Central NOS inhibition differentially affects vasopressin gene expression in hypothalamic nuclei in septic rats.

Our aim was to investigate the effect of central NOS inhibition on hypothalamic arginine vasopressin (AVP) gene expression, hormone release and on the cardiovascular response during experimental sepsis. Male Wistar rats were intracerebroventricularly injected with the non-selective NO synthase (NOS) inhibitor (L-NAME) or aminoguanidine, a selective inhibitor of the inducible isoform (iNOS). After 30 min, sepsis was induced by cecal ligation and puncture (CLP) causing an increase in heart rate (HR), as well as a reduction in median arterial pressure (MAP) and AVP expression ratio (AVP(R)), mainly in the supraoptic nucleus. AVP plasma levels (AVPp) increased in the early but not in the late phase of sepsis. L-NAME pretreatment increased MAP but did not change HR. It also resulted in an increase in AVPp at all time points, except 24h, when it returned to basal levels. AVP(R), however remained reduced in both nuclei. Aminoguanidine pretreatment resulted in increased MAP in the early phase and higher AVP(R) in the supraoptic, but not in the paraventricular nucleus, while AVPp remained elevated at all time points. We suggest that increased central NO production, mainly inducible NOS-derived, reduces AVP gene expression differentially in supraoptic and paraventricular nuclei, and that this may contribute to low AVP plasma levels and hypotension in the late phase of sepsis.

Differential expression of nNOS mRNA and protein in the nucleus tractus solitarii of young and aged Wistar-Kyoto and spontaneously hypertensive rats.

OBJECTIVE: To examine neuronal nitric oxide synthase (nNOS) mRNA and protein in the nucleus tractus solitarii (NTS) and paraventricular hypothalamic nucleus (PVN) of Wistar-Kyoto (WKY) and spontaneously hypertensive rats (SHR) during their life span. METHODS: By means of in situ hybridization and immunohistochemistry, we evaluated nNOS mRNA and protein in the NTS and PVN of 15-day- and 1-, 2-, 4-, 8- and 12-month-old SHR and WKY rats. RESULTS: Two patterns of nNOS expression were observed in two subnuclei of the NTS: medial (NTSm) and central (NTSce). NTSce of the SHR exhibited higher nNOS mRNA and protein expression in all ages analyzed when compared to the age-matched WKY. Increased amounts of nNOS mRNA and protein were seen in the NTSm only during the early life of SHR (15 days to 4 months) when compared to WKY, suggesting a special role of this circuitry before the establishment of hypertension. No changes were seen in nNOS mRNA and protein expression in the PVN of the SHR in comparison to the WKY in all periods. nNOS analysis during the life span showed either a decrease or no change in nNOS mRNA expression in NTS or PVN associated with increased nNOS protein at some analyzed periods, suggesting the differential regulation of nNOS mRNA and protein during aging. CONCLUSIONS: Data suggest that different NTS subnuclei exhibit nNOS changes in different phases of the life of SHR and this might be important during the development of hypertension in these animals.

Neuronal nitric oxide synthase inhibition differentially affects oxytocin and vasopressin secretion in salt loaded rats.

Nitric oxide, an endogenous gas produced by nitric oxide synthase (NOS), has been described as a neuromodulator of hormone secretion, including the neurohypophysial peptides oxytocin (OT) and vasopressin (AVP), hormones involved in the sodium and water homeostasis. The presence of NOS in the hypothalamic nuclei as well as in the circumventricular organs suggests a nitrergic regulation of OT and AVP secretion. Thus, the aim of this study was to evaluate the effect of 7-nitroindazole (7-NI), a selective inhibitor of neuronal NOS, in the plasma OT and AVP levels in rats submitted to a short and long-term salt loading. We also evaluated the NOS activity in the supraoptic (SON) and paraventricular (PVN) hypothalamic nuclei. Our data showed an increase of plasma OT and AVP levels in both short and long-term salt loading. The augment of plasma OT and AVP levels was accompanied by an increase of NOS activity in the SON and PVN. The injection of 7-NI potentiated the increase of plasma OT induced by salt loading, but inhibited the increase of plasma AVP in the same experimental conditions. These results indicate that, under short and prolonged osmotic stimulation, nitric oxide may differentially control the neurohypophysial secretion.

Hypothalamic nuclei nitric oxide synthase expression in rats malnourished during early lactation period.

In humans and other animals, it has been shown that protein malnutrition during the prenatal period leads to permanent changes, which in adulthood may cause chronic diseases. Molecules involved in the control of energy metabolism could be targets to alterations caused by nutritional status. Some hypothalamic nuclei as the paraventricular (PVN), ventro-medial and arcuate are related to energy metabolism regulation. Orexigenic and anorexigenic molecules are involved in this regulation. Some studies have showed that these nuclei present nitric oxide synthase (NOS) and that it is increased in obese rats. Recently it had been shown that rats malnourished during the lactation period presented metabolic alterations that persist in adulthood. The aim of this work was to study the expression of NOS in hypothalamic nuclei of rats submitted to malnutrition during the early lactation period. Rats from post-natal day (P10) to P90 were used. Control dams were fed with regular chow pellets and diet dams were fed with protein-free chow pellets during the first 10 days of lactation. NADPH-diaphorase or immunostaining techniques were used to access NOS expression in hypothalamic nuclei. Our results show a delay in NOS expression in the PVN and VMH of malnourished rats. It may affect the development of the hypothalamic circuitry, leading to a metabolic imprinting.

Deoxycorticosterone stimulates the activity of nicotinamide adenine dinucleotide phosphate-diaphorase/nitric oxide synthase immunoreactivity in hypothalamic nuclei of rats.

Mineralocorticoids (MC) play an important role in development of salt appetite. Part of this effect involves the hypothalamic paraventricular (PVN) and supraoptic (SON) nuclei, in which MC treatment increases arginine vasopressin (AVP) synthesis and release. Since the AVP system is also modulated by nitric oxide (NO), we studied if deoxycorticosterone acetate (DOCA) treatment changed the number of nicotinamide adenine dinucleotide phosphate-diaphorase (NADPH-d) active neurons and neuronal NO synthase (nNOS)-immunoreactive (IR) cells in the PVN and SON. After four injections of DOCA (10 mg/rat per day), rats developed a salt appetite and increased NADPH-d active and nNOS-IR neurons in both nuclei. A single DOCA injection did not change salt consumption or nNOS-IR cells, but increased the number of NADPH-d positive neurons in the PVN only. Therefore, while acute MC treatment stimulated the activity of pre-existing enzyme, chronic steroid treatment recruited additional neurons showing nNOS immunoreactivity/NADPH-d activity. These data suggest a role for NO produced in the PVN and SON in DOCA stimulatory effects on AVP mRNA and salt appetite.

Expression of neuronal nitric oxide synthase mRNA in stress-related brain areas after restraint in rats.

The objective of the present study was to investigate the expression of neuronal nitric oxide synthase (nNOS) mRNA in stress-related areas after restraint. Male Wistar rats (n=4-6/group) submitted to 2 h of restraint during one (acute) or seven (chronic) days were sacrificed 24 h after the last restraint period. In situ hybridisation was performed with oligonucleotide probes radiolabeled with (35)S. Acute restraint induced a significant increase in nNOS mRNA in the paraventricular hypothalamic nucleus (PVN), medial parvocellular part, dorsolateral periaqueductal grey (DLPAG) and medial amygdaloid nucleus, but not in the hippocampal formation. This effect persisted after chronic restraint in the PVN and DLPAG. These results suggest that restraint stress induces changes in gene expression of nNOS in areas related to stress reactions.