction / Q000502:physiology*; D014019:Tissue Donors
chemical_list: D065131:Endothelin B Receptor Antagonists; D019332:Endothelin-1; D016232:Endothelins; D007295:Inositol Phosphates; D010446:Peptide Fragments; D011453:Prostaglandins; D044023:Receptor, Endothelin B; C075771:BQ 3020; D000242:Cyclic AMP