Neuronal nitric oxide synthase modulates basal catecholamine secretion in bovine chromaffin cells.
The role of endogenously produced nitric oxide (NO) in the regulation of basal catecholamine (CA) secretion was studied in chromaffin cells. Treatment of chromaffin cells with nitric oxide synthase (NOS) inhibitors produced a dose-dependent increase in basal catecholamine secretion, which paralleled their ability to inhibit NOS activity. This inhibitory profile was similar to that found in neurons, suggesting the constitutive expression of neuronal NOS (nNOS) in these cells, which was confirmed by Western blot analysis. A study of the kinetics and pharmacology of nNOS activity expressed in chromaffin cells in culture indicated that NOS activity is calcium-dependent, increases with time, and is highly dependent on both intracellular concentrations of L-arginine (K(m) approximately 4 microM, V(max) = 908 +/- 60 pmol/hr x 10(6) cells) and transport of L-arginine into the cells (exhibiting two affinity constants of k(1) = 3.2 +/- 0.3 microM and k(2) = 126 +/- 5.5 microM). The effects of NOS inhibitors on CA secretion were mediated by the L-arginine-NO-cGMP pathway, insofar as exogenous L-arginine was able to partially block the increase in CA secretion evoked by them, and 1H-[1,2,4]oxadiazolo[4,3-a]quinoxaline-1-one (ODQ), a specific inhibitor of guanylate cyclase, and zaprinast, an inhibitor of the cGMP phosphodiesterase, were able to increase and inhibit, respectively, basal CA secretion in a dose-dependent manner. These results suggest that chromaffin cells exhibit a tonic production of NO by nNOS that keeps the basal CA secretion at low levels, and this could be necessary for maintaining a normotensive state.
Department of Biochemistry and Molecular Biology, Faculty of Pharmacy, Complutense University of Madrid, Spain.
S
S
Vicente
M P
MP
González
M J
MJ
Oset-Gasque
D015106:3',5'-Cyclic-GMP Phosphodiesterases / Q000037:antagonists & inhibitors*; D000818:Animals; D001120:Arginine / Q000494:pharmacology; D015153:Blotting, Western; D002395:Catecholamines / Q000378:metabolism*; D002417:Cattle; D018929:Cell Culture Techniques; D019439:Chromaffin Cells / Q000201:enzymology* / Q000378:metabolism; D002956:Citrulline / Q000031:analogs & derivatives* / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D006162:Guanylate Cyclase / Q000037:antagonists & inhibitors / Q000378:metabolism; D007527:Isoenzymes; D009419:Nerve Tissue Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors* / Q000378:metabolism*; D052248:Nitric Oxide Synthase Type I; D010726:Phosphodiesterase Inhibitors / Q000494:pharmacology; D011688:Purinones / Q000494:pharmacology; D013890:Thiourea / Q000031:analogs & derivatives* / Q000494:pharmacology; D013997:Time Factors
D002395:Catecholamines; D007527:Isoenzymes; D009419:Nerve Tissue Proteins; D010726:Phosphodiesterase Inhibitors; D011688:Purinones; C086715:thiocitrulline; D002956:Citrulline; D001120:Arginine; D019001:Nitric Oxide Synthase; D052248:Nitric Oxide Synthase Type I; D015106:3',5'-Cyclic-GMP Phosphodiesterases; D006162:Guanylate Cyclase; C011145:zaprinast; D013890:Thiourea