Cardiovascular effects of the microinjection of L-proline into the third ventricle or the paraventricular nucleus of the hypothalamus in unanesthetized rats

We investigated the cardiovascular effects of the microinjection of L-proline (L-Pro) into the third ventricle (3V) and its peripheral mechanisms. Different doses of L-Pro into the 3V caused dose-related pressor and bradycardiac responses. The pressor response to L-Pro injected into the 3V was potentiated by intravenous pretreatment with the ganglion blocker pentolinium (5 mg/kg), thus excluding any significant involvement of the sympathetic nervous system. Because the response to the microinjection of L-Pro into the 3V was blocked by intravenous pretreatment with the V1-vasopressin receptor antagonist dTyr(CH2)5(Me)AVP (50 mu g/kg), it is suggested that these cardiovascular responses are mediated by a vasopressin release. The pressor response to the microinjection of L-Pro into the 3V was found to be mediated by circulating vasopressin, so, given that the paraventricular nucleus of the hypothalamus (PVN) is readily accessible from the 3V, we investigated whether the PVN could be a site of action for the L-Pro microinjected in the 3V. The microinjection of L-Pro (0.033 mu moles/0.1 mu l) into the PVN caused cardiovascular responses similar to those of injection of the 3V and were also shown to be mediated by vasopressin release. In conclusion, these results show that the microinjection of L-Pro into the 3V causes pressor and bradycardiac responses that could involve stimulation of the magnocellular cells of the PVN and release of vasopressin into the systemic circulation. Also, because the microinjection of L-Pro into the PVN caused a pressor response, this is the first evidence of cardiovascular effects caused by its injection in a supramedullary structure. (c) 2012 Wiley Periodicals, Inc.

Purinergic transmission in the rostral but not caudal medullary raphe contributes to the hypercapnia-induced ventilatory response in unanesthetized rats

The medullary raphe (MR) is a putative central chemoreceptor site, contributing to hypercapnic respiratory responses elicited by changes in brain PCO2/pH. Purinergic mechanisms in the central nervous system appear to contribute to central chemosensitivity. To further explore the role of P2 receptors within the rostral and caudal MR in relation to respiratory control in room air and hypercapnic conditions, we performed microinjections of PPADS, a non-selective P2X antagonist, in conscious rats. Microinjections of PPADS into the rostral or caudal MR produced no changes in the respiratory frequency, tidal volume and ventilation in room air condition. The ventilatory response to hypercapnia was attenuated after microinjection of PPADS into the rostral but not in the caudal MR when compared to the control group (vehicle microinjection). These data suggest that P2X receptors in the rostral MR contribute to the ventilatory response to CO2, but do not participate in the tonic maintenance of ventilation under room air condition in conscious rats. (C) 2012 Elsevier B.V. All rights reserved.

Glutamatergic neurotransmission in the hypothalamus PVN on heart rate variability in exercise trained rats

The paraventricular nucleus of hypothalamus (PVN) is a well known site of integration for autonomic and cardiovascular responses, and the glutamate neurotransmitter plays an important role. The aim of our study was to evaluate the cardiovascular parameters and autonomic modulation by means of spectral analysis after ionotropic glutamate receptor inhibition in the PVN in conscious sedentary (S) or swimming trained (ST) rats. After exercise training protocol, adult male Wistar rats, instrumented with guide cannulae to PVN and artery and vein catheters were submitted to mean arterial pressure (MAP) and heart rate (HR) recording. At baseline, physical training induced a resting bradycardia (S: 379 +/- 3, ST: 349 +/- 2 bpm, P<0.05) and promoted adaptations in HRV characterized by an increase of HF in normalized values and a decrease of LF in absolute and normalized units compared with the sedentary group. Microinjection of kynurenic acid (KYNA) in the PVN of sedentary and trained rats promoted decreases in MAP and HR, but the decrease in HR was smaller in the trained animals (Delta HRS: -48 +/- 7, ST: -28 +/- 4 bpm, P<0.05). Furthermore, the differences in baseline parameters of pulse interval, found between sedentary and trained animals, disappeared after KYNA microinjection in the PVN. Our data suggest that the cardiovascular and autonomic adaptations to the heart induced by exercise training may involve glutamatergic mechanisms in the PVN. (C) 2012 Elsevier B.V. All rights reserved.

Effect of the Single or Combined Administration of Cocaine and Testosterone on Cardiovascular Function and Baroreflex Activity in Unanesthetized Rats

Abuse of cocaine and androgenic-anabolic steroids has become a serious public health problem. Despite reports of an increase in the incidence of simultaneous illicit use of these substances, potential toxic interactions between cocaine and androgenic-anabolic steroids in the cardiovascular system are unknown. In the present study, we investigated the effect of single or combined administration of testosterone and cocaine for 1 or 10 consecutive days on basal cardiovascular parameters, baroreflex activity, and hemodynamic responses to vasoactive agents in unanesthetized rats. Ten-day combined administration of testosterone and cocaine increased baseline arterial pressure. Changes in arterial pressure were associated with altered baroreflex activity and impairment of both hypotensive response to intravenous sodium nitroprusside and pressor effect of intravenous phenylephrine. Chronic single administration of either testosterone or cocaine did not affect baseline arterial pressure. However, testosterone-treated animals presented rest bradycardia, cardiac hypertrophy, alterations in baroreflex activity, and enhanced response to sodium nitroprusside. Repeated administration of cocaine affected baroreflex activity and impaired vascular responsiveness to both sodium nitroprusside and phenylephrine. One-day single or combined administration of the drugs did not affect any parameter investigated. In conclusion, the present results suggest a potential interaction between toxic effects of cocaine and testosterone on the cardiovascular activity. Changes in baseline arterial pressure after combined administration of these 2 drugs may result from alterations in baroreflex activity and impairment of vascular responsiveness to vasoactive agents.

Lateral septal area alpha(1)-and alpha(2)-adrenoceptors differently modulate baroreflex activity in unanaesthetized rats

The lateral septal area (LSA) is a limbic structure involved in autonomic, neuroendocrine and behavioural responses. An inhibitory influence of the LSA on baroreflex activity has been reported; however, the local neurotransmitter involved in this modulation is still unclear. In the present study, we verified the involvement of local LSA adrenoceptors in modulating cardiac baroreflex activity in unanaesthetized rats. Bilateral microinjection of the selective a1-adrenoceptor antagonist WB4101 (10 nmol in a volume of 100 nl) into the LSA decreased baroreflex bradycardia evoked by blood pressure increases, but had no effect on reflex tachycardia evoked by blood pressure decreases. Nevertheless, bilateral administration of the selective a2-adrenoceptor antagonist RX821002 (10 nmol in 100 nl) increased baroreflex tachycardia without affecting reflex bradycardia. Treatment of the LSA with a cocktail containing WB4101 and RX821002 decreased baroreflex bradycardia and increased reflex tachycardia. The non-selective beta-adrenoceptor antagonist propranolol (10 nmol in 100 nl) did not affect either reflex bradycardia or tachycardia. Microinjection of noradrenaline into the LSA increased reflex bradycardia and decreased the baroreflex tachycardic response, an opposite effect compared with those observed after double blockade of a1- and a2-adrenoceptors, and this effect of noradrenaline was blocked by local LSA pretreatment with the cocktail containing WB4101 and RX821002. The present results provide advances in our understanding of the baroreflex neural circuitry. Taken together, data suggest that local LSA a1- and a2-adrenoceptors modulate baroreflex control of heart rate differently. Data indicate that LSA a1-adrenoceptors exert a facilitatory modulation on baroreflex bradycardia, whereas local a2-adrenoceptors exert an inhibitory modulation on reflex tachycardia.

Central nitrergic system regulation of neuroendocrine secretion, fluid intake and blood pressure induced by angiotensin-II

Background: Nitric oxide (NO) synthesis has been described in several circumventricular and hypothalamic structures in the central nervous system that are implicated in mediating central angiotensin-II (ANG-II) actions during water deprivation and hypovolemia. Neuroendocrine and cardiovascular responses, drinking behavior, and urinary excretions were examined following central angiotensinergic stimulation in awake freely-moving rats pretreated with intracerebroventricular injections of N omega-nitro-L-arginine methyl ester (L-NAME, 40 mu g), an inhibitor of NO synthase, and L-arginine (20 ug), a precursor of NO. Results: Injections of L-NAME or ANG-II produced an increase in plasma vasopressin (VP), oxytocin (OT) and atrial natriuretic peptide (ANP) levels, an increase in water and sodium intake, mean arterial blood pressure and sodium excretion, and a reduction of urinary volume. L-NAME pretreatment enhanced the ANG-II response, while L-arginine attenuated VP and OT release, thirst, appetite for sodium, antidiuresis, and natriuresis, as well as pressor responses induced by ANG-II. Discussion and conclusion: Thus, the central nitrergic system participates in the angiotensinergic responses evoked by water deprivation and hypovolemia to refrain neurohypophysial secretion, hydromineral balance, and blood pressure homeostasis.

Influence of Plasma Protein Binding on Pharmacodynamics: Estimation of In Vivo Receptor Affinities of beta Blockers Using a New Mechanism-Based PK-PD Modelling Approach

The objective of this investigation was to examine in a systematic manner the influence of plasma protein binding on in vivo pharmacodynamics. Comparative pharmacokinetic-pharmacodynamic studies with four beta blockers were performed in conscious rats, using heart rate under isoprenaline-induced tachycardia as a pharmacodynamic endpoint. A recently proposed mechanism-based agonist-antagonist interaction model was used to obtain in vivo estimates of receptor affinities (K(B),(vivo)). These values were compared with in vitro affinities (K(B),(vitro)) on the basis of both total and free drug concentrations. For the total drug concentrations, the K(B),(vivo) estimates were 26, 13, 6.5 and 0.89 nM for S(-)-atenolol, S(-)-propranolol, S(-)-metoprolol and timolol. The K(B),(vivo) estimates on the basis of the free concentrations were 25, 2.0, 5.2 and 0.56 nM, respectively. The K(B),(vivo)-K(B),(vitro) correlation for total drug concentrations clearly deviated from the line of identity, especially for the most highly bound drug S(-)-propranolol (ratio K(B),(vivo)/K(B),(vitro) similar to 6.8). For the free drug, the correlation approximated the line of identity. Using this model, for beta-blockers the free plasma concentration appears to be the best predictor of in vivo pharmacodynamics. (C) 2008 Wiley-Liss, Inc. and the American Pharmacists Association J Pharm Sci 98:3816-3828, 2009

Effect of the duration of daily aerobic physical training on cardiac autonomic adaptations

The present study has investigated in conscious rats the influence of the duration of physical training sessions on cardiac autonomic adaptations by using different approaches; 1) double blockade with methylatropine and propranolol; 2) the baroreflex sensitivity evaluated by alternating bolus injections of phenylephrine and sodium nitroprusside; and 3) the autonomic modulation of HRV in the frequency domain by means of spectral analysis. The animals were divided into four groups: one sedentary group and three training groups submitted to physical exercise (swimming) for 15, 30, and 60 min a day during 10 weeks. All training groups showed similar reduction in intrinsic heart rate (IHR) after double blockade with methylatropine and propranolol. However, only 30-min and 60-min physical training presented an increase in the vagal autonomic component for determination of basal heart rate (HR) in relation to group sedentary. Spectral analysis of HR showed that the 30-min and 60-min physical training presented the reduction in low-frequency oscillations (LF = 0.20-0.75 Hz) and the increase in high-frequency oscillations (HF = 0.75-2.5 Hz) in normalized units. These both groups only showed an increased baroreflex sensitivity to tachycardiac responses in relation to group sedentary, however when compared, the physical training of 30-min exhibited a greater gain. In conclusion, cardiac autonomic adaptations, characterised by the increased predominance of the vagal autonomic component, were not proportional to the duration of daily physical training sessions. In fact, 30-minute training sessions provided similar cardiac autonomic adaptations, or even more enhanced ones, as in the case of baroreflex sensitivity compared to 60-minute training sessions. (C) 2010 Elsevier B.V. All rights reserved.

The Essential Oil of Eucalyptus tereticornis and its Constituents, alpha- and beta-Pinene, Show Accelerative Properties on Rat Gastrointestinal Transit

The essential oil of Eucalyptus tereticornis (EOET) has pharmacological activities but their effects on the gastrointestinal tract are yet unknown. It possesses alpha- and beta-pinene as minor constituents, isomers largely used as food or drink additives. In this work, we studied their actions on gut motility. After feeding with a liquid test meal, conscious rats received perorally EOET, alpha-, or beta-pinene, and the fractional dye retention was determined. EOET and its constituents decreased the gastric retention. In anesthetized rats, pinenes increased gastric tonus, while enhancing the meal progression in the small intestine of conscious rats. Both alpha- and beta-pinene contracted gastric strips in vitro but relaxed the duodenum. Conversely, EOET relaxed both the gastric and duodenal strips. In conclusion, EOET accelerates the gastric emptying of liquid, and part of its action is attributed to the contrasting effects induced by alpha- and beta-pinene on the gut.

Validation of ACB in vitro and in vivo as a biomagnetic method for measuring stomach contraction

Background The aim of this study was to validate a biomagnetic method (alternate current biosusceptometry, ACB) for monitoring gastric wall contractions in rats. Methods In vitro data were obtained to establish the relationship between ACB and the strain-gauge (SG) signal amplitude. In vivo experiments were performed in pentobarbital-anesthetized rats with SG and magnetic markers previously implanted under the gastric serosa or after ingestion of magnetic material. Gastric motility was quantified from the tracing amplitudes and frequency profiles obtained by Fast Fourier Transform. Key Results The correlation between in vitro signal amplitudes was strong (R = 0.989). The temporal cross-correlation coefficient between the ACB and SG signal amplitude was higher (P < 0.0001) in the postprandial (88.3 +/- 9.1 V) than in the fasting state (31.0 +/- 16.9 V). Irregular signal profiles, low contraction amplitudes, and smaller signal-to-noise ratios explained the poor correlation between techniques for fasting-state recordings. When a magnetic material was ingested, there was also strong correlation in the frequency and signal amplitude and a small phase-difference between the techniques. The contraction frequencies using ACB were 0.068 +/- 0.007 Hz (postprandial) and 0.058 +/- 0.007 Hz (fasting) (P < 0.002) and those using SG were 0.066 +/- 0.006 Hz (postprandial) and 0.059 +/- 0.008 Hz (fasting) (P < 0.005). Conclusions & Inferences In summary, ACB is reliable for monitoring gastric wall contractions using both implanted and ingested magnetic materials, and may serve as an accurate and sensitive technique for gastrointestinal motility studies.

Focal CO2 dialysis in raphe obscurus does not stimulate ventilation but enhances the response to focal CO2 dialysis in the retrotrapezoid nucleus

Simultaneous inhibition of the retrotrapezoid nucleus (RTN) and raphe obscurus (ROb) decreased the systemic CO2 response by 51%, an effect greater than inhibition of RTN (- 24%) or ROb (0%) alone, suggesting that ROb modulates chemoreception by interaction with the RTN (19). We investigated this interaction further by simultaneous dialysis of artificial cerebrospinal fluid equilibrated with 25% CO2 in two probes located in or adjacent to the RTN and ROb in conscious adult male rats. Ventilation was measured in a whole body plethysmograph at 30 C. There were four groups (n = 5): 1) probes correctly placed in both RTN and ROb (RTN-ROb); 2) one probe correctly placed in RTN and one incorrectly placed in areas adjacent to ROb (RTN-peri-ROb); 3) one probe correctly placed in ROb and one probe incorrectly placed in areas adjacent to RTN (peri-RTN-ROb); and 4) neither probe correctly placed (peri-RTN-peri-ROb). Focal simultaneous acidification of RTN-ROb significantly increased ventilation ((V) over dot E) up to 22% compared with baseline, with significant increases in both breathing frequency and tidal volume. Focal acidification of RTN-peri-ROb increased (V) over dot E significantly by up to 15% compared with baseline. Focal acidification of ROb and peri-RTN had no significant effect. The simultaneous acidification of regions just outside the RTN and ROb actually decreased (V) over dot E by up to 11%. These results support a modulatory role for the ROb with respect to central chemoreception at the RTN.

Serotonergic neurons in the nucleus raphe obscurus contribute to interaction between central and peripheral ventilatory responses to hypercapnia

Serotonergic (5-HT) neurons in the nucleus raphe obscurus (ROb) are involved in the respiratory control network. However, it is not known whether ROb 5-HT neurons play a role in the functional interdependence between central and peripheral chemoreceptors. Therefore, we investigated the role of ROb 5-HT neurons in the ventilatory responses to CO(2) and their putative involvement in the central-peripheral CO(2) chemoreceptor interaction in unanaesthetised rats. We used a chemical lesion specific for 5-HT neurons (anti-SERT-SAP) of the ROb in animals with the carotid body (CB) intact or removed (CBR). Pulmonary ventilation (V (E)), body temperature and the arterial blood gases were measured before, during and after a hypercapnic challenge (7% CO(2)). The lesion of ROb 5-HT neurons alone (CB intact) or the lesion of 5-HT neurons of ROb+CBR did not affect baseline V (E) during normocapnic condition. Killing ROb 5-HT neurons (CB intact) significantly decreased the ventilatory response to hypercapnia (p < 0.05). The reduction in CO(2) sensitivity was approximately 15%. When ROb 5-HT neurons lesion was combined with CBR (anti-SERT-SAP+CBR), the V (E) response to hypercapnia was further decreased (-31.2%) compared to the control group. The attenuation of CO(2) sensitivity was approximately 30%, and it was more pronounced than the sum of the individual effects of central (ROb lesion; -12.3%) or peripheral (CBR; -5.5%) treatments. Our data indicate that ROb 5-HT neurons play an important role in the CO(2) drive to breathing and may act as an important element in the central-peripheral chemoreception interaction to CO(2) responsiveness.

Propyretic role of the locus coeruleus nitric oxide pathway

Nitric oxide has been reported to modulate fever in the brain. However, the sites where NO exerts this modulation remain somewhat unclear. Locus coeruleus (LC) neurons express not only nitric oxide synthase (NOS) but also soluble guanylyl cyclase (sGC). In the present study, we evaluated in vivo and ex vivo the putative role of the LC NO-cGMP pathway in fever. To this end, deep body temperature was measured before and after pharmacological modulations of the pathway. Moreover, nitrite/nitrate (NOx) and cGMP levels in the LC were assessed. Conscious rats were microinjected within the LC with a non-selective NOS inhibitor (NG-monomethyl-l-arginine acetate), a NO donor (NOC12), a sGC inhibitor (1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one) or a cGMP analogue (8-bromo-cGMP) and injected intraperitoneally with endotoxin. Inhibition of NOS or sGC before endotoxin injection significantly increased the latency to the onset of fever. During the course of fever, inhibition of NOS or sGC attenuated the febrile response, whereas microinjection of NOC12 or 8-bromo-cGMP increased the response. These findings indicate that the LC NO-cGMP pathway plays a propyretic role. Furthermore, we observed a significant increase in NOx and cGMP levels, indicating that the febrile response to endotoxin is accompanied by stimulation of the NO-cGMP pathway in the LC.

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.

Influence of angiotensin II receptor subtypes of the paraventricular nucleus on the physiological responses induced by angiotensin II injection into the medial septal area

OBJECTIVE: We determined the effects of losartan and PD 123319 (antagonists of the AT1 and AT2 angiotensin receptors, respectively), and [Sar¹, Ala8] ANG II (a relatively peptide antagonist of angiotensin receptors) injected into the paraventricular nucleus (PVN) on water and 3% NaCl intake, and the diuretic, natriuretic, and pressor effects induced by administration of angiotensin II (ANG II) into the medial septal area (MSA) of conscious rats. METHODS: Holtzman rats were used . Animals were anesthetized with tribromoethanol (20 mg) per 100 grams of body weight, ip. A stainless steel guide cannula was implanted into the MSA and PVN. All drugs were injected in 0.5-mul volumes for 10-15 seconds. Seven days after brain surgery, water and 3% NaCl intake, urine and sodium excretion, and arterial blood pressure were measured. RESULTS: Losartan (40 nmol) and [Sar¹, Ala8] ANG II (40 nmol) completely eliminated whereas PD 123319 (40 nmol) partially blocked the increase in water and sodium intake and the increase in arterial blood pressure induced by ANG II (10 nmol) injected into the MSA. The PVN administration of PD 123319 and [Sar¹, Ala8] ANG II blocked whereas losartan attenuated the diuresis and natriuresis induced by MSA administration of ANG II. CONCLUSION: MSA involvement with PVN on water and sodium homeostasis and arterial pressure modulation utilizing ANGII receptors is suggested.