5-HT1A, but not 5-HT2 and 5-HT7, receptors in the nucleus raphe magnus modulate hypoxia-induced hyperpnoea

Aim: In the present study, we assessed the role of 5-hydroxytryptamine (5-HT) receptors (5-HT1A, 5-HT2 and 5-HT7) in the nucleus raphe magnus (NRM) on the ventilatory and thermoregulatory responses to hypoxia. Methods: To this end, pulmonary ventilation (V-E) and body temperature (T-b) of male Wistar rats were measured in conscious rats, before and after a 0.1 mu L microinjection of WAY-100635 (5-HT1A receptor antagonist, 3 mu g 0.1 mu L-1, 56 mM), ketanserin (5-HT2 receptor antagonist, 2 mu g 0.1 mu L-1, 36 mM) and SB269970 (5-HT7 receptor antagonist, 4 mu g 0.1 mu L-1, 103 mM) into the NRM, followed by 60 min of severe hypoxia exposure (7% O-2). Results: Intra-NMR microinjection of vehicle (control rats) or 5-HT antagonists did not affect V-E or T-b during normoxic conditions. Exposure of rats to 7% O-2 evoked a typical hypoxia-induced anapyrexia after vehicle microinjections, which was not affected by microinjection of WAY-100635, SB269970 or ketanserin. The hypoxia-induced hyperpnoea was not affected by SB269970 and ketanserin intra-NMR. However, the treatment with WAY-100635 intra-NRM attenuated the hypoxia-induced hyperpnoea. Conclusion: These data suggest that 5-HT acting on 5-HT1A receptors in the NRM increases the hypoxic ventilatory response.

Ace gene dosage influences the development of renovascular hypertension

P>1. Clinical and experimental evidence highlights the importance of the renin-angiotensin system in renovascular hypertension. Furthermore, genetic factors affecting angiotensin-converting enzyme (ACE) could influence the development of renovascular hypertension. 2. To test the effect of small gene perturbations on the development of renovascular hypertension, mice harbouring two or three copies of the Ace gene were submitted to 4 weeks of two-kidney, one-clip (2K1C) hypertension. Blood pressure (BP), cardiac hypertrophy, baroreflex sensitivity and blood pressure and heart rate variability were assessed and compared between the different groups. 3. The increase in BP induced by 2K1C was higher in mice with three copies of the Ace gene compared with mice with only two copies (46 vs 23 mmHg, respectively). Moreover, there was a 3.8-fold increase in the slope of the left ventricle mass/BP relationship in mice with three copies of the Ace gene. Micewith three copies of the Ace gene exhibited greater increases in cardiac and serum ACE activity than mice with only two copies of the gene. Both baroreflex bradycardia and tachycardia were significantly depressed in mice with three copies of the Ace gene after induction of 2K1C hypertension. The variance in basal systolic BP was greater in mice with three copies of the Ace gene after 2K1C hypertension compared with those with only two copies of the gene (106 vs 54%, respectively). In addition, the low-frequency component of the pulse interval was higher mice with three copies of the Ace gene after 2K1C hypertension compared with those with only two (168 vs 86%, respectively). Finally, in mice with three copies of the Ace gene, renovascular hypertension induced a 6.1-fold increase in the sympathovagal balance compared with a 3.2-fold increase in mice with only two copies of the gene. 4. Collectively, these data provide direct evidence that small genetic disturbances in ACE levels per se have an influence on haemodynamic, cardiac mass and autonomic nervous system responses in mice under pathological perturbation.

Involvement of the AT(1) receptor in the venoconstriction induced by angiotensin II in both the inferior vena cava and femoral vein

Although angiotensin II-induced venoconstriction has been demonstrated in the rat vena cava and femoral vein, the angiotensin II receptor subtypes (AT(1) or AT(2)) that mediate this phenomenon have not been precisely characterized. Therefore, the present study aimed to characterize the pharmacological receptors involved in the angiotensin II-induced constriction of rat venae cavae and femoral veins, as well as the opposing effects exerted by locally produced prostanoids and NO upon induction of these vasomotor responses. The obtained results suggest that both AT(1) and AT(2) angiotensin II receptors are expressed in both veins. Angiotensin II concentration-response curves were shifted toward the right by losartan but not by PD 123319 in both the vena cava and femoral vein. Moreover, it was observed that both 10(-5) M indomethacin and 10(-4) M L-NAME improve the angiotensin II responses in the vena cava and femoral vein. In conclusion, in the rat vena cava and femoral vein, angiotensin II stimulates AT(1) but not AT(2) to induce venoconstriction, which is blunted by vasodilator prostanoids and NO. (C) 2010 Elsevier Inc. All rights reserved.

INHIBITION OF THE CAUDAL PRESSOR AREA REDUCES CARDIORESPIRATORY CHEMOREFLEX RESPONSES

The caudal pressor area (CPA) is a brainstem area located close to the spinal cord. The activation of the CPA increases sympathetic activity and mean arterial pressure (MAP) by mechanisms dependent on the commissural nucleus of the solitary tract (commNTS) and rostroventrolateral medulla, however, the signals that activate the CPA to produce these responses are still unknown. Therefore, in the present study, we investigated the activity of glutamatergic and GABAergic mechanisms from the CPA and commNTS in rats exposed to hypoxia and the effects of the inhibition of CPA neurons on cardiorespiratory responses to peripheral chemoreceptor activation with i.v. sodium cyanide (NaCN). Male Sprague-Dawley rats (250-280 g, n=5-8/group) were used. In conscious rats, most of the commNTS neurons (66 +/- 11%) and part of the CPA neurons (36 +/- 7%) activated by hypoxia (8% O2) were glutamatergic (contained VGLUT2mRNA). Small part of the neurons activated during hypoxia was GABAergic (contained GAD-67mRNA) in the commNTS (9 +/- 4%) or the CPA (6 +/- 2%). In urethane anesthetized rats, the inhibition of CPA neurons with bilateral injections of muscimol (GABA-A agonist, 2 mM) reduced baseline MAP, splanchnic sympathetic nerve discharge (SND) and phrenic nerve discharge (PND). Muscimol into the CPA also reduced by around 50% the pressor and sympathoexcitatory responses and the increase in PND to peripheral chemoreceptor activation with NaCN (50 mu g/kg i.v.), without changing sympathetic baroreflex responses. These data suggest that CPA mechanisms facilitate cardiorespiratory responses to peripheral chemoreflex activation. Immunohistochemistry results also suggest that at least part of the CPA mechanisms activated by hypoxia is glutamatergic. (C) 2011 IBRO. Published by Elsevier Ltd. All rights reserved.

Switching control of sympathetic activity from forebrain to hindbrain in chronic dehydration

We investigated the mechanisms responsible for increased blood pressure and sympathetic nerve activity (SNA) caused by 2-3 days dehydration (DH) both in vivo and in situ preparations. In euhydrated (EH) rats, systemic application of the AT(1) receptor antagonist Losartan and subsequent pre-collicular transection (to remove the hypothalamus) significantly reduced thoracic (t) SNA. In contrast, in DH rats, Losartan, followed by pre-collicular and pontine transections, failed to reduce tSNA, whereas transection at the medulla-spinal cord junction massively reduced tSNA. In DH but not EH rats, selective inhibition of the commissural nucleus tractus solitarii (cNTS) significantly reduced tSNA. Comparable data were obtained in both in situ and in vivo (anaesthetized/conscious) rats and suggest that following chronic dehydration, the control of tSNA transfers from supra-brainstem structures (e. g. hypothalamus) to the medulla oblongata, particularly the cNTS. As microarray analysis revealed up-regulation of AP1 transcription factor JunD in the dehydrated cNTS, we tested the hypothesis that AP1 transcription factor activity is responsible for dehydration-induced functional plasticity. When AP1 activity was blocked in the cNTS using a viral vector expressing a dominant negative FosB, cNTS inactivation was ineffective. However, tSNA was decreased after pre-collicular transection, a response similar to that seen in EHrats. Thus, the dehydration-induced switch in control of tSNA from hypothalamus to cNTS seems to be mediated via activation of AP1 transcription factors in the cNTS. If AP1 activity is blocked in the cNTS during dehydration, sympathetic activity control reverts back to forebrain regions. This unique reciprocating neural structure-switching plasticity between brain centres emphasizes the multiple mechanisms available for the adaptive response to dehydration.

Hemodynamic responses to aortic depressor nerve stimulation in conscious L-NAME-induced hypertensive rats

Durand MT, Castania JA, Fazan R Jr, Salgado MC, Salgado HC. Hemodynamic responses to aortic depressor nerve stimulation in conscious L-NAME-induced hypertensive rats. Am J Physiol Regul Integr Comp Physiol 300: R418-R427, 2011. First published November 24, 2010; doi: 10.1152/ajpregu.00463.2010.-The present study investigated whether baroreflex control of autonomic function is impaired when there is a deficiency in NO production and the role of adrenergic and cholinergic mechanisms in mediating reflex responses. Electrical stimulation of the aortic depressor nerve in conscious normotensive and nitro-L-arginine methyl ester (L-NAME)-induced hypertensive rats was applied before and after administration of methylatropine, atenolol, and prazosin alone or in combination. The hypotensive response to progressive electrical stimulation (5 to 90 Hz) was greater in hypertensive (-27 +/- 2 to -64 +/- 3 mmHg) than in normotensive rats (-17 +/- 1 to -46 +/- 2 mmHg), whereas the bradycardic response was similar in both groups (-34 +/- 5 to -92 +/- 9 and -21 +/- 2 to -79 +/- 7 beats/min, respectively). Methylatropine and atenolol showed no effect in the hypotensive response in either group. Methylatropine blunted the bradycardic response in both groups, whereas atenolol attenuated only in hypertensive rats. Prazosin blunted the hypotensive response in both normotensive (43%) and hypertensive rats (53%) but did not affect the bradycardic response in either group. Prazosin plus angiotensin II, used to restore basal arterial pressure, provided hemodynamic responses similar to those of prazosin alone. The triple pharmacological blockade abolished the bradycardic response in both groups but displayed similar residual hypotensive response in hypertensive (-13 +/- 2 to -27 +/- 2 mmHg) and normotensive rats (-10 +/- 1 to -25 +/- 3 mmHg). In conclusion, electrical stimulation produced a well-preserved baroreflex-mediated decrease in arterial pressure and heart rate in conscious L-NAME-induced hypertensive rats. Moreover, withdrawal of the sympathetic drive played a role in the reflex bradycardia only in hypertensive rats. The residual fall in pressure after the triple pharmacological blockade suggests the involvement of a vasodilatory mechanism unrelated to NO or deactivation of alpha(1)-adrenergic receptor.

Pimaradienoic acid inhibits vascular contraction and induces hypotension in normotensive rats

The present investigation was designed to investigate the effect of the diterpene ent-pimara-8(14),15-dien-19-oic acid (pimaradienoic acid, PA) on smooth muscle extracellular Ca2+ influx. To this end, the effect of PA on phenylephrine- and KCI-induced increases in cytosolic calcium concentration ([Ca2+](c)) measured by the variation in the ratio of fluorescence intensities (R340/ 380 nm) of Fura-2, was analysed. Whether bolus injection of PA could induce hypotensive responses in conscious normotensive rats was also evaluated. PA inhibited the contraction induced by phenylephrine (0.03 or 10 mu mol L-1) and KCI (30 or 90 mmol L-1) in endothelium-denuded rat aortic rings in a concentration dependent manner. Pre-treatment with PA (110, 100, 200 mu mol L-) attenuated the contraction induced by CaCl2 (0.5 nmol L(-)1 or 2.5 mmol L-1) in denuded rat aorta exposed to Ca2+- free medium containing phenylephrine (0.1 mu mol L-1) or KCI (30 mmol L-1). Interestingly, the inhibitory effect displayed by PA on CaCl2-induced contraction was more pronounced when KCI was used as the stimulant. Phenylephrine- and KCI-induced increases in (Ca2+,](c) were inhibited by PA. Similarly, verapamil, a Ca2+-channel blocker, also inhibited the increase in [Ca2+](c) induced by either phenylephrine or KCI. Finally, bolus injection of PA (1-15 mg kg(-1)) produced a dose-dependent decrease in mean arterial pressure in conscious normotensive rats. The results provide the first direct evidence that PA reduces vascular contractility by reducing extracellular Ca2+ influx through smooth muscle cellular membrane, a mechanism that could mediate the hypotensive response induced by this diterpene in normotensive rats.

Hypotensive effect of the nitrosyl ruthenium complex nitric oxide donor in renal hypertensive rats

We have described a new compound (trans-[RuCl([15]ane N(4))NO](2+)), which in vitro releases NO by the action of a reducing agent such as catecholamines. We investigated the effect of this NO donor in lowering the mean arterial pressure (MAP) in severe and moderate renal hypertensive 2K-1C rats. MAP was measured before and after intravenous in bolus injection of the compound in conscious 2K-1C and normotensive (2K) rats. In the hypertensive rats (basal 196.70 +/- 8.70 mmHg, n=5), the MAP was reduced in -34.25 +/- 13.50 mmHg(P < 0.05) 6 h after administration of 10 mmol/L/Kg of the compound in bolus. In normotensive rats the compound had no effect. We have also studied the effect of the injection of 0.1 mmol/L/Kg in normotensive (basal 118.20 +/- 11.25 mmHg, n = 4), moderate (basal 160.90 +/- 2.30 mmHg, n = 6), and severe hypertensive rats (basal 202.46 +/- 16.74 mmHg, n = 6). The compound at the dose of 0.1 mmol/L/Kg did not have effect (P> 0.05) on MAP of normotensive and moderate hypertensive rats. However, in the severe hypertensive rats (basal 202.46 +/- 16.70 mmHg, n = 6) there was a significant reduction on the MAP of -28.64 +/- 12.45 mmHg. The NO donor reduced the MAP of all hypertensive rats in the dose of 10 mmol/L/Kg and in the severe hypertensive rats at the dose of 0.1 mmol/L/Kg. The compound was not cytotoxic to the rat aortic vascular smooth muscle cells in the concentration of 0.1 mmol/LKg that produced the maximum relaxation. (C) 2008 Elsevier Inc. All rights reserved.

Gender-specific vascular effects elicited by chronic ethanol consumption in rats: a role for inducible nitric oxide synthase

Background and purpose: Epidemiological data suggest that the risk of ethanol-associated cardiovascular disease is greater in men than in women. This study investigates the mechanisms underlying gender-specific vascular effects elicited by chronic ethanol consumption in rats. Experimental approach: Vascular reactivity experiments using standard muscle bath procedures were performed on isolated thoracic aortae from rats. mRNA and protein for inducible NO synthase (iNOS) and for endothelial NOS (eNOS) was assessed by RT-PCR or western blotting, respectively. Key results: In male rats, chronic ethanol consumption enhanced phenylephrine-induced contraction in both endothelium-intact and denuded aortic rings. However, in female rats, chronic ethanol consumption enhanced phenylephrine-induced contraction only in endothelium denuded aortic rings. After pre-incubation of endothelium-intact rings with L-NAME, both male and female ethanol-treated rats showed larger phenylephrine-induced contractions in aortic rings, compared to the control group. Acetylcholine-induced relaxation was not affected by ethanol consumption. The effects of ethanol on responses to phenylephrine were similar in ovariectomized (OVX) and intact (non-OVX) female rats. In the presence of aminoguanidine, but not 7-nitroindazole, the contractions to phenylephrine in rings from ethanol-treated female rats were greater than that found in control tissues in the presence of the inhibitors. mRNA levels for eNOS and iNOS were not altered by ethanol consumption. Ethanol intake reduced eNOS protein levels and increased iNOS protein levels in aorta from female rats. Conclusions and implications: Gender differences in the vascular effects elicited by chronic ethanol consumption were not related to ovarian hormones but seemed to involve the upregulation of iNOS.

Bed nucleus of the stria terminalis alpha(1)-adrenoceptor modulates baroreflex cardiac component in unanesthetized rats

The bed nucleus of stria terminalis (BST) has a tonic modulating role on the baroreflex parasympathetic component. In the present study, we verified that local BST-adrenoceptors modulate baroreflex-evoked bradycardiac responses in unanesthetized rats. Bilateral microinjection of the selective alpha(1)-adrenoceptor antagonist WB4101 (15 nmol/100 nL) into the BST increased the gain of reflex bradycardia in response to mean arterial pressure increases caused by intravenous (i.v.) infusion of phenylephrine, suggesting that BST alpha(1)-adrenoceptors modulate baroreflex bradycardiac response. Bilateral microinjection of either the selective alpha(2)-adrenoceptor antagonist RX821002 (15 nmol/100 nL) or the non-selective beta-adrenoceptor antagonist propranolol (15 nmol/100 nL) into the BST had not affected baroreflex bradycardia. Animals were pretreated intravenously with the cholinergic muscarinic receptor antagonist homatropine methyl bromide (HMB, 1.5 mg/Kg) to test the hypothesis that activation of alpha(1)-adrenoceptors in the BST would modulate the baroreflex parasympathetic component. Baroreflex bradycardiac responses evoked before and after BST treatment with WB4101 were no longer different when rats were pretreated with HMB. These results suggest that parasympathetic activation accounts for the effects saw after BST pharmacological manipulation and ruling out the possibility of a sympathetic withdraw. In conclusion, our data point out that local alpha(1)-adrenoceptors mediate the BST tonic influence on the baroreflex bradycardiac response modulating parasympathetic cardiac activity. (C) 2008 Elsevier B.V. All rights reserved.

INTERMITTENT ACTIVATION OF PERIPHERAL CHEMORECEPTORS IN AWAKE RATS INDUCES Fos EXPRESSION IN ROSTRAL VENTROLATERAL MEDULLA-PROJECTING NEURONS IN THE PARAVENTRICULAR NUCLEUS OF THE HYPOTHALAMUS

Despite the well-established sympathoexcitation evoked by chemoreflex activation, the specific sub-regions of the CNS underlying such sympathetic responses remain to be fully characterized. In the present study we examined the effects of intermittent chemoreflex activation in awake rats on Fos-immunoreactivity (Fos-ir) in various subnuclei of the paraventricular nucleus of the hypothalamus (PVN), as well as in identified neurosecretory preautonomic PVN neurons. In response to intermittent chemoreflex activation, a significant increase in the number of Fos-ir cells was found in autonomic-related PVN subnuclei, including the posterior parvocellular, ventromedial parvocellular and dorsal-cap, but not in the neurosecretory magnocellular-containing lateral magnocellular subnucleus. No changes in Fos-ir following chemoreflex activation were observed in the anterior PVN subnucleus. Experiments combining Fos immunohistochemistry and neuronal tract tracing techniques showed a significant increase in Fos-ir in rostral ventrolateral medulla (RVLM)-projecting (PVN-RVLM), but not in nucleus of solitarii tract (NTS)-projecting PVN neurons. In summary, our results support the involvement of the PVN in the central neuronal circuitry activated in response to chemoreflex activation, and indicate that PVN-RVLM neurons constitute a neuronal substrate contributing to the sympathoexcitatory component of the chemoreflex. Published by Elsevier Ltd on behalf of IBRO.

The variability of baroreflex sensitivity in juvenile, spontaneously hypertensive rats

In this study the baroreflex sensitivity of conscious, juvenile, spontaneously hypertensive rats (SHRs) was compared. The study population consisted of 19 eight-week-old male SHRs. The baroreflex sensitivity was quantified as the derivative of the variation in heart rate (HR) and the variation of mean arterial pressure (baroreflex sensitivity = Delta HR/Delta MAP). MAP was manipulated with sodium nitroprusside (SNP) and phenylephrine (PHE), administered via an inserted cannula in the right femoral vein. The SHRs were divided into four groups: (1) low bradycardic baroreflex (LB) where the baroreflex gain (BG) was between 0 and 1 bpm/mmHg with PHE; (2) high bradycardic baroreflex (HB), where the BG was < -1 bpm/mmHg with PHE; (3) low tachycardic baroreflex (LT) where the BC was between 0 and 3 bpm/mmHg with SNP; (4) high tachycardic baroreflex (HT) where the BG was > 3 bpm/mmHg with SNP. We noted that 36.8% of the rats presented with an increased bradycardic reflex, while 27.8% demonstrated an attenuated tachycardic reflex. No significant alterations were noted regarding the basal MAP and HR. There were significant differences in the baroreflex sensitivity between SHRs in the same laboratory. One should be careful when interpreting studies employing the SHR as a research model.

Treatment with paracetamol, ketorolac or etoricoxib did not hinder alveolar bone healing: a histometric study in rats

Prostaglandins control osteoblastic and osteoclastic function under physiological or pathological conditions and are important modulators of the bone healing process. The non-steroidal anti-inflammatory drugs (NSAIDs) inhibit cyclooxygenase (COX) activity and consequently prostaglandins synthesis. Experimental and clinical evidence has indicated a risk for reparative bone formation related to the use of non-selective (COX-1 and COX-2) and COX-2 selective NSAIDs. Ketorolac is a non-selective NSAID which, at low doses, has a preferential COX-1 inhibitory effect and etoricoxib is a new selective COX-2 inhibitor. Although literature data have suggested that ketorolac can interfere negatively with long bone fracture healing, there seems to be no study associating etoricoxib with reparative bone formation. Paracetamol/acetaminophen, one of the first choices for pain control in clinical dentistry, has been considered a weak anti-inflammatory drug, although supposedly capable of inhibiting COX-2 activity in inflammatory sites. OBJECTIVE: The purpose of the present study was to investigate whether paracetamol, ketorolac and etoricoxib can hinder alveolar bone formation, taking the filling of rat extraction socket with newly formed bone as experimental model. MATERIAL AND METHODS: The degree of new bone formation inside the alveolar socket was estimated two weeks after tooth extraction by a differential point-counting method, using an optical microscopy with a digital camera for image capture and histometry software. Differences between groups were analyzed by ANOVA after confirming a normal distribution of sample data. RESULTS AND CONCLUSIONS: Histometric results confirmed that none of the tested drugs had a detrimental effect in the volume fraction of bone trabeculae formed inside the alveolar socket.

Cola beverage consumption delays alveolar bone healing: a histometric study in rats

Epidemiological studies have suggested that cola beverage consumption may affect bone metabolism and increase bone fracture risk. Experimental evidence linking cola beverage consumption to deleterious effects on bone is lacking. Herein, we investigated whether cola beverage consumption from weaning to early puberty delays the rate of reparative bone formation inside the socket of an extracted tooth in rats. Twenty male Wistar rats received cola beverage (cola group) or tap water (control group) ad libitum from the age of 23 days until tooth extraction at 42 days and euthanasia 2 and 3 weeks later. The neoformed bone volume inside the alveolar socket was estimated in semi-serial longitudinal sections using a quantitative differential point-counting method. Histological examination suggested a decrease in the osteogenic process within the tooth sockets of rats from both cola groups, which had thinner and sparser new bone trabeculae. Histometric data confirmed that alveolar bone healing was significantly delayed in cola-fed rats at three weeks after tooth extraction (ANOVA, p = 0.0006, followed by Tukey's test, p < 0.01). Although the results of studies in rats cannot be extrapolated directly to human clinical dentistry, the present study provides evidence that cola beverage consumption negatively affect maxillary bone formation.

Effects of detachment and repositioning of the medial pterygoid muscle on the growth of the maxilla and mandible of young rats

PURPOSE: To analyze the effects of detachment and repositioning of the medial pterygoid muscle on the growth of the maxilla and mandible of young rats through cephalometry. METHODS: Thirty one-month-old Wistar rats were used, distributed into three groups: experimental, sham-operated and control. In the experimental group, unilateral detachment and repositioning of the medial pterygoid muscle was performed. The sham-operated group only underwent surgical access, and the control group did not undergo any procedure. The animals were sacrificed at the age of three months. Their soft tissues were removed and the mandible was disarticulated. Radiographs of the skull in axial projection and the hemimandibles in lateral projection were obtained, and cephalometry was performed. The values obtained were subjected to statistical analyses among the groups and between the sides in each group. RESULTS: There were significant differences in the length of the mandible relative to the angular process in the experimental group and in the height of the mandibular body in the sham-operated group. CONCLUSION: The experimental detachment and repositioning of the medial pterygoid muscle during the growth period in rats affected the growth of the angle region, resulting in asymmetry of the mandible.