GABAergic and glutamatergic transmission reveals novel cardiovascular and urinary bladder control features in the shell nucleus accumbens.

The shell Nucleus Accumbens (NAcc) projects to the lateral preoptic area, which is involved in the central micturition control and receives inputs from medullary areas involved in cardiovascular control. We investigated the role of GABAergic and glutamatergic transmission in the shell NAcc on intravesical pressure (IP) and cardiovascular control. Male Wistar rats with guide cannulas implanted bilaterally in the shell NAcc 7 days prior to the experiments were anesthetized with 2% isoflurane in 100% O2 and subjected to cannulation of the femoral artery and vein for mean arterial pressure (MAP) and heart rate recordings (HR) and infusion of drugs, respectively. The urinary bladder (UB) was cannulated for IP measurement. A Doppler flow probe was placed around the renal arterial for renal blood flow (RBF) measurement. After the baseline MAP, HR, IP and RBF recordings for 15 min, GABA or bicuculline methiodate (BMI) or L-glutamate or kynurenic acid (KYN) or saline (vehicle) were bilaterally injected into the shell NAcc and the variables were measured for 30 min. Data are as mean ± SEM and submitted to Student́s t test. GABA injections into the shell NAcc evoked a significant fall in MAP and HR and increased IP and RC compared to saline. L-glutamate in the shell NAcc increased MAP, HR and IP and reduced RC. Injections of BMI and KYN elicited no changes in the variables recorded. Therefore, the GABAergic and glutamatergic transmissions in neurons in the shell NAcc are involved in the neural pathways responsible for the central cardiovascular control and UB regulation.

Sympathetic and angiotensinergic activity in spontaneously hypertensive rats treated with 3-amino-1,2,4-triazole.

Previous studies from our laboratory have shown that the pressor response to intracerebroventricular (icv) administered ANG II in normotensive rats or spontaneously hypertensive rats (SHRs) is attenuated by increased central H2O2 concentration, produced either by direct H2O2 icv injection or by increased endogenous H2O2 centrally in response to local catalase inhibition with 3-amino-1,2,4-triazole (ATZ). In the present study, we evaluated the effects of ATZ administered peripherally on arterial pressure and sympathetic and angiotensinergic activity in SHRs. Male SHRs weighing 280-330 g were used. Mean arterial pressure (MAP) and heart rate (HR) were recorded in conscious freely moving SHRs. Acute intravenous injection of ATZ (300 mg/kg of body weight) did not modify MAP and HR during the next 4 h, however, the treatment with ATZ (300 mg/kg of body weight twice per day) for 3 days reduced MAP (144 ± 6, vs. saline, 183 ± 13 mmHg), without changing HR. Intravenous hexamethonium (ganglionic blocker) produced a smaller decrease in MAP 4 h after ATZ (-25 ± 3, vs saline -38 ± 4 mmHg). Losartan (angiotensinergic AT1 receptor blocker) produced a significant depressor response 4 h after ATZ (-22 ± 4, vs. saline: -2 ± 4 mmHg) and in 3-day ATZ treated SHRs (-25 ± 5, vs. saline: -9 ± 4 mmHg). The results suggest that the treatment with ATZ reduces sympathetic activity in SHRs and simultaneously increases angiotensinergic activity.

Palatability profile in spontaneously hypertensive rats.

The spontaneously hypertensive rats (SHRs) have enhanced palatability for NaCl taste as measured by the increased number of hedonic versus aversive responses to intraoral infusion (1 mL/1 min) of 0.3 M NaCl, in a taste reactivity test in euhydrated condition or after 24 h of water deprivation + 2 h of partial rehydration (WD-PR). SHRs also ingested more sucrose than normotensive rats, without differences in quinine hydrochloride intake. Here, we investigated the palatability of SHRs (n = 8-10) and normotensive Holtzman rats (n = 8-10) to sucrose and quinine sulphate infused intraorally in the same conditions that NaCl palatability was increased in SHRs. SHRs had similar number of hedonic responses to 2% sucrose in euhydrated condition (95 ± 19) or after WD-PR (142 ± 25), responses increased when compared with normotensive rats in euhydrated condition (13 ± 3) or after WD-PR (21 ± 6). SHRs also showed increased number of aversive responses to 1.4 mM quinine sulphate compared with normotensive rats, whether in euhydrated condition (86 ± 6, vs. normotensive: 54 ± 7) or after WD-PR (89 ± 9, vs. normotensive: 40 ± 9). The results suggest that similar to NaCl taste, sweet taste responses are increased in SHRs and resistant to challenges in bodily fluid balance. They also showed a more intense aversive response in SHRs to bitter taste compared with normotensives. This suggests that the enhanced response of SHRs to taste rewards does not correspond to a decreased response to a typical aversive taste.

Chronic administration of catalase inhibitor attenuates hypertension in renovascular hypertensive rats.

AIMS: Reactive oxygen species like hydrogen peroxide (H2O2) are produced endogenously and may participate in intra- and extracellular signaling, including modulation of angiotensin II responses. In the present study, we investigated the effects of chronic subcutaneous (sc) administration of the catalase inhibitor 3-amino-1,2,4-triazole (ATZ) on arterial pressure, autonomic modulation of arterial pressure, hypothalamic expression of AT1 receptors and neuroinflammatory markers and fluid balance in 2-kidney, 1clip (2K1C) renovascular hypertensive rats. MATERIALS AND METHODS: Male Holtzman rats with a clip occluding partially the left renal artery and chronic sc injections of ATZ were used. KEY FINDINGS: Subcutaneous injections of ATZ (600 mg/kg of body weight/day) for 9 days in 2K1C rats reduced arterial pressure (137 ± 8, vs. saline: 182 ± 8 mmHg). ATZ also reduced the sympathetic modulation and enhanced the parasympathetic modulation of pulse interval, reducing the sympatho-vagal balance. Additionally, ATZ reduced mRNA expression for interleukins 6 and IL-1ß, tumor necrosis factor-α, AT1 receptor (0.77 ± 0.06, vs. saline: 1.47 ± 0.26 fold change), NOX 2 (0.85 ± 0.13, vs. saline: 1.75 ± 0.15 fold change) and the marker of microglial activation, CD 11 (0.47 ± 0.07, vs. saline, 1.34 ± 0.15 fold change) in the hypothalamus of 2K1C rats. Daily water and food intake and renal excretion were only slightly modified by ATZ. SIGNIFICANCE: The results suggest that the increase of endogenous H2O2 availability with chronic treatment with ATZ had an anti-hypertensive effect in 2K1C hypertensive rats. This effect depends on decreased activity of sympathetic pressor mechanisms and mRNA expression of AT1 receptors and neuroinflammatory markers possibly due to reduced angiotensin II action.

Involvement of V1-type vasopressin receptors on NaCl intake by hyperosmotic rats treated with muscimol in the lateral parabrachial nucleus.

GABAA receptor activation with agonist muscimol in the lateral parabrachial nucleus (LPBN) induces 0.3 M NaCl intake. In the present study, we investigated water and 0.3 M NaCl intake in male adult rats treated with losartan (angiotensin AT1 receptor antagonist) or MeT-AVP (V1-type vasopressin receptor antagonist) combined with muscimol or methysergide (5-HT2 antagonist) into the LPBN in rats treated with intragastric 2 M NaCl. After 2 M NaCl load and bilateral injections of muscimol (0.5 nmol/0.2 µL) into the LPBN, rats ingested water and 0.3 M NaCl. The pre-treatment of the LPBN with MeT-AVP (1 nmol/0.2 µL) but not losartan (50 µg/0.2 µL) in muscimol treated rats reduced 0.3 M NaCl intake. The pre-treatment of the LPBN with MeT-AVP did not modify the increased 0.3 M NaCl intake in rats treated with methysergide (4 µg/0.2 µL), suggesting that the effect of MeT-AVP was not due to non-specific inhibition of ingestive behavior. The results suggest that endogenous vasopressin in the LPBN facilitates the effects of GABAergic activation driving cell-dehydrated male rats to ingest 0.3 M NaCl.

A critique on the theory of homeostasis.

The objective of this critique is to demonstrate that the theory of "internal environment" (TIE) does not support the theory of "homeostasis" (TOH). We review and conclude that remains valid the concept of "internal environment", which corresponds anatomically to the extracellular fluid (ECF) that bathes tissue cells. The Claude Bernard's classification of "life", a corollary of the TIE under a strict "reactive" paradigm, we then interpret as a classification of how animals behave in response to environmental changes. According to such interpretation, the two theories agree that, when facing changes in the external environment, animals with "free" behavior regulate essential metabolism factors present in the ECF. These are "internalized environmental factors" or IEF (temperature, O2, water, and basic organic and inorganic "nutrients"), a marine legacy of the evolution of the body fluid compartments. However, we show that have empirical and logical shortcomings key inferences derived from the TIE. Such inferences representing traditional premises of TOH we summarize here in two axioms: "if free behavior then regulated IEF" and "all behavioral mechanisms regulate the IEF". In addition, whereas "stability" means "free behavior versus dormancy" in TIE, it means "tissue cells that resist destruction" in TOH. This leads to inevitable contradictions, here discussed at length, that reduce the scope of TOH. We might be in need of a theory that considers not only where TIE and TOH are superficially valid, but also where they crucially diverge, in order to explain "stability" as applied to physiology and behavior.

Anti-hypertensive effect of hydrogen peroxide acting centrally.

Intracerebroventricular (icv) injection of hydrogen peroxide (H2O2) or the increase of endogenous H2O2 centrally produced by catalase inhibition with 3-amino-1,2,4-triazole (ATZ) injected icv reduces the pressor responses to central angiotensin II (ANG II) in normotensive rats. In the present study, we investigated the changes in the arterial pressure and in the pressor responses to ANG II icv in spontaneously hypertensive rats (SHRs) and 2-kidney, 1-clip (2K1C) hypertensive rats treated with H2O2 injected icv or ATZ injected icv or intravenously (iv). Adult male SHRs or Holtzman rats (n = 5-10/group) with stainless steel cannulas implanted in the lateral ventricle were used. In freely moving rats, H2O2 (5 µmol/1 µl) or ATZ (5 nmol/1 µl) icv reduced the pressor responses to ANG II (50 ng/1 µl) icv in SHRs (11 ± 3 and 17 ± 4 mmHg, respectively, vs. 35 ± 6 mmHg) and 2K1C hypertensive rats (3 ± 1 and 16 ± 3 mmHg, respectively, vs. 26 ± 2 mmHg). ATZ (3.6 mmol/kg of body weight) iv alone or combined with H2O2 icv also reduced icv ANG II-induced pressor response in SHRs and 2K1C hypertensive rats. Baseline arterial pressure was also reduced (-10 to -15 mmHg) in 2K1C hypertensive rats treated with H2O2 icv and ATZ iv alone or combined and in SHRs treated with H2O2 icv alone or combined with ATZ iv. The results suggest that exogenous or endogenous H2O2 acting centrally produces anti-hypertensive effects impairing central pressor mechanisms activated by ANG II in SHRs or 2K1C hypertensive rats.

Catalase blockade reduces the pressor response to central cholinergic activation.

Intracerebroventricular (icv) injection of hydrogen peroxide (H2O2), a reactive oxygen species, or the blockade of catalase (enzyme that degrades H2O2 into H2O and O2) with icv injection of 3-amino-1,2,4-triazole (ATZ) reduces the pressor effects of angiotensin II also injected icv. In the present study, we investigated the effects of ATZ injected icv or intravenously (iv) on the pressor responses induced by icv injections of the cholinergic agonist carbachol, which similar to angiotensin II induces pressor responses that depend on sympathoexcitation and vasopressin release. In addition, the effects of H2O2 icv on the pressor responses to icv carbachol were also tested to compare with the effects of ATZ. Normotensive non-anesthetized male Holtzman rats (280-300 g, n = 8-9/group) with stainless steel cannulas implanted in the lateral ventricle were used. Previous injection of ATZ (5 nmol/1 µl) or H2O2 (5 µmol/1 µl) icv similarly reduced the pressor responses induced by carbachol (4 nmol/1 µl) injected icv (13 ± 4 and 12 ± 4 mmHg, respectively, vs. vehicle + carbachol: 30 ± 5 mmHg). ATZ (3.6 mmol/kg of body weight) injected iv also reduced icv carbachol-induced pressor responses (21 ± 2 mmHg). ATZ icv or iv and H2O2 icv injected alone produced no effect on baseline arterial pressure. The treatments also produced no significant change of heart rate. The results show that ATZ icv or iv reduced the pressor responses to icv carbachol, suggesting that endogenous H2O2 acting centrally inhibits the pressor mechanisms (sympathoactivation and/or vasopressin release) activated by central cholinergic stimulation.

Central muscarinic and LPBN mechanisms on sodium intake.

Central cholinergic activation stimulates water intake, but also NaCl intake when the inhibitory mechanisms are blocked with injections of moxonidine (α2 adrenergic/imidazoline agonist) into the lateral parabrachial nucleus (LPBN). In the present study, we investigated the involvement of central M1 and M2 muscarinic receptors on NaCl intake induced by pilocarpine (non-selective muscarinic agonist) intraperitoneally combined with moxonidine into the LPBN or by muscimol (GABAA agonist) into the LPBN. Male Holtzman rats with stainless steel cannulas implanted bilaterally in the LPBN and in the lateral ventricle were used. Pirenzepine (M1 muscarinic antagonist, 1 nmol/1 µl) or methoctramine (M2 muscarinic antagonist, 50 nmol/1 µL) injected intracerebroventricularly (i.c.v.) reduced 0.3 M NaCl and water intake in rats treated with pilocarpine (0.1 mg/100 g of body weight) injected intraperitoneally combined with moxonidine (0.5 nmol/0.2 µL) into the LPBN. In rats treated with muscimol (0.5 nmol/0.2 µL) into the LPBN, methoctramine i.c.v. also reduced 0.3 M NaCl and water intake, however, pirenzepine produced no effect. The results suggest that M1 and M2 muscarinic receptors activate central pathways involved in the control of water and sodium intake that are under the influence of the LPBN inhibitory mechanisms.

Opioid and α2 adrenergic mechanisms are activated by GABA agonists in the lateral parabrachial nucleus to induce sodium intake.

The activation of GABA, opioid or α2 adrenergic mechanisms in the lateral parabrachial nucleus (LPBN) facilitates hypertonic NaCl intake in rats. In the present study, we combined opioid or α2 adrenergic antagonists with GABA agonists into the LPBN in order to investigate if NaCl intake caused by GABAergic activation in normohydrated rats depends on opioid or α2-adrenergic mechanisms in this area. Male Holtzman rats with stainless steel cannulas implanted bilaterally in the LPBN were used. Bilateral injections of muscimol or baclofen (GABAA and GABAB agonists, respectively, 0.5 nmol/0.2 µl) into the LPBN induced strong ingestion of 0.3 M NaCl (45.8 ±â€¯7.3 and 21.8 ±â€¯4.8 ml/240 min, respectively) and water intake (22.7 ±â€¯3.4 and 6.6 ±â€¯2.5 ml/240 min, respectively). Naloxone (opioid antagonist, 150 nmol/0.2 µl) into the LPBN abolished 0.3 M NaCl and water intake to muscimol (2.0 ±â€¯0.6 and 0.9 ±â€¯0.2 ml/240 min, respectively) or baclofen (2.3 ±â€¯1.1 and 0.8 ±â€¯0.4 ml/240 min, respectively). RX 821002 (α2 adrenoceptor antagonist, 10 nmol/0.2 µl) into the LPBN reduced 0.3 M NaCl intake induced by the injections of muscimol or baclofen (26.6 ±â€¯8.0 and 10.1 ±â€¯4.9 ml/240 min, respectively). RX 821002 reduced water intake induced by muscimol (7.7 ±â€¯2.9 ml/240 min), not by baclofen. The results suggest that sodium intake caused by gabaergic activation in the LPBN in normohydrated rats is totally dependent on the activation of opioid mechanisms and partially dependent on the activation of α2 adrenergic mechanisms in the LPBN.

Enhanced angiotensin II induced sodium appetite in renovascular hypertensive rats.

Renovascular hypertensive 2-kidney, 1-clip (2K1C) rats have an increased activity of the renin-angiotensin system and an initial transitory increase in daily water and NaCl intake. However, the dipsogenic and natriorexigenic responses to angiotensin II (ANG II) have not been tested yet in 2K1C rats. Therefore, in the present study, we evaluated water and 0.3 M NaCl intake induced by water deprivation (WD)-partial rehydration (PR) or intracerebroventricular (icv) ANG II in 2K1C rats. In addition, the cardiovascular changes to these treatments were also evaluated. Male Holtzman rats received a silver clip around the left renal artery to induce 2K1C renovascular hypertension. In the 5th week, a group of animals received a guide cannula in the lateral ventricle for icv injections. Daily water intake increased from the 3rd week after surgery and remained elevated until the 6th week (last recording week), whereas daily 0.3 M NaCl intake transiently increased from the 2nd to the 5th week after surgery. On the 6th week, in spite of comparable daily 0.3 M NaCl intake between 2K1C and sham rats, WD-PR and icv ANG II induced an increased 0.3 M NaCl intake in 2K1C rats. Water intake induced by WD-PR, not by icv ANG II, also increased in 2K1C rats. The increase in arterial pressure to WD-PR or icv ANG II was similar in sham and 2K1C rats. Therefore, these results suggest that 2K1C rats are more responsive to the natriorexigenic effects of ANG II, whereas other responses to ANG II are not modified.

The lateral parabrachial nucleus and central angiotensinergic mechanisms in the control of sodium intake induced by different stimuli.

Angiotensin II (ANG II) is a typical facilitatory stimulus for sodium appetite. Surprisingly, hyperosmolarity and central cholinergic stimulation, two classical antinatriorexigenic stimuli, also facilitate NaCl intake when they are combined with injections of the α2-adrenoceptor/imidazoline agonist moxonidine into the lateral parabrachial nucleus (LPBN). In the present study, we tested the relative importance of central angiotensinergic and cholinergic mechanisms for the control of water and NaCl intake by combining different dipsogenic or natriorexigenic stimuli with moxonidine injection into the LPBN. Adult male Holtzman rats (n=9-10/group) with stainless steel cannulas implanted in the lateral ventricle and LPBN were used. Bilateral injections of moxonidine (0.5 nmol) into the LPBN increased water and 0.3M NaCl intake in rats that received furosemide+captopril injected subcutaneously, ANG II (50ng) or carbachol (cholinergic agonist, 4 nmol) injected intracerebroventricularly (icv) or 2M NaCl infused intragastrically (2ml/rat). Losartan (AT1 antagonist, 100µg) or atropine (muscarinic antagonist, 20 nmol) injected icv abolished the effects on water and 0.3M NaCl of moxonidine combined to either 2M NaCl intragastrically or carbachol icv. However, atropine icv did not change 0.3M NaCl intake produced by direct central action of ANG II like that induced by ANG II icv or furosemide+captopril combined with moxonidine into the LPBN. The results suggest that different stimuli, including hyperosmolarity and central cholinergic stimulation, share central angiotensinergic activation as a common mechanism to facilitate sodium intake, particularly when they are combined with deactivation of the LPBN inhibitory mechanisms.

Lateral parabrachial nucleus and opioid mechanisms of the central nucleus of the amygdala in the control of sodium intake.

Facilitatory and inhibitory mechanisms in the central nucleus of the amygdala (CeA) and the lateral parabrachial nucleus (LPBN), respectively, are important for the control of sodium and water intake. Here we investigated the importance of the opioid mechanisms in the CeA for water and 0.3M NaCl intake in euhydrated or hyperosmotic rats treated with injections of muscimol (GABAA agonist) or moxonidine (α2 adrenergic/imidazoline agonist) into the LPBN, respectively. Male Holtzman rats (n=4-8/group) with stainless steel cannulas implanted bilaterally in the CeA and in the LPBN were used. The ingestion of 0.3M NaCl and water by euhydrated rats treated with muscimol (0.5nmol/0.2µl) into the LPBN (29.4±2.7 and 15.0±2.4ml/4h, respectively) was abolished by the previous injections of naloxone (opioid antagonist, 40µg/0.2µl) into the CeA (0.7±0.3 and 0.3±0.1ml/4h, respectively). The ingestion of 0.3M NaCl by rats treated with intragastric 2M NaCl (2ml/rat) combined with moxonidine (0.5nmol/0.2µl) into the LPBN (17.0±3.8ml/2h) was also strongly reduced by the previous injections of naloxone into the CeA (3.2±2.5ml/2h). Sucrose intake was not affected by naloxone injections into the CeA, which minimized the possibility of non-specific inhibition of ingestive behaviors with this treatment. The present results suggest that opioid mechanisms in the CeA are essential for hypertonic NaCl intake when the LPBN inhibitory mechanisms are deactivated or attenuated with injections of muscimol or moxonidine in this area.

Short-term cross-sensitizion of need-free sugar intake by combining sodium depletion and hypertonic NaCl intake.

History of sodium depletion cross-sensitizes the effects of drugs of abuse. The objective of the present study was to find out if history of sodium depletion also cross-sensitizes a natural reward such as sugar intake in the rat. Sodium depletion was induced by furosemide combined with removal of ambient sodium for 24 h; it was repeated seven days later. The depletion was immediately followed by 0.3 M NaCl intake in a sodium appetite test (active sodium repletion). Seven days after the last depletion, hydrated and fed (need-free) sucrose-naïve animals were offered 10% sucrose in a first 2-h sucrose test. The sucrose test was repeated once a day in a series of five consecutive days. History of sodium depletion enhanced sucrose intake in the first and second tests; it had no effect from the third to fifth sucrose test. The effect on the initial sucrose intake tests disappeared if the rats did not ingest 0.3 M NaCl in the sodium appetite test. Prior experience with sucrose intake in need-free conditions had no effect on sodium appetite. History of intracellular dehydration transiently influenced sucrose intake in the first sucrose test. We found no evidence for thirst sensitization. We conclude that history of dehydration, particularly that resulting from sodium depletion, combined to active sodium repletion, produced short-term cross-sensitization of sucrose intake in sucrose-naïve rats. The results suggest that the cross-sensitization of sucrose intake related with acquisition of sugar as a novel nutrient rather than production of lasting effects on sugar rewarding properties.

Water deprivation-partial rehydration induces sensitization of sodium appetite and alteration of hypothalamic transcripts.

iSodium intake occurs either as a spontaneous or induced behavior, which is enhanced, i.e., sensitized, by repeated episodes of water deprivation followed by subsequent partial rehydration (WD-PR). In the present work, we examined whether repeated WD-PR alters hypothalamic transcripts related to the brain renin-angiotensin system (RAS) and apelin system in male normotensive Holtzman rats (HTZ). We also examined whether the sodium intake of a strain with genetically inherited high expression of the brain RAS, the spontaneously hypertensive rat (SHR), responds differently than HTZ to repeated WD-PR. We found that repeated WD-PR, besides enhancing spontaneous and induced 0.3 M NaCl intake, increased the hypothalamic expression of angiotensinogen, aminopeptidase N, and apelin receptor transcripts (43%, 60%, and 159%, respectively) in HTZ at the end of the third WD-PR. Repeated WD-PR did not change the daily spontaneous 0.3 M NaCl intake and barely changed the need-induced 0.3 M NaCl intake of SHR. The same treatment consistently enhanced spontaneous daily 0.3 M NaCl intake in the normotensive Wistar-Kyoto rats. The results show that repeated WD-PR produces alterations in hypothalamic transcripts and also sensitizes sodium appetite in HTZ. They suggest an association between the components of hypothalamic RAS and the apelin system, with neural and behavioral plasticity produced by repeated episodes of WD-PR in a normotensive strain. The results also indicate that the inherited hyperactive brain RAS is not a guarantee for sensitization of sodium intake in the male adult SHR exposed to repeated WD-PR.

Participation of α2 -adrenoceptors in sodium appetite inhibition during sickness behaviour following administration of lipopolysaccharide.

Sickness behaviour, a syndrome characterized by a general reduction in animal activity, is part of the active-phase response to fight infection. Lipopolysaccharide (LPS), an effective endotoxin to model sickness behaviour, reduces thirst and sodium excretion, and increases neurohypophysial secretion. Here we review the effects of LPS on thirst and sodium appetite. Altered renal function and hydromineral fluid intake in response to LPS occur in the context of behavioural reorganization, which manifests itself as part of the syndrome. Recent data show that, in addition to its classical effect on thirst, non-septic doses of LPS injected intraperitoneally produce a preferential inhibition of intracellular thirst versus extracellular thirst. Moreover, LPS also reduced hypertonic NaCl intake in sodium-depleted rats that entered a sodium appetite test. Antagonism of α2 -adrenoceptors abolished the effect of LPS on sodium appetite. LPS and cytokine transduction potentially recruit brain noradrenaline and α2 -adrenoceptors to control sodium appetite and sickness behaviour.

Importance of the central nucleus of the amygdala on sodium intake caused by deactivation of lateral parabrachial nucleus.

The lateral parabrachial nucleus (LPBN) and the central nucleus of the amygdala (CeA) are important central areas for the control of sodium appetite. In the present study, we investigated the importance of the facilitatory mechanisms of the CeA on NaCl and water intake produced by the deactivation of LPBN inhibitory mechanisms. Male Holtzman rats (n=7-14) with stainless steel cannulas implanted bilaterally in the CeA and LPBN were used. Bilateral injections of moxonidine (α2-adrenoceptor/imidazoline agonist, 0.5 nmol/0.2 µl) into the LPBN increased furosemide+captopril-induced 0.3M NaCl (29.7 ± 7.2, vs. vehicle: 4.4 ± 1.6 ml/2h) and water intake (26.4 ± 6.7, vs. vehicle: 8.2 ± 1.6 ml/2h). The GABAA agonist muscimol (0.25 nmol/0.2 µl) injected bilaterally into the CeA abolished the effects of moxonidine into the LPBN on 0.3M NaCl (2.8 ± 1.6 ml/2h) and water intake (3.3 ± 2.3 ml/2h). Euhydrated rats treated with muscimol (0.5 nmol/0.2 µl) into the LPBN also ingested 0.3M NaCl (19.1 ± 6.4 ml/4h) and water (8.8 ± 3.2 ml/4h). Muscimol (0.5 nmol/0.2 µl) into the CeA also abolished 0.3M NaCl (0.1 ± 0.04 ml/4h) and water intake (0.1 ± 0.02 ml/4h) in euhydrated treated with muscimol into the LPBN. The present results show that neuronal deactivation of the CeA abolishes NaCl intake produced by the blockade of LPBN inhibitory mechanisms, suggesting an interaction between facilitatory mechanisms of the CeA and inhibitory mechanisms of the LPBN in the control of NaCl intake.

Mapping brain Fos immunoreactivity in response to water deprivation and partial rehydration: Influence of sodium intake.

Water deprivation (WD) followed by water intake to satiety, produces satiation of thirst and partial rehydration (PR). Thus, WD-PR is a natural method to differentiate thirst from sodium appetite. WD-PR also produces Fos immunoreactivity (Fos-ir) in interconnected areas of a brain circuit postulated to subserve sodium appetite. In the present work, we evaluated the effect of sodium intake on Fos-ir produced by WD-PR in brain areas operationally defined according to the literature as either facilitatory or inhibitory to sodium intake. Isotonic NaCl was available for ingestion in a sodium appetite test performed immediately after a single episode of WD-PR. Sodium intake decreased Fos-ir in facilitatory areas such as the lamina terminalis (particularly subfornical organ and median preoptic nucleus), central amygdala and hypothalamic parvocellular paraventricular nucleus in the forebrain. Sodium intake also decreased Fos-ir in inhibitory areas such as the area postrema, lateral parabrachial nucleus and nucleus of the solitary tract in the hindbrain. In contrast, sodium intake further increased Fos-ir that was activated by water deprivation in the dorsal raphe nucleus, another inhibitory area localized in the hindbrain. WD-PR increased Fos-ir in the core and shell of the nucleus accumbens. Sodium intake reduced Fos-ir in both parts of the accumbens. In summary, sodium intake following WD-PR reduced Fos-ir in most facilitatory and inhibitory areas, but increased Fos-ir in another inhibitory area. It also reduced Fos-ir in a reward area (accumbens). The results suggest a functional link between sodium intake and the activity of the hindbrain-forebrain circuitry subserving reward and sodium appetite in response to water deprivation.