Effects of escalating versus fixed ethanol exposure on ∆FosB expression in the mesocorticolimbic pathway in adolescent and adult rats.

Background: We have reported induction of ∆FosB in adolescent rats that drank less ethanol than adults yet exhibited a progressive increase in ethanol intake.Objective: To test the hypothesis that an escalating pattern of ethanol exposure is more effective to induce ∆FosB expression [at prelimbic cortex (PrL), nucleus accumbens core and shell, striatum, basolateral amygdala (BLA) and central amygdala (CeC)] than a pattern equated for number of exposures yet employing a fixed ethanol dose.Methods: Adolescent and adult (Exp. 1, n = 48) male and female (n = 24 of each sex) or only adult male (Exp. 2, n = 36) Wistar rats were intermittently intubated with vehicle, escalating (from 0.5 to 2.5 g/kg) or fixed (2.0 g/kg) doses of ethanol, across 18 sessions. ∆FosB induction was assessed using immunohistochemistry. Ethanol intake, anxiety and risk-taking were assessed (in adults only) via two-bottles tests and the multivariate concentric square field.Results: Both patterns heightened ∆FosB levels similarly in adolescents and adults and in males and females. Fixed dosing induced ∆FosB in all areas (p < .05) except the CeC, whereas the escalating pattern induced ∆FosB in the PrL and BLA only (p < .05). Ethanol intake was initially lower in ethanol pre-exposed subjects than in control subjects (p < .05). Rats exposed to the fixed pattern exhibited enhanced risk-taking behavior (p < .05).Conclusions: The results agree with studies showing ethanol-mediated induction of ∆FosB in reward areas and indicate that, following ethanol intubations, this induction is similar in adolescents and adults. The induction of ∆FosB seems not necessarily associated with susceptibility for ethanol intake.

Sex chromosome complement involvement in angiotensin receptor sexual dimorphism.

This study aimed to define whether sex chromosome complement (SCC) may differentially modulate sex differences in relative gene expression of basal Agtr1a, Agtr2, and Mas1 receptors at fore/hindbrain nuclei and at medulla/cortical kidney. Samples were collected from gonadectomized male (XX and XY) and female (XX and XY) mice of the "four core genotypes" model. At brain level, a SCC effect at the area postrema was demonstrated. An increase in mRNA level of Agtr1a and Agtr1a/Agtr2 ratio in XY-SCC mice was associated with a decrease in Mas1 compared to XX-SCC mice. In the renal cortex, a SCC effect for Agtr2 and Mas1 was observed. Regardless of sex (male or female), XX-SCC mice expressed higher levels of mRNA Agtr2 and Mas1 than XY-SCC mice {F(1,12) = 6,126,p < 0.05; F(1,21) = 5,143,p < 0.05}. Furthermore, XX-female mice showed a significant increase in Mas1 expression compared to XY-female mice. These results reveal a SCC modulatory effect at central and kidney level on angiotensin receptor expression, with an enhancement of the vasodilatory arm in XX-mice and an increase in the vasoconstriction arm in XY-mice, which may underlie sex differences in the regulation of arterial pressure.

Body sodium overload modulates the firing rate and fos immunoreactivity of serotonergic cells of dorsal raphe nucleus.

In order to determine whether serotonergic (5HT) dorsal raphe nucleus (DRN) cells are involved in body sodium status regulation, the effect of a s.c. infusion of either 2 M or 0.15 M NaCl on 5HT DRN neuron firing was studied using single unit extracellular recordings. In separate groups of 2 M and 0.15 M NaCl-infused rats, water intake, oxytocin (OT) plasma concentration, urine and plasma sodium and protein concentrations were also measured. Also, to determine the involvement of particular brain nuclei and neurochemical systems in body sodium overload (SO), animals from both groups were perfused for brain immunohistochemical detection of Fos, Fos-OT and Fos-5HT expression. SO produced a significant increase in serotonergic DRN neuron firing rate compared to baseline and 0.15 M NaCl-infused rats. As expected, 2 M NaCl s.c. infusion also induced a significant increase of water intake, diuresis and natriuresis, plasma sodium concentration and osmolality, even though plasma volume did not increase as indicated by changes in plasma protein concentration. The distribution of neurons along the forebrain and brainstem expressing Fos after SO showed the participation of the lamina terminalis, extended amygdala, supraoptic and paraventricular hypothalamic nuclei in the neural network that controls osmoregulatory responses. Both Fos-OT immunoreactive and plasma OT concentration increased after s.c. hypertonic sodium infusion. Finally, matching the "in vivo" electrophysiological study, SO doubled the number of Fos-5HT immunolabeled cells within the DRN. In summary, the results characterize the behavioral, renal and endocrine responses after body sodium overload without volume expansion and specify the cerebral nuclei that participate at different CNS levels in the control of these responses. The electrophysiological approach also allows us to determine in an "in vivo" model that DRN 5HT neurons increase their firing frequency during an increase in systemic sodium concentration and osmolality, possibly to modulate sodium and water intake/excretion and avoid extracellular volume expansion.

Activation of lateral parabrachial afferent pathways and endocrine responses during sodium appetite regulation.

Modulation of salt appetite involves interactions between the circumventricular organs (CVOs) receptive areas and inhibitory hindbrain serotonergic circuits. Recent studies provide support to the idea that the serotonin action in the lateral parabrachial nucleus (LPBN) plays an important inhibitory role in the modulation of sodium appetite. The aim of the present work was to identify the specific groups of neurons projecting to the LPBN that are activated in the course of sodium appetite regulation, and to analyze the associated endocrine response, specifically oxytocin (OT) and atrial natriuretic peptide (ANP) plasma release, since both hormones have been implicated in the regulatory response to fluid reestablishment. For this purpose we combined the detection of a retrograde transported dye, Fluorogold (FG) injected into the LPBN with the analysis of the Fos immunocytochemistry brain pattern after sodium intake induced by sodium depletion. We analyzed the Fos-FG immunoreactivity after sodium ingestion induced by peritoneal dialysis (PD). We also determined OT and ANP plasma concentration by radioimmunoassay (RIE) before and after sodium intake stimulated by PD. The present study identifies specific groups of neurons along the paraventricular nucleus, central extended amygdala, insular cortex, dorsal raphe nucleus, nucleus of the solitary tract and the CVOs that are activated during the modulation of sodium appetite and have direct connections with the LPBN. It also shows that OT and ANP are released during the course of sodium satiety and fluid reestablishment. The result of this brain network activity may enable appropriate responses that re-establish the body fluid balance after induced sodium consumption.

Lateral parabrachial afferent areas and serotonin mechanisms activated by volume expansion.

Recent evidence has shown that the serotonergic mechanism of the lateral parabrachial nucleus (LPBN) participates in the regulation of renal and hormonal responses to isotonic blood volume expansion (BVE). We investigated the BVE-induced Fos activation along forebrain and hindbrain nuclei and particularly within the serotonergic clusters of the raphé system that directly project to the LPBN. We also examined whether there are changes in the concentration of serotonin (5HT) within the raphé nucleus in response to the same stimulus. With this purpose, we analyzed the cells doubly labeled for Fos and Fluorogold (FG) following BVE (NaCl 0.15 M, 2 ml/100 g b.w., 1 min) 7 days after FG injection into the LPBN. Compared with the control group, blood volume-expanded rats showed a significant greater number of Fos-FG double-labeled cells along the nucleus of the solitary tract, locus coeruleus, hypothalamic paraventricular nucleus, central extended amygdala complex, and dorsal raphé nucleus (DRN) cells. Our study also showed an increase in the number of serotonergic DRN neurons activated in response to isotonic BVE. We also observed decreased levels of 5HT and its metabolite 5-hydroxyindoleacetic acid (measured by high-pressure liquid chromatography) within the raphé nucleus 15 min after BVE. Given our previous evidence on the role of the serotonergic system in the LPBN after BVE, the present morphofunctional findings suggest the existence of a key pathway (DRN-LPBN) that may control BVE response through the modulation of 5HT release.

Dorsal raphe nuclei integrate allostatic information evoked by depletion-induced sodium ingestion.

Structures of the lamina terminalis (LT) sense and integrate information reflecting the state of body water and sodium content. Output from the LT projects into a neural network that regulates body fluid balance. Serotonin (5-HT) and the dorsal raphe nuclei (DRN) have been implicated in the inhibitory control of salt intake (i.e., sodium appetite). Signals arriving from the LT evoked by fluid depletion-induced sodium ingestion interact with this inhibitory serotonergic system. We investigated the role of neurons along the LT that directly project to the DRN. We analyzed the pattern of immunoreactivity (ir) of LT cells double-labeled for Fos (a marker of neural activity) and Fluorogold (FG; a retrograde tracer) following sodium depletion-induced sodium intake. Seven days after injection of FG into the DRN, sodium appetite was induced by furosemide injection and overnight access to only a low sodium diet (Furo-LSD) and distilled water. Twenty-four hours later, access to 0.3 M NaCl was given to depleted or sham-depleted rats and sodium intake was measured over the following 60 min. Ninety minutes after the termination of the intake test, the animals were perfused and their brains were processed for immunohistochemical detection of Fos and FG. Compared to sham-depleted animals there was a significantly greater number of Fos-/FG-ir double-labeled cells in the subfornical organ, the organum vasculosum of the lamina terminalis and the median preoptic nucleus in rats that ingested NaCl. Projections from the LT cells may contribute to inhibitory mechanisms involving 5-HT neurons in the DRN that limit the intake of sodium and prevent excess volume expansion.