Chronic Metformin Administration Does Not Alter Carotid Sinus Nerve Activity in Control Rats.

Metformin is a glucose-lowering, insulin-sensitizing drug that is commonly used in the treatment of type 2 diabetes (T2D). In the last decade, the carotid body (CB) has been described as a metabolic sensor implicated in the regulation of glucose homeostasis, being CB dysfunction crucial for the development of metabolic diseases, such as T2D. Knowing that metformin could activate AMP-activated protein kinase (AMPK) and that AMPK has been described to have an important role in CB hypoxic chemotransduction, herein we have investigated the effect of chronic metformin administration on carotid sinus nerve (CSN) chemosensory activity in basal and hypoxic and hypercapnic conditions in control animals. Experiments were performed in male Wistar rats subjected to 3 weeks of metformin (200 mg/kg) administration in the drinking water. The effect of chronic metformin administration was tested in spontaneous and hypoxic (0% and 5% O2) and hypercapnic (10% CO2) evoked CSN chemosensory activity. Metformin administration for 3 weeks did not modify the basal CSN chemosensory activity in control animals. Moreover, the CSN chemosensory response to intense and moderate hypoxia and hypercapnia was not altered by the chronic metformin administration. In conclusion, chronic metformin administration did not modify chemosensory activity in control animals.

Endogenous hydrogen sulfide maintains eupnea in an in situ arterially perfused preparation of rats.

Hydrogen sulfide (H2S) is constitutively generated in the human body and works as a gasotransmitter in synaptic transmission. In this study, we aimed to evaluate the roles of endogenous H2S in generating eupnea at the respiratory center. We employed an in situ arterially perfused preparation of decerebrated rats and recorded the central respiratory outputs. When the H2S-producing enzyme cystathionine ß-synthase (CBS) was inhibited, respiration switched from the 3-phase eupneic pattern, which consists of inspiration, postinspiration, and expiration, to gasping-like respiration, which consists of inspiration only. On the other hand, when H2S synthesis was inhibited via cystathionine γ-lyase (CSE) or when H2S synthesis was activated via CBS, eupnea remained unchanged. These results suggest that H2S produced by CBS has crucial roles in maintaining the neuronal network to generate eupnea. The mechanism of respiratory pattern generation might be switched from a network-based system to a pacemaker cell-based system in low H2S conditions.

Irradiation of carotid baroreceptor with low-intensity pulsed ultrasound exerts different metabolic protection in perirenal, epididymal white adipose tissue and interscapular brown adipose tissue of obese rats.

This study was designed to clarify whether the irradiation of carotid baroreceptor (CB) with low-intensity pulsed ultrasound (LIPUS) protects against obesity by rebalancing the autonomic nervous system (ANS). Obesity was induced using a high-fat diet (HFD) for 8 weeks in Sprague-Dawley rats. Irradiation with LIPUS was daily (20 minutes a day) applied to the right CB. In our study, LIPUS significantly ameliorated metabolic disorders in obese rats. LIPUS partly restored norepinephrine (NE) and acetylcholine (ACH) levels in the perirenal white adipose tissue (PWAT), epididymal white adipose tissue (EWAT), interscapular brown adipose tissue (IBAT), and plasma of obese rats. LIPUS partially rectified the dysregulated AMP-activated protein kinase (AMPK)/peroxisome proliferator-activated receptor (PPAR) α/É£ pathway in the PWAT, EWAT, and IBAT of obese rats. PPARγ and PPARγ target genes respond more sensitively to HFD and LIPUS in PWAT and EWAT than in IBAT. NE, ACH, uncoupling protein-1, phosphorylated AMPK, PPARα, and PPARα target genes respond more sensitively to HFD and LIPUS in IBAT than in PWAT and EWAT. Conclusion: LIPUS irradiation of CB exerts different metabolic protection in PWAT, EWAT, and IBAT by rebalancing the ANS and rectifying the AMPK/PPARα/É£ pathway in obese rats.

Vesicular glutamate transporter 2-immunoreactive afferent nerve terminals in rat carotid sinus baroreceptors.

Sensory nerve endings respond to various stimuli and subsequently transmit afferent informations to central nervous system, but their responsibility has been suggested to be modulated by glutamate. In the present study, we examined the immunohistochemical localization of vesicular glutamate transporter 1 (vGLUT1) and vGLUT2 in baroreceptor nerve endings immunoreactive for P2X2 and P2X3 purinoceptors in the rat carotid sinus by immunohistochemistry of whole-mount preparations with confocal scanning laser microscopy. P2X3-immunoreactive flat leaf-like axon terminals were immunoreactive to vGLUT2, but not to vGLUT1. Among members of the soluble N-ethylmaleimide-sensitive factor attachment protein receptor complex, immunoreactivities for synaptosomal-associated protein, 25 kDa, Syntaxin1, and vesicle-associated membrane protein 2 (VAMP2) were localized in P2X2- and P2X3-immunoreactive axon terminals. Punctate immunoreactive products for VAMP2 and vGLUT2 were co-localized in axon terminals. These results suggest that vGLUT2 is localized in P2X3-immunoreactive baroreceptor terminals in the carotid sinus, and these terminals may release glutamate by exocytosis in order to modulate baroreceptor function in the carotid sinus.

Decoding Neural Metabolic Markers From the Carotid Sinus Nerve in a Type 2 Diabetes Model.

Recent studies showed that the carotid sinus nerve (CSN) and the sympathetic nervous system (SNS) are overactivated in type 2 diabetes and that restoring the correct CSN neural activity can re-establish the proper metabolism. However, a robust characterization of the relationship between CSN and SNS neural activities and metabolism in type 2 diabetes is still missing. Here, we investigated the relationship between neural activity of CSN and SNS in control rats and in rats with diet-induced type 2 diabetes and the animal condition during metabolic challenges. We found that the diabetic condition can be discriminated on the basis of CSN and SNS neural activities due to a high-frequency shift in both spectra. This shift is suppressed in the SNS in case of CSN denervation, confirming the role of CSN in driving sympathetic overactivation in type 2 diabetes. Interestingly, the Inter-Burst-Intervals (IBIs) calculated from CSN bursts strongly correlate with perturbations in glycaemia levels. This finding, held for both control and diabetic rats, indicates the possibility of detecting metabolic information from neural recordings even in pathological conditions. Our results suggest that CSN activity could serve as a marker to monitor glycaemic alterations and, therefore, it could be used for closed-loop control of CSN neuromodulation. This paves the way to the development of novel and effective bioelectronic therapies for type 2 diabetes.

GABAA receptor, KATP channel and L-type Ca2+ channel is associated with facilitation effect of H2S on the baroreceptor reflex in spontaneous hypertensive rats.

BACKGROUND: We aimed to investigate whether the facilitating effect of H2S on the baroreceptor reflex is associated with the GABAA receptor, KATP channel and L-type Ca2+ channel pathway. METHODS: Spontaneously hypertensive rats (SHRs) and Wistar Kyoto (WKY) rats were used to investigate the facilitating effect of H2S on the baroreceptor reflex by perfusing the isolated carotid sinus. The mechanism by which H2S facilitated the baroreceptor reflex was determined by using Bay K8644 (an agonist of calcium channels), glibenclamide (Gli, a KATP channel blocker), and picrotoxin (PIC, a blocker of γ-aminobutyric acid [GABA]A receptor). RESULTS: As compared with WKY rats, SHRs showed impaired baroreceptor reflex sensitivity, as demonstrated by a right and upward shift of the functional curve for the intrasinus pressure-arterial blood pressure relation. H2S perfusion (25, 50, or 100 µmol/L) dose-dependently ameliorated the impaired sensitivity of the baroreceptor reflex. Bay K8644 (500 nmol/L), Gli (20 µmol/L) and PIC (50 µmol/L) all prevented H2S ameliorating the impaired baroreceptor reflex. CONCLUSIONS: H2S facilitating the baroreceptor reflex might be associated with activating the GABAA receptor, opening the KATP channel, and closing the L-type Ca2+ channel. These areas should provide new targets for preventing and treating hypertension.

Distribution and morphology of baroreceptors in the rat carotid sinus as revealed by immunohistochemistry for P2X3 purinoceptors.

The morphological characteristics of baroreceptors in the rat carotid sinus were reevaluated by whole-mount preparations with immunohistochemistry for P2X3 purinoceptors using confocal scanning laser microscopy. Immunoreactive nerve endings for P2X3 were distributed in the internal carotid artery proximal to the carotid bifurcation, particularly in the region opposite the carotid body. Some pre-terminal axons in nerve endings were ensheathed by myelin sheaths immunoreactive for myelin basic protein. Pre-terminal axons ramified into several branches that extended two-dimensionally in every direction. The axon terminals of P2X3-immunoreactive nerve endings were flat and leaf-like in shape, and extended hederiform- or knob-like protrusions in the adventitial layer. Some axons and axon terminals with P2X3 immunoreactivity were also immunoreactive for P2X2, and axon terminals were closely surrounded by terminal Schwann cells with S100 or S100B immunoreactivity. These results revealed the detailed morphology of P2X3-immunoreactive nerve endings and suggested that these endings respond to a mechanical deformation of the carotid sinus wall with their flat leaf-like terminals.

Peripheral versus central effect of intravenous moxonidine on rat carotid sinus baroreflex-mediated sympathetic arterial pressure regulation.

AIMS: Moxonidine is a centrally acting antihypertensive agent with a selectivity to I1-imidazoline receptors higher than that to α2-adrenergic receptors. The present study aimed to quantify a peripheral effect of moxonidine on carotid sinus baroreflex-mediated sympathetic arterial pressure (AP) regulation separately from its central effect. MAIN METHODS: In eight anesthetized Wistar rats, changes in efferent sympathetic nerve activity (SNA) and AP in response to a carotid sinus pressure input were compared before and during an intravenous administration of moxonidine (100µgkg-1 bolus followed by a continuous infusion at 200µg·kg-1·h-1). KEY FINDINGS: Moxonidine significantly narrowed the range of the AP response (55.3±5.8 to 39.1±6.1mmHg, P<0.05) without changing the minimum AP (77.2±6.4 to 80.7±5.1mmHg, not significant). In the neural arc, moxonidine reduced the minimum SNA (56.6±5.9 to 29.7±6.2%, P<0.05) without affecting the range of the SNA response (45.3±5.5 to 40.2±5.0%, not significant). In the peripheral arc, moxonidine increased the intercept (3.0±8.5 to 51.1±7.2mmHg, P<0.01) and reduced the slope (1.28±0.06 to 0.92±0.15mmHg/%, P<0.05). SIGNIFICANCE: Moxonidine increased AP at any given SNA, suggesting that the peripheral vasoconstrictive effect is stronger than generally recognized. The peripheral vasoconstrictive effect of moxonidine may partly offset the vasodilatory effect attained by centrally-mediated sympathoinhibition.

Endogenous Hydrogen Sulfide Enhances Carotid Sinus Baroreceptor Sensitivity by Activating the Transient Receptor Potential Cation Channel Subfamily V Member 1 (TRPV1) Channel.

BACKGROUND: We aimed to investigate the regulatory effects of hydrogen sulfide (H2S) on carotid sinus baroreceptor sensitivity and its mechanisms. METHODS AND RESULTS: Male Wistar-Kyoto rats and spontaneously hypertensive rats (SHRs) were used in the experiment and were given an H2S donor or a cystathionine-ß-synthase inhibitor, hydroxylamine, for 8 weeks. Systolic blood pressure and the cystathionine-ß-synthase/H2S pathway in carotid sinus were detected. Carotid sinus baroreceptor sensitivity and the functional curve of the carotid baroreceptor were analyzed using the isolated carotid sinus perfusion technique. Effects of H2S on transient receptor potential cation channel subfamily V member 1 (TRPV1) expression and S-sulfhydration were detected. In SHRs, systolic blood pressure was markedly increased, but the cystathionine-ß-synthase/H2S pathway in the carotid sinus was downregulated in comparison to that of Wistar-Kyoto rats. Carotid sinus baroreceptor sensitivity in SHRs was reduced, demonstrated by the right and upward shift of the functional curve of the carotid baroreceptor. Meanwhile, the downregulation of TRPV1 protein was demonstrated in the carotid sinus; however, H2S reduced systolic blood pressure but enhanced carotid sinus baroreceptor sensitivity in SHRs, along with TRPV1 upregulation in the carotid sinus. In contrast, hydroxylamine significantly increased the systolic blood pressure of Wistar-Kyoto rats, along with decreased carotid sinus baroreceptor sensitivity and reduced TRPV1 protein expression in the carotid sinus. Furthermore, H2S-induced enhancement of carotid sinus baroreceptor sensitivity of SHRs could be amplified by capsaicin but reduced by capsazepine. Moreover, H2S facilitated S-sulfhydration of TRPV1 protein in the carotid sinus of SHRs and Wistar-Kyoto rats. CONCLUSIONS: H2S regulated blood pressure via an increase in TRPV1 protein expression and its activity to enhance carotid sinus baroreceptor sensitivity.

Oxidative stress augments chemoreflex sensitivity in rats exposed to chronic intermittent hypoxia.

Chronic exposure to intermittent hypoxia (CIH) elicits plasticity of the carotid sinus and phrenic nerves via reactive oxygen species (ROS). To determine whether CIH-induced alterations in ventilation, metabolism, and heart rate are also dependent on ROS, we measured responses to acute hypoxia in conscious rats after 14 and 21 d of either CIH or normoxia (NORM), with or without concomitant administration of allopurinol (xanthine oxidase inhibitor), combined allopurinol plus losartan (angiotensin II type 1 receptor antagonist), or apocynin (NADPH oxidase inhibitor). Carotid body nitrotyrosine production was measured by immunohistochemistry. CIH produced an increase in the ventilatory response to acute hypoxia that was virtually eliminated by all three pharmacologic interventions. CIH caused a robust increase in carotid body nitrotyrosine production that was greatly attenuated by allopurinol plus losartan and by apocynin but unaffected by allopurinol. CIH caused a decrease in metabolic rate and a reduction in hypoxic bradycardia. Both of these effects were prevented by allopurinol, allopurinol plus losartan, and apocynin.

Neural regulation of hypoxia-inducible factors and redox state drives the pathogenesis of hypertension in a rodent model of sleep apnea.

Obstructive sleep apnea (OSA) is one of the most common causes of hypertension in western societies. OSA causes chronic intermittent hypoxia (CIH) in specialized O2-sensing glomus cells of the carotid body. CIH generates increased reactive oxygen species (ROS) that trigger a feedforward mechanism in which increased intracellular calcium levels ([Ca(2+)]i) trigger increased HIF-1α synthesis and increased HIF-2α degradation. As a result, the normal homeostatic balance between HIF-1α-dependent prooxidant and HIF-2α-dependent antioxidant enzymes is disrupted, leading to further increases in ROS. Carotid body sensory nerves project to the nucleus tractus solitarii, from which the information is relayed via interneurons to the rostral ventrolateral medulla in the brain stem, which sends sympathetic neurons to the adrenal medulla to stimulate the release of epinephrine and norepinephrine, catecholamines that increase blood pressure. At each synapse, neurotransmitters trigger increased [Ca(2+)]i, HIF-1α:HIF-2α, and Nox2:Sod2 activity that generates increased ROS levels. These responses are not observed in other regions of the brain stem that do not receive input from the carotid body or signal to the sympathetic nervous system. Thus sympathetic nervous system homeostasis is dependent on a balance between HIF-1α and HIF-2α, disruption of which results in hypertension in OSA patients.

Coumestrol inhibits carotid sinus baroreceptor activity by cAMP/PKA dependent nitric oxide release in anesthetized male rats.

Phytoestrogens could offer multiple beneficial effects on the cardiovascular system. Here, we have examined the effects of coumestrol (CMT) on carotid baroreceptors activity (CBA) and the possible mechanisms in male rats. The functional parameters of carotid baroreceptors were measured by recording sinus nerve afferent discharge in anesthetized male rats with perfused isolated carotid sinus. The levels of protein expression were determined by using ELISA and Western blotting. CMT (1 to 100µmolL(-1)) inhibited CBA, which shifted the functional curve of the carotid baroreceptor to the right and downward, with a marked decrease in the peak slope and the peak integral value of carotid sinus nerve discharge in a concentration dependent manner. These effects were not blocked by a specific estrogen receptor antagonist ICI 182,780, but were completely abolished by nitric oxide (NO) synthase inhibitor l-NAME (N(G)-nitro-l-arginine methyl ester). Furthermore, a NO donor, SIN-1(3-morpholion-sydnon-imine), could potentiate these inhibitory effects of CMT. CMT stimulated the phosphorylation of Ser(1176)-eNOS (endothelial nitric oxide synthase) in a dose-dependent manner in carotid bifurcation tissue over a perfusion period of 15min. The rapid activation of eNOS by CMT was blocked by a highly selective PKA (protein kinase A) inhibitor H89. In addition, inhibition of PI3K (phosphatidylinositol-3-kinase) and ERK (extracellular signal-regulated kinase) pathways had no effect on eNOS activation by CMT. CMT inhibited CBA via eNOS activation and NO synthesis. These effects were mediated by the cAMP/PKA pathway and were unrelated to the estrogenic effect.

Discharges of aortic and carotid sinus baroreceptors during spontaneous motor activity and pharmacologically evoked pressor interventions.

Our laboratory has demonstrated that the cardiomotor component of aortic baroreflex is temporarily inhibited at the onset of spontaneous motor activity in decerebrate cats, without altering carotid sinus baroreflex. A reason for this dissociation may be attributed to a difference in the responses between aortic nerve activity (AoNA) and carotid sinus nerve activity (CsNA) during spontaneous motor activity. The stimulus-response curves of AoNA and CsNA against mean arterial blood pressure (MAP) were compared between the pressor interventions evoked by spontaneous motor activity and by intravenous administration of phenylephrine or norepinephrine, in which the responses in heart rate (HR) were opposite (i.e., tachycardia vs. baroreflex bradycardia), despite the identical increase in MAP of 34-40 mmHg. In parallel to the pressor response, mean AoNA and CsNA increased similarly by 78-81 and by 88 % of the baseline control, respectively, irrespective of whether the pressor response was evoked by spontaneous motor activity or by a pharmacological intervention. The slope of the stimulus-response curve of the mean AoNA became greater (P < 0.05) during spontaneous motor activity as compared to the pharmacological intervention. On the other hand, the stimulus-response curve of the mean CsNA and its slope were equal (P > 0.05) between the two pressor interventions. Furthermore, the slopes of the stimulus-response curves of both diastolic AoNA and CsNA (defined as the minimal value within a beat) exhibited a greater increase during spontaneous motor activity. All differences in the slopes of the stimulus-response curves were abolished by restraining HR at the intrinsic cardiac frequency. In conclusion, mean mass activities of both aortic and carotid sinus baroreceptors are able to encode the beat-by-beat changes in MAP not only at rest but also during spontaneous motor activity and spontaneous motor activity-related reduction of aortic baroreceptor activity is denied accordingly.

Changes in glutamate receptor subunits within the medulla in goats after section of the carotid sinus nerves.

The mechanisms which contribute to the time-dependent recovery of resting ventilation and the ventilatory CO2 chemoreflex after carotid body denervation (CBD) are poorly understood. Herein we tested the hypothesis that there are time-dependent changes in the expression of specific AMPA, NMDA, and/or neurokinin-1 (NK1R) receptors within respiratory-related brain stem nuclei acutely or chronically after CBD in adult goats. Brain stem tissues were collected acutely (5 days) or chronically (30 days) after sham or bilateral CBD, immunostained with antibodies targeting AMPA (GluA1 or GluA2), NMDA (GluN1), or NK-1 receptors, and optical density (OD) compared. Physiological measurement confirmed categorization of each group and showed ventilatory effects consistent with bilateral CBD (Miller et al. J Appl Physiol 115: 1088-1098, 2013). Acutely after CBD, GluA1 OD was unchanged or slightly increased, but GluA2 and GluN1 OD were reduced 15-30% within the nucleus tractus solitarius (NTS) and in other medullary respiratory nuclei. Chronically after CBD, GluA1 was reduced (P < 0.05) within the caudal NTS and in other nuclei, but there was significant recovery of GluA2 and GluN1 OD. NK1 OD was not significantly different from control after CBD. We conclude that the initial decrease in GluA2 and GluN1 after CBD likely contributes to hypoventilation and the reduced CO2 chemoreflex. The partial recovery of ventilation and the CO2 chemoreflex after CBD parallel a time-dependent return of these receptors to near control levels but likely depend upon additional initiating and maintenance factors for neuroplasticity.

Chronic caffeine intake in adult rat inhibits carotid body sensitization produced by chronic sustained hypoxia but maintains intact chemoreflex output.

Sustained hypoxia produces a carotid body (CB) sensitization, known as acclimatization, which leads to an increase in carotid sinus nerve (CSN) activity and ensuing hyperventilation greater than expected from the prevailing partial pressure of oxygen. Whether sustained hypoxia is physiological (high altitude) or pathological (lung disease), acclimatization has a homeostatic implication because it tends to minimize hypoxia. Caffeine, the most commonly ingested psychoactive drug and a nonselective adenosine receptor antagonist, alters CB function and ventilatory responses when administered acutely. Our aim was to investigate the effect of chronic caffeine intake on CB function and acclimatization using four groups of rats: normoxic, caffeine-treated normoxic, chronically hypoxic (12% O2, 15 days), and caffeine-treated chronically hypoxic rats. Caffeine was administered in drinking water (1 mg/ml). Caffeine ameliorated ventilatory responses to acute hypoxia in normoxic animals without altering the output of the CB (CSN neural activity). Caffeine-treated chronically hypoxic rats exhibited a decrease in the CSN response to acute hypoxia tests but maintained ventilation compared with chronically hypoxic animals. The findings related to CSN neural activity combined with the ventilatory responses indicate that caffeine alters central integration of the CB input to increase the gain of the chemoreflex and that caffeine abolishes CB acclimatization. The putative mechanisms involved in sensitization and its loss were investigated: expression of adenosine receptors in CB (A(2B)) was down-regulated and that in petrosal ganglion (A(2A)) was up-regulated in caffeine-treated chronically hypoxic rats; both adenosine and dopamine release from CB chemoreceptor cells was increased in chronic hypoxia and in caffeine-treated chronic hypoxia groups.

Baroreflex modulation during sleep and in obstructive sleep apnea syndrome.

This review focuses on the complex integration between cardiovascular reflexes and central autonomic influences controlling physiological sleep-dependent changes in arterial blood pressure and heart rate. A brief introduction on the anatomic and functional organization of the arterial baroreflex and the methods available to assess its function in humans is followed by an analysis of the functional interaction between autonomic nervous system and sleep mechanisms at the highest levels of brain organization. An insight into these interactions is important to shed light on the physiopathology of the most frequent complications of obstructive sleep apnea syndrome, such as sustained arterial hypertension, and excessive daytime sleepiness.

Hypoxic intensity: a determinant for the contribution of ATP and adenosine to the genesis of carotid body chemosensory activity.

Excitatory effects of adenosine and ATP on carotid body (CB) chemoreception have been previously described. Our hypothesis is that both ATP and adenosine are the key neurotransmitters responsible for the hypoxic chemotransmission in the CB sensory synapse, their relative contribution depending on the intensity of hypoxic challenge. To test this hypothesis we measured carotid sinus nerve (CSN) activity in response to moderate and intense hypoxic stimuli (7 and 0% O(2)) in the absence and in the presence of adenosine and ATP receptor antagonists. Additionally, we quantified the release of adenosine and ATP in normoxia (21% O(2)) and in response to hypoxias of different intensities (10, 5, and 2% O(2)) to study the release pathways. We found that ZM241385, an A(2) antagonist, decreased the CSN discharges evoked by 0 and 7% O(2) by 30.8 and 72.5%, respectively. Suramin, a P(2)X antagonist, decreased the CSN discharges evoked by 0 and 7% O(2) by 64.3 and 17.1%, respectively. Simultaneous application of both antagonists strongly inhibited CSN discharges elicited by both hypoxic intensities. ATP release by CB increased in parallel to hypoxia intensity while adenosine release increased preferably in response to mild hypoxia. We have also found that the lower the O(2) levels are, the higher is the percentage of adenosine produced from extracellular catabolism of ATP. Our results demonstrate that ATP and adenosine are key neurotransmitters involved in hypoxic CB chemotransduction, with a more relevant contribution of adenosine during mild hypoxia, while vesicular ATP release constitutes the preferential origin of extracellular adenosine in high-intensity hypoxia.

Angiotensin II evokes sensory long-term facilitation of the carotid body via NADPH oxidase.

We previously reported that reactive oxygen species generated by NADPH oxidase 2 (Nox2) induces sensory plasticity of the carotid body, manifested as a progressive increase in baseline sensory activity or sensory long-term facilitation (sLTF). ANG II, a peptide generated within the carotid body, is a potent activator of Nox2. In the present study, we tested the hypothesis that ANG II evokes sLTF of the carotid body via Nox2 activation. Experiments were performed on carotid bodies ex vivo from adult rats and mice. Sensory activity was recorded from the carotid sinus nerve. Repetitive (5 times for 30 s each at 5-min intervals), but not continuous (for 150 s), application of 60 pM ANG II evoked robust sLTF of the carotid body. ACh, ATP, substance P, and KCl, when applied repetitively, stimulated the carotid body but did not evoke sLTF. Reactive oxygen species levels increased in response to repetitive applications of ANG II, and this effect was blocked by apocynin, an inhibitor of Nox2, as well as losartan, an angiotensin type 1 (AT(1)) receptor antagonist. Losartan, apocynin, and 4-(2-aminoethyl)benzenesulfonyl fluoride prevented ANG II-induced sLTF, which was absent in mice deficient in gp91(phox), the catalytic subunit of the Nox2 complex. These results demonstrate that repetitive application of ANG II induces sLTF of the carotid body via activation of Nox2 by AT(1) receptors.

Reflexes from pulmonary arterial baroreceptors in dogs: interaction with carotid sinus baroreceptors.

In contrast to the reflex vasodilatation occurring in response to stimulation of baroreceptors in the aortic arch, carotid sinuses and coronary arteries, stimulation of receptors in the wall of pulmonary arteries results in reflex systemic vasoconstriction. It is rare for interventions to activate only one reflexogenic region, therefore we investigated how these two types of reflexes interact. In anaesthetized dogs connected to cardiopulmonary bypass, reflexogenic areas of the carotid sinuses, aortic arch and coronary arteries and the pulmonary artery were subjected to independently controlled pressures. Systemic perfusion pressure (SPP) measured in the descending aorta (constant flow) provided an index of systemic vascular resistance. In other experiments, sympathetic efferent neural activity was recorded in fibres dissected from the renal nerve (RSNA). Physiological increases in pulmonary arterial pressure (PAP) induced significant increases in SPP (+39.1 ± 10.4 mmHg) and RSNA (+17.6 ± 2.2 impulses s(−1)) whereas increases in carotid sinus pressure (CSP) induced significant decreases in SPP (−42.6 ± 10.8 mmHg) and RSNA (−42.8 ± 18.2 impulses s(−1)) (P < 0.05 for each comparison; paired t test). To examine possible interactions, PAP was changed at different levels of CSP in both studies. With CSP controlled at 124 ± 2 mmHg, the threshold, 'set point' and saturation pressures of the PAP­SPP relationship were higher than those with CSP at 60 ± 1 mmHg; this rightward shift was associated with a significant decrease in the reflex gain. Similarly, increasing CSP produced a rightward shift of the PAP­RSNA relationship, although the effect on reflex gain was inconsistent. Furthermore, the responses to changes in CSP were influenced by setting PAP at different levels; increasing the level of PAP from 5 ± 1 to 33 ± 3 mmHg significantly increased the set point and threshold pressures of the CSP­SPP relationship; the reflex gain was not affected. These results indicate the existence of interaction between pulmonary arterial and carotid sinus baroreceptor reflexes; physiological and pathological states that alter the stimulus to one may alter the reflex responses from the other.

Hypoxia-induced hypothermia mediated by noradrenaline and nitric oxide in the rostromedial preoptic area.

Central nitric oxide (NO) has an important role in hypothermia induced by hypoxia as well as in that elicited by noradrenaline (NA) microinjected into the rostromedial preoptic area (POA) of the hypothalamus. Here, I tested the hypothesis that activation of adrenoceptors and NO in the rostromedial POA is involved in hypoxia-induced hypothermia in urethane-chloralose-anesthetized, neuromuscularly blocked, artificially ventilated rats. Hypoxic ventilation (10% O2-90% N2, 5 min) evoked an increase in the tail skin temperature and a decrease in the colonic temperature, though these changes occurred at 30 s to 7 min after returning the rats to ventilation with room air. These responses were eliminated by prior bilateral transection of the carotid sinus nerves, but not by bilateral cervical vagotomy, suggesting the involvement of activated carotid chemoreceptors in the hypoxic ventilation-induced hypothermia. Such responses were also greatly attenuated by the microinjection of an NO synthase (NOS) inhibitor, NG-monomethyl-L-arginine (L-NMMA, 25 nmol), but not by that of its inactive enantiomer, NG-monomethyl-D-arginine (D-NMMA, 25 nmol), into the NA-sensitive, hypothermia-inducing site in the rostromedial POA. Pretreatment with the α1-adrenoceptor blocker prazosin (50 pmol), but not vehicle saline, also greatly attenuated the hypoxic ventilation-induced heat loss responses. These results suggest that this hypoxia-induced hypothermia was mediated, at least in part, by activation of α1-adrenoceptors and NOS in the rostromedial POA.