Rapid expression of neuronal and inducible nitric oxide synthases during post-ischemic reperfusion in rat brain

Objective: To determine whether neuronal and inducible nitric oxide synthase (nNOS and iNOS) isoforms are expressed within cortical neurons during early reperfusion after focal cerebral ischemia. Methods: Male spontaneously hypertensive rats underwent occlusion of the left middle cerebral artery for 2 h. Coronal brain sections with normal and ischemic cortex were obtained after 15 min or 1, 6 or 24 h of reperfusion. Immunohistochemical and double-label immunofluorescent techniques were used to confirm cellular identity and localize nNOS and iNOS. Results: Immunoreactive nNOS was identified within isolated neurons in layer V of normal cortex. However, the number of nNOS-immunoreactive neurons in ischemic cortex rose markedly at 15 min and persisted for 24 h (P less than or equal to0.001 at each time point when compared to normal cortex). Cells that were immunoreactive fur nNOS appeared in perivascular clusters within ischemic brain at all sampling times. Immunoreactive iNOS was also expressed within neurons in ischemic cortex, peaking after 15 min of reperfusion (P less than or equal to0.01). Although nNOS-immunoreactive neurons were observed in random numbers within normal tissue throughout reperfusion, iNOS-immunoreactive neurons increased steadily in the same region (P less than or equal to0.05). Conclusions: Ischemic neurons become immunoreactive for both nNOS and iNOS during early reperfusion. Expression of iNOS immunoreactivity in unaffected neurons may reflect transcription of immediate early genes in response to stimulatory neurotransmission from ischemic cortex. (C) 2001 Elsevier Science B.V. All rights reserved.