EFFECTS OF MAGNESIUM ON NITRIC-OXIDE SYNTHASE ACTIVITY IN ENDOTHELIAL-CELLS

Magnesium modulates endothelium-dependent vasodilation in intact blood vessels. Therefore, the effects of magnesium on nitric oxide (NO) rel ease by isolated endothelial cells and nitric oxide synthase (NOS) act ivity in endothelial cell homogenates were studied. Unstimulated and b radykinin-stimulated NO release by porcine aortic endothelial cell (PA EC) monolayers were unaffected by 30 min of exposure to magnesium conc entrations varying from 0.010 to 10.0 mM. In contrast, when A-23187-st imulated cells were exposed to 0.01, 3.16, and 10.0 mM MgCl2, NO relea se was decreased by 11.3 +/- 1.8, 11.7 +/- 3.0, and 20.3 +/- 7.2%, res pectively, compared with cells exposed to 1.0 mM MgCl2 (P < 0.01). The se data suggested that a change in the intracellular magnesium concent ration had an effect on NO release, in contrast to a change in the ext racellular concentration, which did not have an effect. To further ass ess this possibility, crude NOS extracts were prepared from PAEC and e xposed to MgCl2. NOS activity was measured via the conversion of L-[H- 3]arginine to L-[H-3]citrulline. Increasing the concentration of MgCl2 by 1.0, 3.16, and 10.0 mM caused a 16.0 +/- 6.8, 17.1 +/- 1.7, and 38 .6 +/- 5.3% decrease in citrulline formation, respectively (P < 0.05), suggesting a direct inhibition of NOS by MgCl2. No significant differ ence in the degree of inhibition of NOS activity was found between MgS O4 and MgCl2, thus ruling out a nonspecific chloride effect. In additi on, increasing the concentration of NaCl to 15 mM had no effect on NOS activity, ruling out a nonspecific osmotic effect [101.6 +/- 10.5% of control activity, P = not significant (NS)]. Finally, in cell monolay ers exposed to A-23187, 3.16 mM CaCl2 overcame the inhibition of NO re lease by 3.16 mM MgCl2 (100.7 +/- 7.5% of NO released by cells exposed to 1.0 and 1.87 mM CaCl2, P = NS), suggesting that magnesium antagoni zes calcium-dependent NO release by endothelial cells. In conclusion, 1) extracellular magnesium does not acutely affect NO release by PAEC, 2) magnesium inhibits PAEC NOS activity in a dose-dependent manner, 3 ) the inhibitory effect of MgCl2 is specific to magnesium and not due to an effect of either increased osmotic strength or chloride anion, 4 ) the inhibitory effect of magnesium is probably due to competitive an tagonism of intracellular calcium, and 5) intracellular magnesium conc entration may be an important regulator of NOS activity.