Functional identification of phosphodiesterase activity in human trabecular meshwork cells.
The phosphodiesterases (PDE) activity in human trabecular meshwork cells (HTM-3) was investigated in this study in order to better understand the signal transduction pathways in the conventional outflow tract of the eye. Agonists (isoproterenol or nitroprusside) were used to stimulate adenylyl cyclase and guanylyl cyclase, respectively, in the absence and presence of nonselective IBMX or PDE5 specific inhibitors E4021 (1). The subcellular distribution of cAMP and cGMP PDEs was determined directly by PDE enzyme assays using HTM-3 cells. Levels of cyclic nucleotides were measured in the same cells by radioimmunoassay (RIA). Isoproterenol alone elevated cAMP levels, and this response was enhanced by IBMX. Nitroprusside alone caused no increase in basal cGMP levels but, in the presence of E4021, nitroprusside produced significant, dose-related elevation of cGMP levels. Subcellular distribution experiments indicated that the greatest activity for PDEs resided in the supernatant fraction. In conclusion, HTM-3 cells contain PDEs that degrade both cyclic nucleotides. The PDE activities reside predominantly in the supernatant, but the PDE activity for degrading cGMP is more pronounced. Moreover, results with E4021 suggest that PDE5 activity could play a critical role in modulating cGMP-related activity in the trabecular meshwork.
Zhou
L
L
Department of Pharmacology and Toxicology, Morehouse School of Medicine, Atlanta, Georgia, USA. lzhou3@emory.edu
Thompson
W J
WJ
Potter
D E
DE
D015056:1-Methyl-3-isobutylxanthine / Q000494:pharmacology; D015105:3',5'-Cyclic-AMP Phosphodiesterases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D015106:3',5'-Cyclic-GMP Phosphodiesterases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D000262:Adenylyl Cyclases / Q000378:metabolism; D002478:Cells, Cultured; D000242:Cyclic AMP / Q000378:metabolism; D006152:Cyclic GMP / Q000378:metabolism; D006162:Guanylate Cyclase / Q000378:metabolism; D006801:Humans; D007545:Isoproterenol / Q000494:pharmacology; D009599:Nitroprusside / Q000494:pharmacology; D010880:Piperidines / Q000494:pharmacology; D011799:Quinazolines / Q000494:pharmacology; D015398:Signal Transduction / Q000502:physiology*; D014129:Trabecular Meshwork / Q000166:cytology / Q000201:enzymology*
C091517:E 4021; D010880:Piperidines; D011799:Quinazolines; D009599:Nitroprusside; D000242:Cyclic AMP; D015105:3',5'-Cyclic-AMP Phosphodiesterases; D015106:3',5'-Cyclic-GMP Phosphodiesterases; D000262:Adenylyl Cyclases; D006162:Guanylate Cyclase; D006152:Cyclic GMP; D007545:Isoproterenol; D015056:1-Methyl-3-isobutylxanthine