RESUMEN

Background: Hypertension is characterized by upregulation of the renin-angiotensin system, increased blood-brain barrier (BBB) permeability, microglia activation within autonomic nuclei, and an intense sympathoexcitation. There is no information on the interplay of these events during the development of neurogenic hypertension. We sought to identify the interaction and time-course changes of Ang II availability, barrier dysfunction, microglia activation, and autonomic imbalance within autonomic areas during the development of neurogenic hypertension. Methods: Sequential changes of hemodynamic/autonomic parameters, BBB permeability, microglia structure/density (IBA-1), and angiotensin II (Ang II) immunofluorescence were evaluated within the paraventricular hypothalamic nucleus, nucleus of the solitary tract, and rostral ventrolateral medulla of Wistar and spontaneously hypertensive rats (SHRs) aged 4 weeks, 5 weeks, 6 weeks, 8 weeks, and 12 weeks. The somatosensory cortex and hypoglossal nucleus were also analyzed as non-autonomic control areas. Results: Increased brain Ang II availability (4th-5th week) was the first observed change, followed by the incipient BBB leakage and increased microglia density (6th week). From the 5th-6th weeks on, BBB leakage, Ang II, and IBA-1 densities increased continuously, allowing a parallel increase in both Ang II-microglia colocalization and the transition of microglial cells from highly ramified in the basal surveillant condition (4th-5th week) to shorter process arbors, fewer endpoints, and enlarged soma in the disease-associate condition (6th week to the 12th week). Simultaneously with increased Ang II-microglia colocalization and microglia morphologic phenotypic changes, sympathetic activity and pressure variability increased, autonomic control deteriorated, and blood pressure increased. These responses were not specific for autonomic nuclei but also occurred at a lower magnitude in the somatosensory cortex and hypoglossal nucleus, indicating the predominance of hypertension-induced effects on autonomic areas. No changes were observed in age-matched controls where Ang II density did not change. Conclusion: Brain Ang II density is the initial stimulus to drive coordinated changes in BBB permeability and microglial reactivity. Increased BBB dysfunction allows access of plasma Ang II and increases its local availability and the colocalization and activation of microglial cells. It is a potent stimulus to augments vasomotor sympathetic activity, autonomic imbalance, and pressure elevation during the establishment of hypertension.

RESUMEN

Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) is an enveloped, plus-stranded RNA virus responsible for the Coronavirus Disease 2019 (COVID-19). Patients infected with COVID-19 may be asymptomatic or have symptoms ranging from mild manifestations to severe cases of the disease that could lead to death. The SARS-CoV-2 genome encodes 4 structural proteins, including the Spike protein (S), the Nucleocapsid protein (N), Membrane protein (M) and, the Envelope protein (E). The N protein forms a major component of the ribonucleoprotein complex within the virus particle and play a vital role in its transcription and replication. Nevertheless, the S protein was the most important protein in the development of vaccines against COVID-19. However, the decrease in number of registered immunizations against the disease and the rapid drop in neutralizing antibody titers together with looser preventive measures for virus transmission, favored the rapid appearance of new variants of concerns (VOCs) that primarily show mutations in the S protein. This fact makes the N protein a good candidate for the development of diagnostic tests, due to its stability, amino acid conservation, high immunogenicity, and the smaller likelihood of mutation. With the aim of developing a new diagnostic kit based on the N protein, we evaluated the humoral response in female Wistar rats against this target. Three constructions of the N protein were used to inoculate the animals: the full-length protein (Cfull), the N- (NTD), and the C-terminal (CTD) portion of the protein. The immunizations induced the animal's immune response, with specific polyclonal IgG antibodies against the Cfull protein and its fragments. There were not non-specific bind to the protein used as negative control. Anti-Cfull antibodies demonstrated high efficiency in binding to the NTD protein and the antibodies present in the anti-CTD and anti-NTD sera have recognized the Cfull protein, but they were not able to recognize the NTD and CTD proteins, respectively. Our results indicate an efficient protocol for obtaining high antibody titers against the N recombinant protein of SARS-CoV-2 and its fragments highlighting the Cfull protein, which can be used in the development of new diagnostic kits.

Asunto(s)

COVID-19 , SARS-CoV-2 , Humanos , Femenino , Animales , Ratas , SARS-CoV-2/genética , COVID-19/diagnóstico , Anticuerpos Antivirales , Vacunas contra la COVID-19 , Ratas Wistar , Anticuerpos Neutralizantes , Proteínas de la Nucleocápside/genética , Pruebas Diagnósticas de Rutina , Prueba de COVID-19

RESUMEN

Hypertension augments while exercise training corrects the increased vesicle trafficking (transcytosis) across the blood-brain barrier (BBB) within preautonomic areas and the autonomic imbalance. There is no information on a possible mechanism(s) conditioning these effects. Knowing that Mfsd2a is the major transporter of docosahexaenoic acid (DHA) and that Mfsd2a knockout mice exhibited leaky BBB, we sought to identify its possible involvement in hypertension- and exercise-induced transcytosis across the BBB. Spontaneously hypertensive rats (SHR) and Wistar rats were submitted to treadmill training (T) or kept sedentary (S) for 4 wk. Resting hemodynamic/autonomic parameters were recorded in conscious chronically cannulated rats. BBB permeability within the hypothalamic paraventricular nucleus (PVN) was evaluated in anesthetized rats. Brains were harvested for Mfsd2a and caveolin-1 (an essential protein for vesicle formation) expression. SHR-S versus Wistar-S exhibited elevated arterial pressure (AP) and heart rate (HR), increased vasomotor sympathetic activity, reduced cardiac parasympathetic activity, greater pressure variability, reduced HR variability, and depressed baroreflex control. SHR-S also showed increased BBB permeability, reduced Mfsd2a, and increased caveolin-1 expression. SHR-T versus SHR-S exhibited increased Mfsd2a density, reduced caveolin-1 protein expression, and normalized PVN BBB permeability, which were accompanied by resting bradycardia, partial AP drop, reduced sympathetic and normalized cardiac parasympathetic activity, increased HR variability, and reduced pressure variability. No changes were observed in Wistar-T versus Wistar-S. Training is an efficient tool to rescue Mfsd2a expression, which by transporting DHA into the endothelial cell reduces caveolin-1 availability and vesicles' formation. Exercise-induced Mfsd2a normalization is an important mechanism to correct both BBB function and autonomic control in hypertensive subjects.

Asunto(s)

Hipertensión , Simportadores , Animales , Ratas , Barrera Hematoencefálica/metabolismo , Capilares/metabolismo , Caveolina 1/genética , Caveolina 1/metabolismo , Núcleo Hipotalámico Paraventricular/metabolismo , Ratas Endogámicas SHR , Ratas Wistar , Simportadores/metabolismo

RESUMEN

Introduction: Chronic hypertension is accompanied by either blood-brain barrier (BBB) leakage and autonomic dysfunction. There is no consensus on the mechanism determining increased BBB permeability within autonomic areas. While some reports suggested tight junction's breakdown, others indicated the involvement of transcytosis rather than paracellular transport changes. Interestingly, exercise training was able to restore both BBB permeability and autonomic control of the circulation. We sought now to clarify the mechanism(s) governing hypertension- and exercise-induced BBB permeability. Methods: Spontaneously hypertensive rats (SHR) and normotensive controls submitted to 4-week aerobic training (T) or sedentary protocol (S) were chronically cannulated for baseline hemodynamic and autonomic recordings and evaluation of BBB permeability. Brains were harvested for measurement of BBB function (FITC-10 kDa leakage), ultrastructural analysis of BBB constituents (transmission electron microscopy) and caveolin-1 expression (immunofluorescence). Results: In SHR-S the increased pressure, augmented sympathetic vasomotor activity, higher sympathetic and lower parasympathetic modulation of the heart and the reduced baroreflex sensitivity were accompanied by robust FITC-10kDa leakage, large increase in transcytotic vesicles number/capillary, but no change in tight junctions' density within the paraventricular nucleus of the hypothalamus, the nucleus of the solitary tract and the rostral ventrolateral medulla. SHR-T exhibited restored BBB permeability and normalized vesicles counting/capillary simultaneously with a normal autonomic modulation of heart and vessels, resting bradycardia and partial pressure reduction. Caveolin-1 expression ratified the counting of transcellular, not other cytoplasmatic vesicles. Additionally, T caused in both groups significant increases in tight junctions' extension/capillary border. Discussion: Data indicate that transcytosis, not the paracellular transport, is the primary mechanism underlying both hypertension- and exercise-induced BBB permeability changes within autonomic areas. The reduced BBB permeability contributes to normalize the autonomic control of the circulation, which suppresses pressure variability and reduces the occurrence of end-organ damage in the trained SHR. Data also disclose that hypertension does not change but exercise training strengthens the resistance of the paracellular pathway in both strains.

RESUMEN

Although hypertension disrupts the blood-brain barrier (BBB) integrity within the paraventricular nucleus of hypothalamus (PVN) and increases the leakage into the brain parenchyma, exercise training (T) was shown to correct it. Since there is scarce and contradictory information on the mechanism(s) determining hypertension-induced BBB deficit and nothing is known about T-induced improvement, we sought to evaluate the paracellular and transcellular transport across the BBB within the PVN in both conditions. Spontaneously hypertensive rats (SHR) and WKY submitted to 4-wk aerobic T or sedentary (S) protocol were chronically catheterized for hemodynamic recordings at rest and intra-arterial administration of dyes (Rhodamine-dextran 70 kDa + FITC-dextran 10 kDa). Brains were harvesting for FITC leakage examination, qPCR evaluation of different BBB constituents and protein expression of caveolin-1 and claudin-5, the main markers of transcytosis and paracellular transport, respectively. Hypertension was characterized by increased arterial pressure and heart rate, augmented sympathetic modulation of heart and vessels, and reduced cardiac parasympathetic control, marked FITC extravasation into the PVN which was accompanied by increased caveolin-1 gene and protein expression, without changes in claudin-5 and others tight junctions' components. SHR-T vs. SHR-S showed a partial pressure reduction, resting bradycardia, improvement of autonomic control of the circulation simultaneously with correction of both FITC leakage and caveolin-1 expression; there was a significant increase in claudin-5 expression. Caveolin-1 content was strongly correlated with improved autonomic control after exercise. Data indicated that within the PVN the transcytosis is the main mechanism governing both hypertension-induced BBB leakage, as well as the exercise-induced correction.

Asunto(s)

Barrera Hematoencefálica/metabolismo , Capilares/metabolismo , Permeabilidad Capilar , Caveolina 1/metabolismo , Claudina-5/metabolismo , Terapia por Ejercicio , Hipertensión/terapia , Núcleo Hipotalámico Paraventricular/irrigación sanguínea , Condicionamiento Físico Animal , Uniones Estrechas/metabolismo , Transcitosis , Animales , Barrera Hematoencefálica/fisiopatología , Capilares/fisiopatología , Sistema Cardiovascular/inervación , Caveolina 1/genética , Claudina-5/genética , Modelos Animales de Enfermedad , Hipertensión/metabolismo , Hipertensión/fisiopatología , Masculino , Esfuerzo Físico , Ratas Endogámicas SHR , Ratas Endogámicas WKY , Sistema Nervioso Simpático/fisiopatología

RESUMEN

Histidine-containing dipeptides (HCDs) are abundantly expressed in striated muscles. Although important properties have been ascribed to HCDs, including H+ buffering, regulation of Ca2+ transients and protection against oxidative stress, it remains unknown whether they play relevant functions in vivo. To investigate the in vivo roles of HCDs, we developed the first carnosine synthase knockout (CARNS1-/-) rat strain to investigate the impact of an absence of HCDs on skeletal and cardiac muscle function. Male wild-type (WT) and knockout rats (4 months-old) were used. Skeletal muscle function was assessed by an exercise tolerance test, contractile function in situ and muscle buffering capacity in vitro. Cardiac function was assessed in vivo by echocardiography and cardiac electrical activity by electrocardiography. Cardiomyocyte contractile function was assessed in isolated cardiomyocytes by measuring sarcomere contractility, along with the determination of Ca2+ transient. Markers of oxidative stress, mitochondrial function and expression of proteins were also evaluated in cardiac muscle. Animals were supplemented with carnosine (1.8% in drinking water for 12 weeks) in an attempt to rescue tissue HCDs levels and function. CARNS1-/- resulted in the complete absence of carnosine and anserine, but it did not affect exercise capacity, skeletal muscle force production, fatigability or buffering capacity in vitro, indicating that these are not essential for pH regulation and function in skeletal muscle. In cardiac muscle, however, CARNS1-/- resulted in a significant impairment of contractile function, which was confirmed both in vivo and ex vivo in isolated sarcomeres. Impaired systolic and diastolic dysfunction were accompanied by reduced intracellular Ca2+ peaks and slowed Ca2+ removal, but not by increased markers of oxidative stress or impaired mitochondrial respiration. No relevant increases in muscle carnosine content were observed after carnosine supplementation. Results show that a primary function of HCDs in cardiac muscle is the regulation of Ca2+ handling and excitation-contraction coupling.

Asunto(s)

Carnosina , Dipéptidos , Animales , Anserina , Histidina , Masculino , Músculo Esquelético , Miocitos Cardíacos , Ratas

RESUMEN

Remodeling of capillary rarefaction and deleterious arteries are characteristic hallmarks of hypertension that are partially corrected by exercise training. In addition, experimental evidence showed capillary rarefaction within the brain cortex and reduced cerebral blood flow. There is no information on hypertension- and exercise-induced effects on capillary profile and function within preautonomic nuclei. We sought now to evaluate the effects of hypertension and exercise training (T) on the capillary network within hypothalamic paraventricular (PVN) and solitary tract (NTS) nuclei, and on the remodeling of brain arteries. Age-matched spontaneously hypertensive rats (SHR) and Wistar-Kyoto (WKY), submitted to moderate T or kept sedentary (S) for three months, were chronically cannulated for hemodynamic recordings at rest. Rats were anesthetized for i.v. administration of fluorescein isothiocyanate (FITC)-dextran (capillary volume/density measurements) or 4% paraformaldehyde perfusion (basilar, middle, and posterior arteries' morphometry) followed by brain harvesting and processing. Other groups of conscious rats had carotid blood flow (CBF, ultrasound flowmeter) acquired simultaneously with hemodynamic recordings at rest and exercise. SHR-S exhibited elevated pressure and heart rate, reduced CBF, increased wall/lumen ratio of arteries, but no capillary rarefaction within the PVN and NTS. T improved performance gain and caused resting bradycardia in both groups; reduction of pressure and sympathetic vasomotor activity and normalization of the wall/lumen ratio were only observed in SHR-T. T groups responded with marked PVN and NTS capillary angiogenesis and augmented CBF during exercise; to avoid overperfusion at rest, reduced basal CBF was observed only in WKY-T. Data indicated that the absence of SHR-S capillary rarefaction and the intense SHR-T angiogenesis within autonomic areas associated with correction of deleterious arteries' remodeling are essential adjustments to hypertension and exercise training, respectively. These adaptive responses maintain adequate baseline perfusion in SHR-S and SHR-T preautonomic nuclei, augmenting it in exercised rats when a well-coordinated autonomic control is required.

RESUMEN

Aerobic exercise training improves the autonomic control of the circulation. Emerging evidence has shown that exercise induces neuroplastic adaptive changes in preautonomic circuitry controlling sympathetic/parasympathetic outflow to heart and vessels. The mechanisms underlying neuronal plasticity are, however, incompletely understood. Knowing that sinoaortic denervation blocks training-induced cardiovascular benefits, we investigate whether baroreceptors' and chemoreceptors' signaling are able to drive neuronal plasticity within medullary and supramedullary pathways controlling autonomic outflow. Male Wistar rats submitted to sinoaortic denervation (SAD) or dopamine ß-hydroxylase-saporin lesion (DBHx) and respective controls (SHAM) were allocated to training (T) or sedentary (S) protocols for 8 weeks. After hemodynamic measurements at rest, rats were deeply anesthetized for brain harvesting. The density of DBH and oxytocin (OT) cell bodies and terminals were analyzed in brainstem and hypothalamic brain areas (double immunofluorescence reactions, optic and confocal microscopy). In SHAM rats training augmented the density of DBH+ neurons in the nucleus of solitary tract, increased the density of ascending NORergic projections and the number of DBH+ boutons contacting preautonomic OT+ neurons into paraventricular hypothalamic preautonomic nuclei, augmented the density of local OTergic neurons and enhanced the density of OT+ terminals targeting brainstem autonomic areas. These plastic changes occurred simultaneously with reduced sympathetic/increased parasympathetic activity, augmented baroreflex sensitivity and reduced resting heart rate. SAD reduced the density of both DBH+ fibers ascending from brainstem to paraventricular nucleus of hypothalamus and preautonomic OT+ neurons projecting to the brainstem, abrogated training-induced plastic changes and autonomic adaptive responses without changing the treadmill performance. Minor neuroplastic changes with preserved baroreflex sensitivity were observed in trained rats after partial selective disruption of ascending NORergic projections. Our data indicated that afferent inputs conveyed by arterial baroreceptors and chemoreceptors are the main stimuli to drive both inactivity-induced and activity-dependent neuroplasticity within the autonomic circuitry.

RESUMEN

GABAergic inhibitory input within the paraventricular hypothalamic nucleus (PVN) plays a key role in restraining sympathetic outflow. Although experimental evidence has shown depressed GABAA receptor function plus sympathoexcitation in hypertension and augmented GABA levels with reduced sympathetic activity after exercise training (T), the mechanisms underlying T-induced effects remain unclear. Here we investigated in T and sedentary (S) SHR and WKY: (1) time-course changes of hemodynamic parameters and PVN glutamic acid decarboxylase (GAD) isoforms' expression; (2) arterial pressure (AP) and heart rate (HR) responses, sympathetic/parasympathetic modulation of heart and vessels and baroreflex sensitivity to GABAA receptor blockade within the PVN. SHR-S versus WKY-S exhibited higher AP and HR, increased sympathetic reduced parasympathetic modulation, smaller baroreflex sensitivity, and reduced PVN GAD65 immunoreactivity. SHR-T and WKY-T showed prompt maintained increase (2-8 weeks) in GAD65 expression (responsible for GABA vesicular pool synthesis), which occurred simultaneously with HR reduction in SHR-T and preceded MAP fall in SHR-T and resting bradycardia in WKY-T. There was no change in GAD67 expression (mainly involved with GABA metabolic pool). Resting HR in both groups and basal MAP in SHR were negatively correlated with PVN GAD65 expression. Normalized baroreflex sensitivity and autonomic control observed only in SHR-T were due to recovery of GABAA receptor function into the PVN since bicuculline administration abolished these effects. Data indicated that training augments in both groups the expression/activity of GABAergic neurotransmission within presympathetic PVN neurons and restores GABAA receptors' function specifically in the SHR, therefore strengthening GABAergic modulation of sympathetic outflow in hypertension.

Asunto(s)

Glutamato Descarboxilasa/genética , Hipertensión/metabolismo , Núcleo Hipotalámico Paraventricular/metabolismo , Esfuerzo Físico , Receptores de GABA-A/metabolismo , Animales , Barorreflejo , Presión Sanguínea , Glutamato Descarboxilasa/metabolismo , Hipertensión/fisiopatología , Isoenzimas/genética , Isoenzimas/metabolismo , Masculino , Ratas , Ratas Endogámicas SHR , Ratas Wistar , Sistema Nervioso Simpático/fisiopatología

RESUMEN

Neuroanatomical studies associating neuronal tract tracing and immunohistochemistry identified reciprocal (ascending noradrenergic/descending oxytocinergic, OTergic) connections between brainstem cardiovascular nuclei and the paraventricular hypothalamic nucleus (PVN). Previous functional studies indicated that exercise training (T) augmented the expression/activity of OTergic pathway and improve the autonomic control of the heart. Knowing that ageing is associated with autonomic dysfunction and sinoaortic denervation blocked T-induced beneficial effects, we hypothesized that T was able to reduce age-dependent impairment by improving the afferent signaling to PVN and augmenting OTergic modulation of cardiovascular control. We evaluated the combined effects of T and age on plastic remodeling of ascending dopamine ß-hydroxylase (DBH+) and descending OT+ pathways and correlated them with cardiovascular parameters. Male Wistar rats were submitted to T or kept sedentary for 8 weeks. After evaluating arterial pressure, heart rate (HR), their variabilities and spectral components in conscious rats at rest, brains were harvested to analyze the plastic remodeling of brain autonomic nuclei (immunofluorescence + confocal microscopy). The density of DBH+ neurons within the nucleus of solitary tract (NTS) and caudal ventrolateral medulla, the number of DBH+ terminals overlapping OT+ neurons in PVN preautonomic nuclei, as well as the density of OT+ neurons and their projections to NTS and dorsal motor nucleus of the vagus were markedly reduced in S rats during 8-weeks of inactivity In contrast, these effects were completely blocked by T and reversed to a large augmentation of DBH+ and OT+ densities in both cell bodies and terminals within autonomic nuclei and target areas. All plastic changes observed correlated positively with parasympathetic activity to the heart (HF-PI, but not with LF-PI) and negatively with resting HR. Data indicate that T, by increasing beneficial neuroplastic adaptive changes within brainstem-PVN reciprocal network, abrogates age-dependent deleterious remodeling and augments parasympathetic modulation of the heart, therefore improving autonomic function. This article is protected by copyright. All rights reserved.

RESUMEN

The blood-brain barrier (BBB) is a complex multicellular structure acting as selective barrier controlling the transport of substances between these compartments. Accumulating evidence has shown that chronic hypertension is accompanied by BBB dysfunction, deficient local perfusion and plasma angiotensin II (Ang II) access into the parenchyma of brain areas related to autonomic circulatory control. Knowing that spontaneously hypertensive rats (SHR) exhibit deficient autonomic control and brain Ang II hyperactivity and that exercise training is highly effective in correcting both, we hypothesized that training, by reducing Ang II content, could improve BBB function within autonomic brain areas of the SHR. After confirming the absence of BBB lesion in the pre-hypertensive SHR, but marked fluorescein isothiocyanate dextran (FITC, 10 kD) leakage into the brain parenchyma of the hypothalamic paraventricular nucleus (PVN), nucleus of the solitary tract, and rostral ventrolateral medulla during the established phase of hypertension, adult SHR, and age-matched WKY were submitted to a treadmill training (T) or kept sedentary (S) for 8 weeks. The robust FITC leakage within autonomic areas of the SHR-S was largely reduced and almost normalized since the 2nd week of training (T2). BBB leakage reduction occurred simultaneously and showed strong correlations with both decreased LF/HF ratio to the heart and reduced vasomotor sympathetic activity (power spectral analysis), these effects preceding the appearance of resting bradycardia (T4) and partial pressure fall (T8). In other groups of SHR-T simultaneously infused with icv Ang II or saline (osmotic mini-pumps connected to a lateral ventricle cannula) we proved that decreased local availability of this peptide and reduced microglia activation (IBA1 staining) are crucial mechanisms conditioning the restoration of BBB integrity. Our data also revealed that Ang II-induced BBB lesion was faster within the PVN (T2), suggesting the prominent role of this nucleus in driven hypertension-induced deficits. These original set of data suggest that reduced local Ang II content (and decreased activation of its downstream pathways) is an essential and early-activated mechanism to maintain BBB integrity in trained SHR and uncovers a novel beneficial effect of exercise training to improve autonomic control even in the presence of hypertension.

RESUMEN

KEY POINTS: Heart Failure (HF) is accompanied by reduced ventricular function, activation of compensatory neurohormonal mechanisms and marked autonomic dysfunction characterized by exaggerated sympathoexcitation and reduced parasympathetic activity. With 6 weeks of exercise training, HF-related loss of choline acetyltransferase (ChAT)-positive vagal preganglionic neurones is avoided, restoring the parasympathetic tonus to the heart, and the immunoreactivity of dopamine ß-hydroxylase-positive premotor neurones that drive sympathetic outflow to the heart is reduced. Training-induced correction of autonomic dysfunction occurs even with the persistence of abnormal ventricular function. Strong positive correlation between improved parasympathetic tonus to the heart and increased ChAT immunoreactivity in vagal preganglionic neurones after training indicates this is a crucial mechanism to restore autonomic function in heart failure. ABSTRACT: Exercise training is an efficient tool to attenuate sympathoexcitation, a hallmark of heart failure (HF). Although sympathetic modulation in HF is widely studied, information regarding parasympathetic control is lacking. We examined the combined effects of sympathetic and vagal tonus to the heart in sedentary (Sed) and exercise trained (ET) HF rats and the contribution of respective premotor and preganglionic neurones. Wistar rats submitted to coronary artery ligation or sham surgery were assigned to training or sedentary protocols for 6 weeks. After haemodynamic, autonomic tonus (atropine and atenolol i.v.) and ventricular function determinations, brains were collected for immunoreactivity assays (choline acetyltransferase, ChATir; dopamine ß-hydroxylase, DBHir) and neuronal counting in the dorsal motor nucleus of vagus (DMV), nucleus ambiguus (NA) and rostroventrolateral medulla (RVLM). HF-Sed vs. SHAM-Sed exhibited decreased exercise capacity, reduced ejection fraction, increased left ventricle end diastolic pressure, smaller positive and negative dP/dt, decreased intrinsic heart rate (IHR), lower parasympathetic and higher sympathetic tonus, reduced preganglionic vagal neurones and ChATir in the DMV/NA, and increased RVLM DBHir. Training increased treadmill performance, normalized autonomic tonus and IHR, restored the number of DMV and NA neurones and corrected ChATir without affecting ventricular function. There were strong positive correlations between parasympathetic tonus and ChATir in NA and DMV. RVLM DBHir was also normalized by training, but there was no change in neurone number and no correlation with sympathetic tonus. Training-induced preservation of preganglionic vagal neurones is crucial to normalize parasympathetic activity and restore autonomic balance to the heart even in the persistence of cardiac dysfunction.

Asunto(s)

Fibras Autónomas Preganglionares/fisiología , Insuficiencia Cardíaca/fisiopatología , Neuronas/fisiología , Condicionamiento Físico Animal , Nervio Vago/fisiología , Animales , Presión Sanguínea , Corazón/inervación , Frecuencia Cardíaca , Masculino , Ratas , Ratas Wistar , Nervio Vago/citología

RESUMEN

We showed previously that sino-aortic denervation prevented training-induced plasticity in pre-autonomic oxytocinergic neurons and blocked the beneficial effects of training. In this study, we investigate the combined effect of training and removal of specific chemoreceptor afferents on both cardiovascular parameters and oxytocin (OT) gene and protein expression within the hypothalamic paraventricular nucleus (PVN). Wistar rats and spontaneously hypertensive rats (SHRs) underwent carotid body denervation or sham surgery and were trained or kept sedentary for 3 months. After haemodynamic measurements at rest, rats were anaesthetized for brain perfusion. Fresh (perfused with PBS) and fixed brains (perfused with 4% paraformaldehyde) were processed for PVN OT mRNA (real-time PCR) and OT immunoreactivity within PVN subnuclei. In sham-operated rats, training improved treadmill performance and reduced resting heart rate (Wistar, -8%; SHRs, -10%), with a reduction in blood pressure only in SHRs (-8%). Training was accompanied by increased PVN OT mRNA expression (twofold increase in sham-operated SHRs) and peptide density in the posterior, ventromedial and dorsal cap PVN subnuclei (on average 70% increase in both strains), with significant correlations between OT content and training-induced resting bradycardia in sham-operated groups. Carotid body denervation did not interfere with the performance gain, abolished chemoreflex activation (without changing baroreflex control) and blocked training-induced cardiovascular adaptations and training-induced changes in PVN OT content in both strains. After carotid body denervation, there was no correlation between OT mRNA or OT immunoractivity and resting heart rate. The chronic absence of chemoreceptor inputs uncovers an unknown role of chemoreceptor signalling in driving the plasticity/activity of PVN oxytocinergic pre-autonomic neurons, thus mediating training-induced cardiovascular adaptive responses.

Asunto(s)

Cuerpo Carotídeo/metabolismo , Hipertensión/metabolismo , Plasticidad Neuronal , Neuronas/metabolismo , Oxitocina/metabolismo , Núcleo Hipotalámico Paraventricular/metabolismo , Esfuerzo Físico , Adaptación Fisiológica , Animales , Desnervación Autonómica , Barorreflejo , Presión Sanguínea , Modelos Animales de Enfermedad , Regulación de la Expresión Génica , Frecuencia Cardíaca , Hipertensión/genética , Hipertensión/fisiopatología , Inmunohistoquímica , Masculino , Oxitocina/genética , Núcleo Hipotalámico Paraventricular/fisiopatología , ARN Mensajero/metabolismo , Ratas , Ratas Endogámicas SHR , Ratas Wistar , Reacción en Cadena en Tiempo Real de la Polimerasa , Conducta Sedentaria , Transducción de Señal , Factores de Tiempo

RESUMEN

Salivary gland function is regulated by both the sympathetic and parasympathetic nervous systems. Previously we showed that the basal sympathetic outflow to the salivary glands (SNA(SG)) was higher in hypertensive compared to normotensive rats and that diabetes reduced SNA(SG) discharge at both strains. In the present study we sought to investigate how SNA(SG) might be modulated by acute changes in the arterial pressure and whether baroreceptors play a functional role upon this modulation. To this end, we measured blood pressure and SNA(SG) discharge in Wistar-Kyoto rats (WKY-intact) and in WKY submitted to sinoaortic denervation (WKY-SAD). We made the following three major observations: (i) in WKY-intact rats, baroreceptor loading in response to intravenous infusion of the phenylephrine evoked an increase in SNA(SG) spike frequency (81%, p<0.01) accompanying the increase mean arterial pressure (ΔMAP: +77 ± 14 mmHg); (ii) baroreceptor unloading with sodium nitroprusside infusion elicited a decrease in SNA(SG) spike frequency (17%, p<0.01) in parallel with the fall in arterial blood pressure (ΔMAP: -30 ± 3 mmHg) in WKY-intact rats; iii) in the WKY-SAD rats, phenylephrine-evoked rises in the arterial pressure (ΔMAP: +56 ± 6 mmHg) failed to produce significant changes in the SNA(SG) spike frequency. Taken together, these data show that SNA(SG) increases in parallel with pharmacological-induced pressor response in a baroreceptor dependent way in anaesthetised rats. Considering the key role of SNA(SG) in salivary secretion, this mechanism, which differs from the classic cardiac baroreflex feedback loop, strongly suggests that baroreceptor signalling plays a decisive role in the regulation of salivary gland function.

Asunto(s)

Vías Aferentes/fisiología , Barorreflejo/fisiología , Presorreceptores/metabolismo , Glándulas Salivales/fisiología , Sistema Nervioso Simpático/fisiología , Potenciales de Acción/efectos de los fármacos , Vías Aferentes/efectos de los fármacos , Anestésicos Intravenosos/farmacología , Animales , Antihipertensivos/farmacología , Barorreflejo/efectos de los fármacos , Presión Sanguínea/efectos de los fármacos , Desnervación/métodos , Frecuencia Cardíaca/efectos de los fármacos , Nitroprusiato/farmacología , Fenilefrina/farmacología , Presorreceptores/efectos de los fármacos , Ratas , Ratas Endogámicas WKY , Glándulas Salivales/efectos de los fármacos , Sistema Nervioso Simpático/lesiones , Simpatomiméticos/farmacología , Uretano/farmacología

RESUMEN

We showed previously that oxytocinergic (OTergic) projections from the hypothalamic paraventricular nucleus (PVN) to the dorsal brain stem mediate training-induced heart rate (HR) adjustments and that beneficial effects of training are blocked by sinoaortic denervation (SAD; Exp Physiol 94: 630-640; 1103-1113, 2009). We sought now to determine the combined effect of training and SAD on PVN OTergic neurons in spontaneously hypertensive rats (SHR) and Wistar-Kyoto (WKY) rats. Rats underwent SAD or sham surgery and were trained (55% of maximal capacity) or kept sedentary for 3 mo. After hemodynamic measurements were taken at rest, rats were deeply anesthetized. Fresh brains were frozen and sliced to isolate the PVN; samples were processed for OT expression (real-time PCR) and fixed brains were processed for OT immunofluorescence. In sham rats, training improved treadmill performance and increased the gain of baroreflex control of HR. Training reduced resting HR (-8%) in both groups, with a fall in blood pressure (-10%) only in SHR rats. These changes were accompanied by marked increases in PVN OT mRNA expression (3.9- and 2.2-fold in WKY and SHR rats, respectively) and peptide density in PVN OTergic neurons (2.6-fold in both groups), with significant correlations between OT content and training-induced resting bradycardia. SAD abolished PVN OT mRNA expression and markedly reduced PVN OT density in WKY and SHR. Training had no effect on HR, PVN OT mRNA, or OT content following SAD. The chronic absence of inputs from baroreceptors and chemoreceptors uncovers the pivotal role of afferent signaling in driving both the plasticity and activity of PVN OTergic neurons, as well as the beneficial effects of training on cardiovascular control.

Asunto(s)

Vías Autónomas/metabolismo , Plasticidad Neuronal/fisiología , Neuronas Aferentes/fisiología , Oxitocina/metabolismo , Condicionamiento Físico Animal/fisiología , Transducción de Señal/fisiología , Animales , Presión Sanguínea/fisiología , Desnervación , Modelos Animales de Enfermedad , Frecuencia Cardíaca/fisiología , Hipertensión/fisiopatología , Masculino , Modelos Animales , Núcleo Hipotalámico Paraventricular/fisiología , Ratas , Ratas Endogámicas SHR , Ratas Endogámicas WKY , Nodo Sinoatrial/fisiología , Nodo Sinoatrial/cirugía , Núcleo Supraóptico/fisiología

RESUMEN

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.

Asunto(s)

Dosificación de Gen , Predisposición Genética a la Enfermedad , Hipertensión Renovascular/genética , Peptidil-Dipeptidasa A/genética , Animales , Animales Modificados Genéticamente , Arritmias Cardíacas/genética , Sistema Nervioso Autónomo/fisiopatología , Barorreflejo/genética , Presión Sanguínea/genética , Estudios de Asociación Genética , Corazón/fisiopatología , Hemodinámica/genética , Hipertensión Renovascular/sangre , Hipertensión Renovascular/metabolismo , Hipertensión Renovascular/fisiopatología , Hipertrofia Ventricular Izquierda/genética , Pulmón/enzimología , Masculino , Ratones , Miocardio/enzimología , Peptidil-Dipeptidasa A/sangre , Peptidil-Dipeptidasa A/metabolismo , Índice de Severidad de la Enfermedad , Sistema Nervioso Simpático/fisiopatología , Nervio Vago/fisiopatología

RESUMEN

We investigate whether arterial baroreceptors mediate the training-induced blood pressure fall and resting bradycardia in hypertensive (SHR) and normotensive rats (WKY). Male SHR and WKY rats, submitted to sino-aortic denervation (SAD) or sham surgery (SHAM group), were allocated to training (T; 55% of maximal exercise capacity) or sedentary (S) protocols for 3 months. Rats were instrumented with arterial and venous catheters for haemodynamic measurements at rest (power spectral analysis) and baroreceptor testing. Kidney and skeletal muscles were processed for morphometric analysis of arterioles. Elevated mean arterial pressure (MAP) and heart rate (HR) in SHAM SHRS were accompanied by increased sympathetic variability and arteriolar wall/lumen ratio [+3.4-fold on low-frequency (LF) power and +70%, respectively, versus WKYS, P < 0.05]. Training caused significant HR (approximately 9% in WKY and SHR) and MAP reductions (-8% in the SHR), simultaneously with improvement of baroreceptor reflex control of HR (SHR and WKY), LF reduction (with a positive correlation between LF power and MAP levels in the SHR) and normalization of wall/lumen ratio of the skeletal muscle arterioles (SHR only). In contrast, SAD increased pressure variability in both strains of rats, causing reductions in MAP (-13%) and arteriolar wall/lumen ratio (-35%) only in the SHRS. Training effects were completely blocked by SAD in both strains; in addition, after SAD the resting MAP and HR and the wall/lumen ratio of skeletal muscle arterioles were higher in SHRT versus SHRS and similar to those of SHAM SHRS. The lack of training-induced effects in the chronic absence of baroreceptor inputs strongly suggests that baroreceptor signalling plays a decisive role in driving beneficial training-induced cardiovascular adjustments.

Asunto(s)

Presión Sanguínea/fisiología , Frecuencia Cardíaca/fisiología , Hipertensión/fisiopatología , Presorreceptores/fisiología , Animales , Arteriolas/patología , Barorreflejo/fisiología , Desnervación , Hipertensión/patología , Masculino , Condicionamiento Físico Animal/fisiología , Ratas , Ratas Endogámicas SHR , Ratas Endogámicas WKY , Transducción de Señal , Nodo Sinoatrial/fisiología , Nodo Sinoatrial/cirugía

RESUMEN

O estudo do sistema cardiovascular do camundongo vem recebendo crescente atenção uma vez que muitas das ferramentas genéticas disponíveis tem sido aplicadas a essa espécie. No estudo da genética da hipertensão, os efeitos sitsêmicos de mutações em genes candidatos tem sido avaliados. Dessa forma a inativação do gene da enzima conversora de angiotensina (ECA) em camundongos induzindo ausência da ECA reduz a pressão arterial (PA), indicando que o gene da ECA é essencial para a manutenção da PA normal. Dessa forma, variações na síntese da enzima conversora da angiotensina (ECA) poderiam estar relacionadas com variações na PA. Entretanto, experimentos anteriores mostraram que a PA de camundongos com diferentes números de cópias do gene da ECA , obtida de maneira indireta, se mantém inalterada, apesar de haver variações da atividade da enzima no soro. O objetivo deste trabalho foi avaliar o controle neurogênico da PA em camundongos normais e transgênicos com diferente número de cópias do gene da ECA, pela medida da PA, freqüência cardíaca (FC) e sensibilidade baroreflexa em condições basais e após a implantação cirúrgica de um clipe na artéria renal. Em uma primeira etapa deste trabalho realizamos experimentos com animais controle (C57BL6/J) nos quais executamos avaliações hemodinâmicas, registro da atividade do nervo depressor aórtico e desnervação sino-aórtica com o objetivo de estabelecer referencias de normalidade para este modelo. Em uma segunda etapa foram estudados camundongos transgênicos (n=27) sendo divididos em 4 grupos de acordo com o número de cópias do gene da enzima conversora da angiotensina (de 1 a 4 cópias). A artéria carótida e veia jugular foram canuladas para registro da pressão intra-arterial e administração de drogas vasoativas. Os sinais de PA e FC foram registrados batimento a batimento com uma freqüência de amostragem de 3000 hertz por canal através do programa AT/CODAS (DataQ Instruments, lnc., Ohio, USA). Em uma terceira etapa foram estudados camundongos transgênicos implantados em sua artéria renal direita com um clipe de prata em formato de "U" com medida de abertura interna de 0,12 mm. Estes animais foram divididos em três grupos: A)animais contendo 2 cópias para o gene da ECA sem clipe na artéria renal (n=5), B) animais contendo 2 cópias para o gene da ECA contendo clipe na artéria renal(n=5) e C) animais de 3 cópias contendo clipe na artéria renal (n=5)…(au)

Asunto(s)

Presión Arterial , Ratones Transgénicos , Peptidil-Dipeptidasa A