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International audience; Major depression is an independent risk factor for the development of cardiovascular disease. This impact of depression on vascular function seems to be mediated by the endothelial dysfunction, defined as an impairment of endothelium-dependent vasorelaxation, which represents a reliable predictor of atherosclerosis and has been regularly found to be associated with depression. This study aimed at investigating aortic vascular reactivity in mice submitted to the unpredictable chronic mild stress (UCMS) procedure, a reliable model of depression. The results confirm the effectiveness of the UCMS procedure to induce neuroendocrine, physical and behavioral depression-like alterations as well as a significant decrease of acetylcholine-induced vasorelaxation without any effect on phenylephrine-induced vasoconstriction. In this study, we reveal an altered vascular reactivity in an animal model of depression, demonstrating an endothelial dysfunction reminiscent to the one found in depressed patients.

International audience; BACKGROUND: Moderate physical activity enhances endothelium-dependent vasorelaxation. Whether the frequency of exercise affects endothelial function is unclear. The purpose of this study was to investigate the effects of various frequencies of training on endothelium-dependent vasorelaxation. DESIGN: Male Wistar rats were trained for 8 weeks on a treadmill at various frequencies [1 (Ex1), 3 (Ex3) or 5 days/week (Ex5)] and compared with age-matched sedentary animals (SED). A control group allowed us to assess endothelial function before the exercise protocol. Rings of thoracic aorta were precontracted with phenylephrine. RESULTS: Endothelium-independent relaxation elicited by sodium nitroprusside was similar in all groups. The maximal response elicited by acetylcholine (ACh) was not different between groups, whereas pD2 values (-logEC50, EC50 being the concentration of ACh that elicited 50% of the maximal response) significantly correlated with frequency of training. nitro-L-arginine methyl ester (L-NAME) reduced the relaxation elicited by 10(-7) mol/l ACh or higher in control and all trained groups, and by 10(-6) mol/l ACh or higher in SED group. Indomethacin inhibited the vasodilating response to 10(-7) mol/l ACh or higher in control, SED and Ex1 groups, and to 10(-8) mol/l or more in Ex3 and Ex5 animals. Tetraethylammonium attenuated the response to 10(-6) mol/l ACh or higher in control and SED groups and to 10(-7) mol/l or more in all trained animals. CONCLUSION: This data suggest that decreased ACh-induced vasorelaxation after physical inactivity may result from impairment of endothelial nitric oxide synthase, prostacyclin and endothelium-derived hyperpolarizing factor pathways. This effect is prevented by training in a frequency-dependent manner.

International audience; We have investigated the possible role of alpha1-adrenoreceptors in regulating the germination of progenitor cells cultured from embryonic rat neocortex. High binding levels of the alpha1-selective radioligand 3[H]prazosin were detected in the forebrain of the rat embryo at E13, and the greatest density of binding sites was localized to the ventricular and subventricular zones. Catecholamine-containing axon terminals were present in these zones in the same period. Germinal neuroepithelial cells retained specific 3[H]prazosin binding in culture. Approximately 25% of cells in culture displayed complex intracellular Ca2+ transients in response to phenylephrine, many of which were abolished with the alpha1B antagonist, chloroethylclonidine. Cultures exhibited concentration-dependent catecholamine stimulation of DNA synthesis mediated by alpha1 receptors in serum-limited conditions. Neuroepithelial cells were labelled via their ventricular processes by intraventricular injection of Fast blue in E13 embryos prior to transfer of the neocortex to dissociated cell culture. Many of labelled cells were present in culture in germinal foci. Some cells which migrated from these foci underwent apoptosis, as determined by TUNEL in situ hybridization. During a transitory period of up to 48 h in culture, alpha1-adrenoreceptor activation by phenylephrine or noradrenaline increased the number of surviving cells. Apoptosis was observed in vivo in both ventricular and subventricular zones of the neocortex from E13 to E15 in increasing numbers. We propose that both the supply of noradrenaline to forebrain germinal cells, and the expression of alpha1-adrenoreceptors on their surface could act to determine whether they die or continue to proliferate.

Gain-of-function mutations in the human WNK1 ( with-no-lysine[K]1 ) gene are responsible for a monogenic form of arterial hypertension, and WNK1 polymorphisms have been associated with common essential hypertension. The role of WNK1 in renal ionic reabsorption has been established, but no investigation of its possible influence on vascular tone, an essential determinant of blood pressure, has been performed until now. WNK1 complete inactivation in the mouse is embryonically lethal. We, thus, examined in Wnk1 +/− haploinsufficient adult mice whether WNK1 could regulate in vivo vascular tone and whether this was correlated with blood pressure variation. Wnk1 +/− mice displayed a pronounced decrease in blood pressure responses in vivo and in vascular contractions ex vivo following α 1 -adrenergic receptor activation with no change in basal blood pressure and renal function. We also observed a major loss of the pressure-induced contractile (myogenic) response in Wnk1 +/− arteries associated with a specific alteration of the smooth muscle cell contractile function. These alterations in vascular tone were associated with a decreased phosphorylation level of the WNK1 substrate SPAK (STE20/SPS1-related proline/alanine-rich kinase) and its target NKCC1 (Na + -K + -2Cl − cotransporter 1) in Wnk1 +/− arteries. Our study identifies a novel and major role for WNK1 in maintaining in vivo blood pressure and vasoconstriction responses specific to α 1 -adrenergic receptor activation. Our findings uncover a vascular signaling pathway linking α 1 -adrenergic receptors and pressure to WNK1, SPAK, and NKCC1 and may, thus, significantly broaden the comprehension of the regulatory mechanisms of vascular tone in arterial hypertension.

International audience; AIMS: Patients affected by Brugada syndrome (BrS) are at risk of sudden cardiac death specifically at rest, when vagal tone is high. The aim of our study was to assess whether a phenylephrine injection, which provokes a baroreflex stimulation, could induce modification of the ST segment elevation and ventricular arrhythmias. METHODS AND RESULTS: Baroreflex test was performed with the administration of phenylephrine (2 microg/kg) to four highly symptomatic patients in a setting fully equipped for cardiac resuscitation. Phenylephrine injection induced a deep vagal stimulation with a decrease in the mean heart rate from 75 +/- 7 to 50 +/- 8 bpm and an increase in the mean systolic blood pressure from 141 +/- 14 to 204 +/- 46 mmHg. ST segment elevation was not modified and no ventricular arrhythmias were induced during the test. CONCLUSION: Although phenylephrine injection induced a major alpha-adrenergic vasoconstriction followed by an arterial baroreflex, this test failed to provoke ventricular arrhythmias or modification of the ST segment elevation in BrS patients.

Objective— Notch3, a member of the evolutionary conserved Notch receptor family, is primarily expressed in vascular smooth muscle cells. Genetic studies in human and mice revealed a critical role for Notch3 in the structural integrity of distal resistance arteries by regulating arterial differentiation and postnatal maturation. Methods and Results— We investigated the role of Notch3 in vascular tone in small resistance vessels (tail and cerebral arteries) and large (carotid) arteries isolated from Notch3 -deficient mice using arteriography. Passive diameter and compliance were unaltered in mutant arteries. Similarly, contractions to phenylephrine, KCl, angiotensin II, and thromboxane A2 as well as dilation to acetylcholine or sodium nitroprusside were unaffected. However, Notch3 deficiency induced a dramatic reduction in pressure-induced myogenic tone associated with a higher flow (shear stress)-mediated dilation in tail and cerebral resistance arteries only. Furthermore, RhoA activity and myosin light chain phosphorylation, measured in pressurized tail arteries, were significantly reduced in Notch3 KO mice. Additionally, myogenic tone inhibition by the Rho kinase inhibitor Y27632 was attenuated in mutant tail arteries. Conclusions— Notch3 plays an important role in the control of vascular mechano-transduction, by modulating the RhoA/Rho kinase pathway, with opposite effects on myogenic tone and flow-mediated dilation in the resistance circulation.

International audience; INTRODUCTION: Dynamic ventricular repolarization assessed by QT/RR slopes studies the effects of modifications in cardiac repolarization independently of variations in RR interval (RR). The effects of changes in sympathetic and vagal activity on the QT/RR slope are controversial. We tested the hypothesis that sympathoexcitation is an important determinant of the QT/RR slope. METHODS AND RESULTS: We compared the effects of a reflex sympathetic activation in response to hypoxia, to the direct effects of the infusion of the beta-adrenergic agent dobutamine, on the QTa (apex) and QTe (end)/RR slopes. Dobutamine was titrated to obtain similar increases in cardiac output than with hypoxia. Cardiac vagal activity was estimated by rMSSD and pNN50. In a second group of healthy subjects, we assessed the effect of a reflex cardiac vagal activation in response to phenylephrine infusion on the same variables. We observed a similar increase in QTa and QTe slopes during hypoxia and dobutamine (both P < 0.017 vs. normoxia), despite divergent changes in cardiac vagal activity, as rMSSD and pNN50 decreased with hypoxia compared to normoxia (P < 0.001) but increased during dobutamine infusion compared to hypoxia (P < 0.017). In contrast, these slopes did not change during the rises in rMSSD and pNN50 elicited by phenylephrine (P > 0.7). CONCLUSION: Beta-adrenergic stimulation induces comparable increases in the QT/RR slopes than hypoxia, but in the presence of a larger cardiac vagal activity. Vagal cardiac activation by phenylephrine does not change the QT slopes. This reveals that the sympathetic system is an important determinant of QT/RR dynamicity in healthy men.

Small increases in extracellular Ca 2+ dilate isolated blood vessels. In the present study, the possibility that a vascular, extracellular Ca 2+ -sensing receptor (CaSR) could mediate these vasodilator actions was investigated. Novel ligands that interact with the CaSR were used in microelectrode recordings from rat isolated mesenteric and porcine coronary arteries. The major findings were that (1) raising extracellular Ca 2+ or adding calindol, a CaSR agonist, produced concentration-dependent hyperpolarizations of vascular myocytes, actions attenuated by Calhex 231, a negative allosteric modulator of CaSR. (2) Calindol-induced hyperpolarizations were inhibited by the intermediate conductance, Ca 2+ -sensitive K + (IK Ca ) channel inhibitors, TRAM-34, and TRAM-39. (3) The effects of calindol were not observed in the absence of endothelium. (4) CaSR mRNA and protein were present in rat mesenteric arteries and in porcine coronary artery endothelial cells. (5) CaSR and IK Ca proteins were restricted to caveolin-poor membrane fractions. We conclude that activation of vascular endothelial CaSRs opens endothelial cell IK Ca channels with subsequent myocyte hyperpolarization. The endothelial cell CaSR may have a physiological role in the control of arterial blood pressure.

International audience; The modulation by 5-HT2 receptors in the nucleus tractus solitarius of the reflex bradycardia evoked by stimulation of peripheral baroreceptors and cardiopulmonary chemoreceptors, and their possible functional interactions with local NMDA receptors, were investigated in pentobarbital- and urethane-anaesthetized rats, respectively. Microinjection of the 5-HT2 receptor agonist, 2,5-dimethoxy-4-iodoamphetamine (0.1-0.5 pmol), into the nucleus tractus solitarius elicited a dose-dependent hypotension and bradycardia. Bilateral microinjections at the same site of a subthreshold dose of 2,5-dimethoxy-4-iodoamphetamine (0.05 pmol) significantly enhanced the aforementioned reflex-evoked bradycardiac responses. In contrast, local bilateral microinjections of the NMDA receptor antagonist, 2-amino-5-phosphonopentanoic acid (500 and 1000 pmol), reduced, in a dose-dependent manner, both reflex-evoked responses. The facilitatory effect of 2,5-dimethoxy-4-iodoamphetamine upon these reflex-evoked bradycardiac responses was prevented by prior local microinjection of low doses of either the selective 5-HT2 receptor antagonist, ketanserin (10 pmol), or 2-amino-5-phosphonopentanoic acid (100 pmol), which, on their own, did not affect the reflex-associated bradycardia. These data suggest that 5-HT2 receptors within the nucleus tractus solitarius participate in a facilitatory modulation of the reflex control of heart rate, probably through functional interactions with local NMDA receptors.

International audience; In the present study we evaluated the effects of bilateral microinjection of muscimol (a GABA(A) receptor agonist) and baclofen (a GABA(B) receptor agonist) into the lateral commissural nucleus tractus solitarii (NTS) of awake rats on the gain of the baroreflex (BG) activated by a short duration (10-15 s) infusion of phenylephrine (Phe, 2.5 microg/0.05 ml, i.v.). Microinjection of muscimol (50 pmol/50 nl, n=8) into the NTS produced a significant increase in baseline mean arterial pressure ((MAP) 122+/-6 vs. 101+/-2 mmHg), no changes in baseline heart rate (HR) and a reduction in BG (-1.59+/-0. 1 vs. -0.69+/-0.1 beats/mmHg). Microinjection of baclofen (6.25 pmol/50 nl, n=6) into the NTS also produced a significant increase in baseline MAP (138+/-5 vs. 103+/-2 mmHg), no changes in baseline HR and a reduction in BG (-1.54+/-0.3 vs. -0.53+/-0.2 beats/mmHg). Considering that the reduction in BG could be secondary to the increase in MAP in response to microinjection of muscimol (n=6) or baclofen (n=7) into the NTS, in these two groups of rats we brought the MAP back to baseline by infusion of sodium nitroprusside (NP, 3.0 microg/0.05 ml, i.v.). Under these conditions, we verified that the BG remained significantly reduced after muscimol (-1.49+/-0.2 vs. -0.35+/-0.2 beats/mmHg) and after baclofen (-1.72+/-0.2 vs. -0.33+/-0.2 beats/mmHg) when compared to control. Reflex tachycardia was observed during the normalization of MAP by NP infusion and, in order to prevent the autonomic imbalance from affecting BG, we used another group of rats treated with atenolol (5 mg/kg, i.v.), a beta1 receptor antagonist. In rats previously treated with atenolol and submitted to NP infusion, we verified that BG remained reduced after microinjection of muscimol or baclofen into the NTS. The data show that activation of GABA(A) and GABA(B) receptors, independently of the changes in the baseline MAP or HR, inhibited the neurons of the NTS involved in the parasympathetic component of the baroreflex.

Our aim was to determine whether the cardiovascular neurones in the rostro-ventrolateral medulla (CV-RVLM neurones) were involved in the sympathoexcitation induced by stimulation of 5-HT3 receptors in the region of the nucleus tractus solitarii (NTS). Experiments were performed in pentobarbitone-anaesthetized rats, artificially ventilated and paralysed with pancuronium bromide. Using extracellular recordings, different types of RVLM neurones were characterized: cardiovascular (CV), ventilation-related and baroreflex-insensitive (unidentified) neurones. The CV-RVLM cells were further subdivided into three populations according to their axonal conduction velocities: A (1.2 ± 0.1 m s−1), B (2.5 ± 0.2 m s−1) and C (6.8 ± 1.1 m s−1). Only the CV-RVLM neurones of the A and B categories were partially inhibited (−30 %) by a hypotensive dose (2.5 μg kg−1 i.v.) of clonidine. Microinjections into the region of the commissural NTS of 1-(m-chlorophenyl)-biguanide (CPBG, 2 nmol), a selective 5-HT3 receptor agonist, elicited an increase in both lumbar sympathetic nerve discharge (SND) and arterial pressure. In addition, this treatment produced a marked excitation of CV-RVLM neurones of the A and B categories, without affecting those of the C type, as well as ventilation-related and unidentified RVLM cells. The activity of the CV neurones in the caudo-ventrolateral part of the medulla oblongata (CV-CVLM) was not modified by 5-HT3 receptor stimulation in the NTS. Prior intra-NTS microinjections of ondansetron (300 pmol, a selective 5-HT3 receptor antagonist) into the region of the commissural NTS prevented the excitation of A and B CV-RVLM neurones induced by CPBG. Intracarotid administration of saline saturated with CO2 (chemoreceptor activation) elicited both an increase in the SND and an excitation of the clonidine-insensitive CV-RVLM neurones of the C type, without affecting A and B neurones. In conclusion, the sympathoexcitation elicited following 5-HT3 receptor stimulation in the region of the commissural NTS of pentobarbitone-anaesthetized rats seems to result from the excitation of two different pools of clonidine-sensitive CV-RVLM neurones. These neurones are apparently not involved in the sympathetic component of the chemoreceptor reflex. The rostro-ventrolateral medulla (RVLM) and nucleus tractus solitarii (NTS) are critically involved in the reflex control of sympathetic activity (Guyenet, Filtz & Donaldson, 1987; Sun & Guyenet, 1987; Spyer, 1994). The RVLM contains neurones that receive a number of inputs both peripheral and central in origin that influence sympathetic nerve activity (Sun & Guyenet, 1987; Spyer, 1994). Previous reports have described two populations of cardiovascular (CV)-RVLM neurones that project to the thoracic spinal cord (Brown & Guyenet, 1985; Sun & Guyenet, 1985). The first population consists of clonidine-sensitive cells with slow-conducting axons. The second population corresponds to cells that do not respond to hypotensive doses of clonidine, and are characterized by a much higher conduction velocity (Sun & Guyenet, 1986). The NTS is the site of termination of afferent fibres arising from arterial baroreceptors (baroreflex), cardiopulmonary chemoreceptors (Bezold-Jarisch reflex) and carotid chemoreceptors (chemoreflex) (Palkovits & Zaborsky, 1977; Kalia & Mesulam, 1980; Jordan & Spyer, 1986). For baro- and Bezold-Jarisch reflexes, which seem to share the same integrating mechanisms and pathways in the brain (Verberne & Guyenet, 1992), second-order neurones project from the NTS to the caudal ventrolateral part of the medulla (CVLM) (Gordon 1987; Guyenet et al. 1987; Verberne & Guyenet, 1992). These neurones exert an excitatory action on the GABAergic CVLM neurones that project to the RVLM where they inhibit the CV-RVLM neurones of this pressor area (Brown & Guyenet, 1985; Sun & Guyenet, 1985; Jeske, Reis & Milner, 1995). Some of the CV-RVLM neurones additionally constitute an efferent link in the sympathetic component of the chemoreflex (Guyenet & Brown, 1986; Sun & Reis, 1995). Indeed, it has been demonstrated that some NTS chemosensitive neurones have axonal projections to the RVLM (Koshiya & Guyenet, 1996). Within the NTS, serotonin (5-hydroxytryptamine, 5-HT) seems to be involved in the reflex control of blood pressure. Studies in both anaesthetized and conscious rats have shown that 5-HT2 receptor stimulation in the NTS elicits the typical CV responses of baroreceptor activation (Merahi, Orer & Laguzzi, 1992a; Callera, Bonagamba, Sevoz, Laguzzi & Machado, 1997a). On the other hand, stimulation of NTS 5-HT3 receptors elicits a chemoreceptor-like increase in arterial pressure and lumbar sympathetic activity (Merahi, Orer, Laporte, Gozlan, Hamon & Laguzzi, 1992b; Callera, Sevoz, Laguzzi & Machado, 1997b). In experiments aimed at analysing the mechanism (s) responsible for this sympatho-excitatory effect, we observed that stimulation of 5-HT3 receptors in the NTS did not inhibit the sympathetic component of the baroreflex (Nosjean, Franc & Laguzzi, 1995). Accordingly, it can be inferred that 5-HT3 receptor-mediated sympathoexcitation is not the consequence of the disruption of tonic baroreceptor sympathoinhibitory messages. However, our finding is compatible with the idea that CV-RVLM neurones may be involved in the sympathoexcitatory effect of 5-HT3 receptor stimulation in the NTS. In other experiments, we also observed that prior microinjections of pressor (nanomolar) doses of 5-HT3 receptor agonists into the NTS did not increase the sympathetic chemoreflex response (Sevoz, Callera, Machado, Hamon & Laguzzi, 1997). However, this observation does not rule out the possibility that under some physiological conditions, 5-HT released in the NTS may excite the NTS-RVLM sympathetic chemoreflex pathway (Koshiya & Guyenet, 1996), and the CV-RVLM neurones involved in this reflex. Indeed, as previously observed with the pressor chemoreflex response (Sun & Reis, 1995), we recently found that the microinjection of kynurenic acid, a glutamate receptor antagonist, into the RVLM blocked the pressor effects elicited by 5-HT3 receptor stimulation in this area (Sevoz, Hamon & Laguzzi, 1996b). In order to elucidate the possible role of CV-RVLM neurones in the sympathetic response to 5-HT3 receptor stimulation in the NTS, we have analysed the effects of intra-NTS microinjections of a potent and selective 5-HT3 receptor agonist, 1-(m-chlorophenyl)-biguanide (CPBG), as well as the effects of chemoreflex activation, on the activity of the different categories of RVLM neurones. In addition, the possible effects of intra-NTS administration of CPBG on the CV-CVLM neurones were also investigated.

International audience; The selectivity of rat auditory cortex neurons for pure tone frequency was studied during and after ionophoretic application (5-40 nA) of noradrenaline in urethane-anaesthetized rats. The dominant effect induced by noradrenaline was a significant decrease in spontaneous (93/268 cells) and evoked activity (133/268 cells) which outlasted the application. In the whole population of cells (n = 268) the signal-to-noise ratio, computed using as the signal either the mean evoked response or the response at the best frequency, was unchanged during noradrenaline application. It was significantly increased only for cells showing significantly decreased spontaneous activity, and was significantly decreased for cells showing increased spontaneous activity. Frequency selectivity was significantly increased for the whole population during and after noradrenaline application. It was also significantly increased for cells showing significantly decreased evoked activity, and was significantly decreased for cells showing increased evoked activity. The noradrenaline-induced inhibition was not blocked by propranolol (beta antagonist); it was blocked by prazosin (alpha1 antagonist) and partly mimicked by phenylephrine (alpha1 agonist). GABA, which also inhibited spontaneous and evoked activity, slightly increased the signal-to-noise ratio and significant increased frequency selectivity. However, when noradrenaline was ejected in the presence of bicuculline at doses that were able to block GABAergic inhibition, the inhibitory effects of noradrenaline on spontaneous and evoked activity were still observed. The possible function of noradrenaline-induced inhibitions in sensory cortices is briefly discussed.