Upregulation of iNOS expression and phosphorylation of eIF-2 alpha are paralleled by suppression of protein synthesis in rat hypothalamus in a closedhead trauma model

When the inducible form of nitric oxide synthase (iNOS) is expressed after challenge to the nervous system, it results in abnormally high concentratio ns of nitric oxide (NO). Under such conditions, NO could phosphorylate the eukaryotic translation initiation factor (eIF)-2(alpha), thus suppressing p rotein synthesis in neurons that play a role in endocrine and autonomic fun ctions. Using the Marmarou model of traumatic brain injury (TBI), we observ ed a rapid increase (at 4 h after TBI) of iNOS mRNA in magno- and parvocell ular supraoptic and paraventricular neurons, declining gradually by similar to 30% at 24 h and by similar to 80% at 48 h. Western analysis indicated a trend towards increased iNOS protein synthesis at 4 h, which peaked at 8 h , and tended to decrease at the later time points. At the same time points, we detected immunocytochemically the phosphorylated form of eIF-2 alpha (e IF-2 alpha [P]) as cytoplasmic and more often as nuclear labeling. The inci dence of double-labeled [iNOS and eIF-2 alpha (P)] neuronal profiles, parti cularly at 24 h and 48 h after TBI, was high. De novo protein synthesis ass essed quantitatively after infusion of S-35 methionine/cysteine was reduced by similar to 20% at 4 h, remained depressed at 24 h, and did not return t o control levels up to 48 h following the trauma. The results suggest that MOS may trigger phosphorylation of eIF-2 alpha, which in turn interferes wi th protein synthesis at the translational (ribosomal complex) and transcrip tional (chromatin) levels. The depression in protein synthesis may include downregulation of iNOS itself, which could be an autoregulatory inhibitory feedback mechanism for NO synthesis. Excessive amounts of NO may also parti cipate in dysfunction of hypothalamic circuits that underlie endocrine and autonomic alterations following TBI.