IMMUNOSUPPRESSANT FK506 ENHANCES PHOSPHORYLATION OF NITRIC-OXIDE SYNTHASE AND PROTECTS AGAINST GLUTAMATE NEUROTOXICITY

Immunosuppressants FK506 and cyclosporin A inhibit neurotoxicity of N- methyl-D-aspartate in primary cortical cultures, while having no effec t on quisqualate- and kainate-mediated neurotoxicity. Rapamycin comple tely reverses the neuroprotective effect of FK506. Both FK506 and cycl osporin A inhibit NMDA-elicited/nitric oxide-mediated increases in cGM P levels in cortical cultures. FK506 has no effect on sodium nitroprus side-induced increases in cGMP. In a stably transfected human kidney 2 93 cell line overexpressing the gene encoding nitric oxide synthase [L -arginine, NADPH:oxygen oxidoreductase (nitric oxide-forming), EC 1.14 .13.39], FK506 inhibits the calcium ionophore A23187, stimulated incre ases in nitrite (a breakdown product of nitric oxide), and potentiates phorbol ester-mediated inhibition of nitrite formation. FK506-mediate d inhibition of nitric oxide formation is completely reversed by rapam ycin. Calcineurin dephosphorylates protein kinase C-mediated phosphory lation of nitric oxide synthase. FK506 prevents the calcineurin-mediat ed dephosphorylation of nitric oxide synthase and thereby diminishes t he enzyme's catalytic activity. These data establish nitric oxide synt hase as a calcineurin substrate. Nitric oxide synthase catalytic activ ity is regulated by the phosphorylation state of the enzyme. Enhanced phosphorylation of nitric oxide synthase diminishes catalytic activity , and dephosphorylation (through activation of calcineurin) enhances c atalytic activity. The neuroprotective effect of FK506 and cyclosporin A presumably involves the inhibition of calcineurin, preventing the d ephosphorylation of nitric oxide synthase and its subsequent activatio n.