This study examined the age-related changes in nitric oxide synthase immunoreactivity (NOS-IR), survival and regeneration of magnocellular neurons in the hypothalamo-neurohypophyseal system (HNS) in rats following hypophysectomy. In adult animal, hypophysectomy induced a significant increase in NOS-IR in the supraoptic (SON), paraventricular nuclei (PVN) and median eminence (ME) by 3 days post-lesion. NOS sustained an increased level until 2 weeks after hypophysectomy and then returned to normal control level. In contrast, at postnatal day 7 (PN7), no obvious increase in NOS-IR was observed in the SON, PVN and ME following the injury compared with age-matched controls. At PN14, the same injury induced an increase in NOS-IR in SON, PVN and ME but the increase was more transient with peak NOS-IR at 3 days and returning to the corresponding control level at 1 week after hypophysectomy. In contrast to a striking age-dependent alteration in NOS-IR in the SON and PVN, hypophysectomy induced substantial degeneration of arginine vasopressin (AVP) and oxytocin (OT) neurons in the SON and PVN in both immature and adult rats and there was no obvious difference in neuronal survival after the same injury among these three groups of different ages by quantitative analysis. Following hypophysectomy, a large number of fibers were observed in the contact zone of the median eminence and the adjacent lumen of the third cerebral ventricle (V3) in adult rats, whereas few fibers could be found in the lumen of the V3 in the immature rats after the same injury. Relationships between NOS induction and magnocellular neuronal survival and regeneration were discussed.