SPONTANEOUS EXPRESSION OF INDUCIBLE NITRIC-OXIDE SYNTHASE IN THE HYPOTHALAMUS AND OTHER BRAIN-REGIONS OF AGING RATS

Our laboratory has demonstrated that aging in Brown-Norway rats is ass ociated with decreased LH pulse amplitude and reduced GnRH and LH resp onsiveness to excitatory amino acids (EAA), presumably through the NMD A receptor (NMDAR). Nitric oxide (NO) is a neurotransmitter postulated to be involved in hypothalamic synaptic events required for normal Gn RH regulation through the activation of neuronal nitric oxide synthase (nNOS). Paradoxically, excessive stimulation of nNOS by NMDAR or the expression of inducible nitric oxide synthase (iNOS) can lead to supra physiological levels of NO acting as effector of apoptosis with result ant decreased regional neuronal function. The aims of this study mere to determine: 1) whether aging in the preoptic area/medial basal hypot halamus is associated with altered NO synthesis; 2) the possible roles of the NMDAR/nNOS cascade and iNOS in this process; and 3) whether al terations in the levels of NOS isoforms are specific to this region of the brain. Brown Norway male rats (N = 5) at ages 1 (immature), 3 (ad ult), and 24 told) months, were used for measuring NMDARs in hypothala mic membranes by the binding of a (3H)-NMDAR ligand. Another series of the same age groups of rats (N = 9) were used to determine by Western blot the contents of NMDAR, nNOS, and iNOS in the hypothalamus, and o nly iNOS in the frontal and parietal cortex, and cerebellum. NOS activ ity was measured in the hypothalamus by the arginine/citrulline assay. A significant decrease of NMDA analog binding was found in the hypoth alamus from old rats as compared with adult (-66%) and immature animal s (-57%), accompanied by a reduction in NMDAR content (-34% and -46%, respectively). NOS activity in the hypothalamus was 67% and 100% highe r in old rats as compared with the other two groups, although no signi ficant differences were observed in nNOS content. However, hypothalami c iNOS increased 3.8- and 7.6-fold in old rats, as compared with adult and immature, respectively. This increase in hypothalamic iNOS was pa ralleled by a rise of iNOS in other brain regions of old rats as compa red respectively to adult and immature animals: 3.9- and 12.8-fold, in the frontal cortex; 2.8- and 2.5-fold, in the parietal cortex; and 3. 1- and 4.8-fold, in the cerebellum. These results show that aging in t his rat model is associated with high NO synthesis in the hypothalamus and other regions of the brain, which is independent of the NMDAR/nNO S cascade. We speculate that increased brain levels of iNOS may lead t o neurotoxicity, which may be involved in GnRH impaired pulsatile secr etion, as well as acting as a possible inducer of age associated neuro nal loss in cognitive related brain areas.