Exercise training reduces sympathetic activity in hypertensive humans and rats. We hypothesized that the swimming exercise would change the neurotransmission in the rostral ventrolateral medulla (RVLM), a key region involved in sympathetic outflow, and hemodynamic control in spontaneously hypertensive rats (SHR) and Wistar-Kyoto (WKY) rats. Bilateral injections of kynurenic acid (KYN) were carried out in the RVLM in sedentary- (S-) or exercised- (E-) SHR and WKY rats submitted to swimming for 6 weeks. Rats were α-chloralose anesthetized and artificially ventilated, with Doppler flow probes around the lower abdominal aorta and superior mesenteric artery. Injections into the RVLM were made before and after i.v. L-NAME (nitric oxide synthase, NOS, inhibitor). Injections of KYN into the RVLM elicited a major vasodilation in the hindlimb more than in the mesenteric artery in E-SHR compared to S-SHR, but similar decrease in arterial pressure was observed in both groups. Injections of KYN into the RVLM after i.v. L-NAME attenuated the hindlimb vasodilation evoked by KYN and increased the mesenteric vasodilation in E-SHR. Swimming exercise can enhance the hindlimb vasodilation mediated by peripheral NO release, reducing the activation of neurons with EAA receptors in the RVLM in SHR.

Maintaining the composition and volume of the extracellular fluid (ECF) and intracellular fluid (ICF) within a restricted range of variation is critical for normal tissue perfusion and cellular function (Strange 1993). The ECF is primarily composed by the vascular and extravascular (interstitial) fluid, and has sodium as a determinant ion of its osmolarity and volume. The ECF/plasma volume ratio is of utmost importance for maintaining vascular capacitance and, consequently, venous return, cardiac output, and arterial pressure (Ramsay 1991). Maintaining the variability of the ECF volume within a strict range is a central goal of homeostatic mechanisms.The central nervous system (CNS) is informed about alterations in volume and tonicity of the ECF through the activity of peripheral and central sensory receptors. Receptors that respond to alterations in the volume of ECF are sensitive to the degree of mechanical distension of blood vessels or cardiac chamber (reviewed by Bourque 2008). They are located in the wall of the atrium and pulmonary vessels (cardiopulmonary receptors), and in the adventitia of the aortic arch/carotid sinus (arterial baroreceptors), or renal afferent arterioles (renal baroreceptors). These receptors mediate central autonomic reflexes through primary connections to the nucleus of the solitary tract (NTS) or mediate renorenal reflexes. Receptors that respond to alterations in ICF are sensitive to ECF tonicity or effective osmotic concentration (Bourque 2008; Kuramochi and Kobayashi 2000; McKinley et al. 1992). They are located in renal, intestinal, and hepatic vessels, in the wall of the gastrointestinal tract and oropharyngeal cavity and in brain circumventricular organs, notably area postrema, subfornical organ (SFO), and organum vasculosum of the lamina terminalis (OVLT). Efferent responses to the activation of these receptors include renal sympathetic nerve activity, hormone secretion [atrial natriuretic peptide (ANP), oxytocin, vasopressin, renin–angiotensin II], and hemodynamic changes. Such responses culminate in controlled alterations of renal sodium and water excretion.Among these several mechanisms and responses, we would like to call attention to those associated with ECF volume expansion. Dehydration is a menace to immediate survival, but volume expansion is associated with long-term changes that affect health, particularly hypertension and heart failure (reviewed by Antunes-Rodrigues et al. 2004; Toney and Stocker 2010; Toney et al. 2010). Conversely, hypertonic NaCl also has a potential therapeutic value to recovery from hemorrhagic shock (Pedrino et al. 2011; Rocha e Silva et al. 1986; Velasco et al. 1980). In this chapter, we will focus on cardiovascular responses to volume expansion and hypertonicity, their sensory afferences, and how these responses are subserved by a brainstem–hypothalamic–preoptic axis and its associated cathecolaminergic pathways.

The paraventricular nucleus of the hypothalamus (PVN) is an important area of the brain involved in the control of cardiovascular system and fluid-electrolyte balance. In the present study we evaluated the effects of hypothalamic disconnection (HD) caudal to PVN in the pressor and dipsogenic responses induced by intracerebroventricular (icv) injections of angiotensin II (ANG II) or carbachol (cholinergic agonist). Male Holtzman rats (280–320 g) with a stainless steel cannula implanted into the lateral ventricle and submitted to sham or HD surgery were used. HD (2 or 15 days) reduced the pressor responses to ANG II (50 ng/1 μl) icv (8 ± 3 and 11 ± 3 mm Hg, respectively, vs. sham: 23 ± 3 and 21 ± 2 mm Hg) or carbachol (4 nmol/1 μl) icv (8 ± 2 and 21 ± 3 mm Hg, respectively, vs. sham: 33 ± 3 and 33 ± 3 mm Hg), without changing baseline arterial pressure. Acutely (2–4 days), HD also reduced water intake to icv ANG II (3.3 ± 2.2 vs. sham: 14.2 ± 3.0 ml/60 min) or carbachol (4.4 ± 1.8 vs. sham: 11.4 ± 1.6 ml/60 min); however, chronically (15–17 days), HD produced no change on ANG II- and carbachol-induced water intake, in spite of the increased daily water intake and urinary volume. The results suggest that medial projections caudal to PVN are important for pressor and dipsogenic responses to central angiotensinergic and cholinergic activation.

Hypernatremia is known to induce renal vasodilation, but the afferent pathways that mediate these responses are not entirely understood. It is known that carotid chemoreceptors are sensitive to changes in plasma sodium concentration, but the physiological significance of this sensitivity is not clear. The present study examines the role of carotid chemoreceptors in cardiovascular responses to intravenous infusion of hypertonic saline (HS). Male Wistar rats (290–350 g) were anesthetized with urethane (1.2 g×kg−1 b.w., i.v.), and instrumented for recording of arterial pressure, renal blood flow (RBF), and renal vascular conductance (RVC). The carotid chemoreceptors were inactivated by bilateral ligation of the carotid body artery (CBA). Sham surgery involved (visualization of these arteries). Thirty min later, HS (3 M NaCl, 1.8 ml×kg−1 b.w., i.v.) was infused in 1 min. These infusions increase plasma osmolality by about 6%. In intact rats (n=8), blood pressure increased slightly (10 min after HS: 9±1.9 mmHg) and transiently (<20 min). HS increased RBF (by 38±4.9%, 10 min after HS) and RVC (by 28±5.1%, at 10 min after HS infusion), and these increases lasted for more than 60 min. In rats submitted to CBA ligation (n=8), the pressor response to HS was larger (24±2.2 mmHg, at 10 min after HS infusion) and prolonged (>60 min). However, increases in RBF and RVC induced by HS were significantly reduced after CBA ligation (RBF 13±5.0% above baseline, RVC 7±4.4% below baseline, 10 min after HS). These results demonstrate that integrity of carotid body afferent is essential for development of the renal vasodilation that follows acute changes in the composition of the extracellular fluid compartment.

Water deprivation and hypernatremia are major challenges for water and sodium homeostasis. Cellular integrity requires maintenance of water and sodium concentration within narrow limits. This regulation is obtained through engagement of multiple mechanisms and neural pathways that regulate the volume and composition of the extracellular fluid. The purpose of this short review is to summarize the literature on central neural mechanisms underlying cardiovascular, hormonal and autonomic responses to circulating volume changes, and some of the findings obtained in the last 12 years by our laboratory. We review data on neural pathways that start with afferents in the carotid body that project to medullary relays in the nucleus tractus solitarii and caudal ventrolateral medulla, which in turn project to the median preoptic nucleus in the forebrain. We also review data suggesting that noradrenergic A1 cells in the caudal ventrolateral medulla represent an essential link in neural pathways controlling extracellular fluid volume and renal sodium excretion. Finally, recent data from our laboratory suggest that these structures may also be involved in the beneficial effects of intravenous infusion of hypertonic saline on recovery from hemorrhagic shock.

The peripheral hyperosmolarity elicited by intravenous infusion of hypertonic saline brings potential benefits to the treatment of hemorrhage. The neural mechanisms involved in these beneficial effects remain unknown. The present study examines the role of carotid chemoreceptors in cardiovascular responses induced by hypertonic saline after hypovolemic hemorrhage in rats. Male Wistar rats (300-400 g) were anesthetized with thiopental, and instrumented for recording of mean arterial pressure. Arterial pressure was reduced to 60 mm Hg by withdrawal of arterial blood over 10 min, and maintained at this level for 60 min by withdrawal or infusion of blood. In control rats (n = 8) with intact chemoreceptors, the subsequent intravenous infusion of hypertonic saline (3M NaCl, 1.8 ml kg(-1) body weight, in 2 min) restored blood pressure (pressure increased from 61 ± 4 to 118 ± 5 mm Hg). In experimental rats (n = 8), the carotid body arteries were tied, 30 min after the beginning of the hypotensive phase, leaving the carotid chemoreceptors ischemic. In these rats, hypertonic saline failed to restore blood pressure (pressure increased from 55 ± 1 to 70 ± 6 mm Hg). These findings suggest that the restoration of blood pressure after hypovolemic hemorrhage induced by hypertonic saline depends on intact carotid chemoreceptors.

Inhibition of the commissural nucleus of the solitary tract (commNTS) induces a fall in sympathetic nerve activity and blood pressure in spontaneously hypertensive rats (SHR), which suggests that this subnucleus of the NTS is a source of sympathoexcitation. Exercise training reduces sympathetic activity and arterial pressure. The purpose of the present study was to investigate whether the swimming exercise can modify the regional vascular responses evoked by inhibition of the commNTS neurons in SHR and normotensive Wistar-Kyoto (WKY) rats. Exercise consisted of swimming, 1 h/day, 5 days/wk for 6 wks, with a load of 2% of the body weight. The day after the last exercise session, the rats were anesthetized with intravenous alpha-chloralose, tracheostomized, and artificially ventilated. The femoral artery was cannulated for mean arterial pressure (MAP) and heart rate recordings, and Doppler flow probes were placed around the lower abdominal aorta and superior mesenteric artery. Microinjection of 50 mM GABA into the commNTS caused similar reductions in MAP in swimming and sedentary SHR (-25 +/- 6 and -30 +/- 5 mmHg, respectively), but hindlimb vascular conductance increased twofold in exercised vs. sedentary SHR (54 +/- 8 vs. 24 +/- 5%). GABA into the commNTS caused smaller reductions in MAP in swimming and sedentary WKY rats (-20 +/- 4 and -16 +/- 2 mmHg). Hindlimb conductance increased fourfold in exercised vs. sedentary WKY rats (75 +/- 2% vs. 19 +/- 3%). Therefore, our data suggest that the swimming exercise induced changes in commNTS neurons, as shown by a greater enhancement of hindlimb vasodilatation in WKY vs. SHR rats in response to GABAergic inhibition of these neurons.

The peripheral hyperosmolarity elicited by intravenous infusion of hypertonic saline (HS) can be beneficial in treating hemorrhagic shock. However, the neural mechanisms involved in this resuscitation remain unknown. The present study sought to determine the effects of selective baroreceptor denervation on arterial blood pressure response during HS resuscitation in rats submitted to hemorrhagic shock. Male Wistar rats (280-320 g) were anesthetized with thiopental sodium (40 mg/kg, i.v.), and the femoral artery and jugular vein were cannulated for MAP and heart rate recording and HS infusion (3 mol/L NaCl; 0.18 mL/100 g body weight, >2 min). Hemorrhagic shock was obtained by withdrawing blood over 30 min until a MAP of 60 mmHg was obtained. This level of MAP was maintained for a further 30 min through subsequent blood withdrawal or reinfusion. Next, animals were divided into selective aortic and/or carotid denervation or sham groups before infusing HS. Results showed that in the sham group (n = 12), HS infusion increased MAP to levels close to baseline (from 65 +/- 3 to 112 +/- 5 mmHg, 10 min after HS). In the aortic denervated group (n = 10), HS infusion also increased MAP (from 54 +/- 3 to 112 +/- 5 mmHg, 10 min after HS). In contrast, in the carotid denervation group (n = 8), the increase in MAP induced by HS infusion was abolished (from 53 +/- 3 to 73 +/- 12 mmHg, 10 min after HS). These results indicate that in hemorrhaged rats, HS infusion produces a pressor effect that is likely to be mediated through carotid rather than aortic baroreceptors.

Several forms of experimental evidence gathered in the last 37 years have unequivocally established that the medulla oblongata harbors the main neural circuits responsible for generating the vasomotor tone and regulating arterial blood pressure. Our current understanding of this circuitry derives mainly from the studies of Pedro Guertzenstein, a former student who became Professor of Physiology at UNIFESP later, and his colleagues. In this review, we have summarized the main findings as well as our collaboration to a further understanding of the ventrolateral medulla and the control of arterial blood pressure under normal and pathological conditions.

Several forms of experimental evidence gathered in the last 37 years have unequivocally established that the medulla oblongata harbors the main neural circuits responsible for generating the vasomotor tone and regulating arterial blood pressure. Our current understanding of this circuitry derives mainly from the studies of Pedro Guertzenstein, a former student who became Professor of Physiology at UNIFESP later, and his colleagues. In this review, we have summarized the main findings as well as our collaboration to a further understanding of the ventrolateral medulla and the control of arterial blood pressure under normal and pathological conditions.

Evidence shows that AngII plays a role in the regulation of RVLM neurons. AngII microinjection into the RVLM of rats results in an increase of the blood pressure (BP) and the sympathetic activity. Further, it is well know that nitric oxide (NO) decreases the release and/or formation of AngII in all areas of the brain stem and reduces AngII receptor binding sites. Thus, the aim is to establish the effect of AngII microinjection into the RVLM of mice and examinate this effect in the absence of neuronal NO. Anesthetized knockout nNOS −/− and wild-type (WT) mice, both of the C57Bl6 strain, were tracheostomized and placed in a stereotaxic frame. Stereotaxic coordinates (bregma): −6,5mm caudal; +1,2mm lateral to midline; and −5,9mm deep. AngII (25pmol) bilateral microinjections were made with the use of glass micropipettes. This data shows that the effects of AngII microinjections into the RVLM of mice are similar to those observed in others animals. However it should be noted that even without the nNO “break”, the BP response was lower in knockout animals, probably implying an unexpected result for the absence of nNO. FAPESP, CNPq, PRONEX

We evaluated if the baroreceptors influence the cNTS in the Hd control during GABAergic blockade in the RVL. Male Wistar rats (300–350g), 10 days sino-aortic denervated (N=6/group), cloralose-anesthetized (60mg/kg, i.v.), tracheostomized, ventilated, and with Doppler flow probes around the superior mesenteric, abdominal aorta and left renal artery were used. Bicuculline (BMI, 2 mM, 60nL) bilaterally into the RVL decreased the hindlimb (HC, −51±11%), mesenteric (MC, −54±9%) and renal (RC, −69±9%) conductances compared to baseline, increasing mean arterial pressure (MAP) (198±9 vs. 107±4 mmHg baseline). After 5 min, muscimol (MS, 6 mM, 60nL) into the midline (m) cNTS produced no changes in the HC, MC and RC, but MAP reduced to 171±8mmHg. Subsequent MS (20nL) bilaterally into the lateral (lat) cNTS increased the HC (167±31%), and RC (210±66%) compared to MS into midline cNTS, reducing MAP to 94±2mmHg. In the control group, BMI into the RVL produced similar Hd changes. Saline into the mcNTS produced small increases in HC (10±8%), MC (17±10%) and RC (18±9%) compared to post-BMI, and no changes on MAP. Saline into the latcNTS did not change HC, MC, and RC, but MAP decreased to 147±10mmHg. The baroreceptors tonically inhibit the latcNTS neurons, which excitate the RVL neurons. Supported by FAPESP, CNPq-PRONEX, NEPAS.

We evaluated the hemodynamic pattern and the contribution of the sympathetic nervous system in conscious and anesthetized (1.4 g/kg urethane, iv) Wistar rats with L-NAME-induced hypertension (20 mg/kg daily). The basal hemodynamic profile was similar for hypertensive animals, conscious (N = 12) or anesthetized (N = 12) treated with L-NAME for 2 or 7 days: increase of total peripheral resistance associated with a decrease of cardiac output (CO) compared to normotensive animals, conscious (N = 14) or anesthetized (N = 14). Sympathetic blockade with hexamethonium essentially caused a decrease in total peripheral resistance in hypertensive animals (conscious, 2 days: from (means +/- SEM) 2.47 +/- 0.08 to 2.14 +/- 0.07; conscious, 7 days: from 2.85 +/- 0.13 to 2.07 +/- 0.33; anesthetized, 2 days: from 3.00 +/- 0.09 to 1.83 +/- 0.25 and anesthetized, 7 days: from 3.56 +/- 0.11 to 1.53 +/- 0.10 mmHg mL-1 min-1) with no change in CO in either group. However, in the normotensive group a fall in CO (conscious: from 125 +/- 4.5 to 96 +/- 4; anesthetized: from 118 +/- 1.5 to 104 +/- 5.5 mL/min) was observed. The responses after hexamethonium were more prominent in the hypertensive anesthetized group. However, no difference was observed between conscious and anesthetized normotensive rats in response to sympathetic blockade. The present study shows that the vasoconstriction in response to L-NAME was mediated by the sympathetic drive. The sympathetic tone plays an important role in the initiation and maintenance of hypertension.

We investigated the role of cNTS in the Hd control during GABAergic blockade in the RVLM in intact rats. Male Wistar rats (300–350g, N=6–7/group), chloralose-anesthetized (60mg/kg, i.v.), tracheostomized, artificially-ventilated, and with miniaturized Doppler flow probes around the superior mesenteric, abdominal aorta and left renal artery were used. Bicuculline (BMI, 2 mM, 60 nL) injected bilaterally into the RVLM decreased the hindlimb (HC, −37±11%), mesenteric (MC, -60±11%) and renal (RC, −48±10%) conductances compared to baseline, and increased MAP (195±9 vs. 119±4 mmHg baseline). The control group also showed similar responses to BMI in the RVL. After 5 min, muscimol (MS, 6 mM, 60 nL) or saline (60 nL, control group) into the midline cNTS similarly increased MC (109±54 and 30±15%, respectively) and RC (40±10 and 71±33%, respectively), but MAP only reduced to 155±7 mmHg with MS. Subsequent MS or saline (20 nL, control group) bilaterally into the lateral cNTS produced no changes on HC, MC or RC, but MAP decreased to 120±3 mmHg with MS. The data suggest that cNTS does not mediate the decrease in HC, MS and RC due to BMI in the RVLM in intact rats. Supported by FAPESP, CNPq-PRONEX, NEPAS.

The splanchnic (SPL) nerve is a postganglionic sympathetic nerve involved in the tonic regulation of the cardiovascular system. Electrical stimulation of this nerve produces mesenteric vasoconstriction and it has been assumed that vasodilatory responses are dependent on inhibition of the vasoconstrictor tone. Several different central stimuli have been shown to dilate the hindquarter vascular bed and constrict the mesenteric vascular bed. To determine whether vasodilatory and vasoconstrictor effects in different vascular beds are elicited by activation of different sympathetic nerves, we investigated the hemodynamic changes in hindquarter, mesenteric and renal vascular beds evoked by electrical stimulation of the SPL nerve. Stimulation of the intact or sectioned SPL nerve in chloralose-anesthetized, artificially ventilated rats evoked increases in the hindquarter vascular conductance and simultaneously decreased the mesenteric and renal vascular conductance. Intravenous (i.v.) administration of l-NAME prior to stimulation of the proximal end of the sectioned SPL nerve abolished the increase in hindquarter conductance, suggesting the involvement of nitric oxide in this response. In assessing the hemodynamic effects of tonic activity on the SPL nerves, no significant changes were observed after unilateral section of the SPL nerve, but bilateral section of the SPL nerves decreased hindquarter conductance and did not significantly change the mesenteric conductance simultaneously. No consistent response was observed in the renal vascular bed after unilateral and subsequent contralateral section of the SPL nerves. These findings demonstrate that electrical stimulation of the SPL nerve produces mesenteric vasoconstriction and simultaneous hindquarter vasodilatation, which is mediated by nitric oxide. Moreover, the present data suggest that SPL nerves may provide a tonic vasodilatory tone in the hindquarter vascular bed and simultaneously a vasoconstrictor tone in another, undetermined vascular bed.

The present study sought to determine the influence of the neuronal isoform of nitric oxide synthase (nNOS) on the baroreflex gain (mean index: bpm/mm Hg) and on the respiratory response to baroreflex activation by using nNOS knockout (nNOS−/−) mice. Experiments were performed with nNOS+/+ wild type (WT; n=12) and nNOS−/− mice (n=11), both of the C57Bl6 strain, randomly assigned to the two groups. Carotid artery and external jugular vein were cannulated under halothane, and after recuperation, urethane was intravenously injected. Baroreflex gain was evaluated by intravenous injection of phenylephrine (PE: 15–18μg/kg) and sodium nitroprusside (SNP: 46–50μg/kg) in WT and nNOS−/− anesthetized mice. At baseline, nNOS−/− mice had a comparable heart rate (604.5±23.6 vs. 618.7±11.2bpm) but higher mean arterial pressure (112.4±6.3 vs. 94.8±3.9mm Hg, P<0.05) than WT mice. Heart rate reflex was significantly reduced (P<0.05) in nNOS−/− mice, tachycardic and bradycardic responses were −1.04±0.3 and −2.03±0.5 in nNOS−/− mice vs. −4±0.7 and −4.52±0.2 in WT mice, respectively. To characterize the effect of arterial pressure changes on respiratory output, PE was injected in a separate group of WT (n=7) and nNOS−/− (n=6) mice. Tidal diaphragm activity decreased in WT mice (P<0.05); however, diaphragm minute respiration and respiratory frequency were not different between the strains of mice. In nNOS−/− group, heart rate reflex by PE injection was significantly reduced (P<0.05). These findings suggest that the absence of nNOS activity leads to an elevation of the baseline blood pressure and also shows the importance of this enzyme on the transmission of baroreflex signals.

Na presente revisão apresentamos dados e elementos sobre o emprego de camundongos no estudo da fisiologia e fisiopatologia cardiovascular. São apontadas as vantagens de seu emprego, como por exemplo, a facilidade de criação e reprodução. Deve se destacar que a possibilidade desses animais serem manipulados por meio da engenharia genética, nos permite obter animais transgênicos, que se caracterizam por manipulação genética visando a introdução de genes de outras espécies, ou, o que é mais comum, a produção de animais knockout. Animais que apresentam deficiência especifica em um gene, não expressando, portanto, determinadas características, como a deficiência do receptor 2A ou a ausência da enzima nNOS. As metodologias e técnicas necessárias para o estudo cardiovascular desses animais são apresentadas e discutidas. São também demonstrados alguns resultados dentre os já obtidos, com o emprego destes animais e uma breve discussão dos mesmos.