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Investigative Ophthalmology & Visual Science, Vol 34, 431-437, Copyright © 1993 by Association for Research in Vision and Ophthalmology

ARTICLES AND REPORTS

Peptides stimulate phosphoinositide hydrolysis in human retinal pigment epithelium

EL Feldman and AE Randolph
Department of Neurology, University of Michigan, Ann Arbor.

PURPOSE. This study examined the effects of peptides on inositol lipid turnover and intracellular calcium (Ca2+) levels in cultured human retinal pigment epithelium (RPE). METHODS. Cultured human RPE were stimulated with bradykinin, arginine vasopressin (AVP), or bombesin. Accumulation of 3H-inositol phosphates in the presence of 10 mmol/l lithium reflected phosphoinositide (PPI) hydrolysis. Peptide-coupled changes in intracellular Ca2+ levels were determined by fura-2 fluorescence. RESULTS. Bradykinin increased PPI hydrolysis to greater than 300% of basal with an EC50 of 300 nM. Bradykinin-coupled PPI turnover was linear up to 60 min and was blocked by > 90% by the B2 antagonist [Thi5,8,D-Phe7]-bradykinin. AVP stimulated PPI turnover in a linear manner by 260% with an EC50 of 800 nM. The V1 receptor antagonist [beta-mercapto-beta,beta-cyclopentamethylenepropionyl1,-O-Me- Tyr2, A rg8]- vasopressin completely inhibited AVP-induced PPI hydrolysis. Bombesin enhanced PPI hydrolysis by 185% with an EC50 of 1 nM. Stimulation was linear up to 60 min and was blocked by > 90% by the bombesin antagonist [Leu13-psi(CH2NH)-Leu14] bombesin. In fura-2-loaded human RPE cells, the resting intracellular Ca2+ concentration was 105 +/- 32 nM (n = 29). Bradykinin increased peak intracellular Ca2+ to 764 +/- 71 nM, AVP to 310 +/- 42 nM, and bombesin to 234 +/- 20.4 nM. CONCLUSIONS. Peptide-stimulated inositol lipid turnover is coupled to cytosolic Ca2+ flux in cultured human RPE.




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Copyright © 1993 by the Association for Research in Vision and Ophthalmology