Nitric oxide synthase induction by ouabain in vascular smooth muscle cellsfrom normotensive and hypertensive rats

Objective To investigate the effect of ouabain on inducible nitric oxide sy nthase (iNOS) activity and expression in cytokine-stimulated vascular smoot h muscle cells (VSMC) from normotensive Wistar-Kyoto (WKY) rats and spontan eously hypertensive rats (SHR), Methods VSMC were treated for 24 h and afterwards, nitric oxide (NO) releas e was determined by the production of nitrite, a stable metabolite of NO. A ctivity of iNOS was measured by the conversion of [H-3]-L-arginine to [H-3] -L-citrulline and iNOS protein expression by Western blotting. Results Ouabain (0.01-1 mmol/l) further enhanced interleukin-1 beta (II-1 b eta)-induced nitrite production by WKY and SHR VSMC, although a more pronou nced effect was observed in SHR cells (maximum response 52.1 +/- 5.2 and 71 .2 +/- 6.4% of II-1 beta effect in WKY and SHR cells, respectively). Such r esponse on NO release was mimicked by the calcium ionophore A 23187 (0.01-1 mu mol/l) and abolished by the voltage-operated calcium channels (VOCC) ni fedipine (0.1 mu mol/l). Expression of iNOS showed that ouabain increased t he synthesis of the enzyme in WKY and SHR VSMC stimulated with II-1 beta, a nd this effect was higher in SHR cells. The increased iNOS expression was s ignificantly reduced by nifedipine, Conclusions Ouabain stimulation of iNOS expression and activity in II-1 bet a-stimulated VSMCs from WKY rats and SHR seems to be related to increased i ntracellular calcium influx through VOCC. The more pronounced effect observ ed in SHR VSMC could be explained by an altered calcium entry in the hypert ensive strain. J Hypertens 2000, 18:877-884 (C) Lippincott Williams & Wilki ns.