Silvana S. Meyrelles

Mechanisms determining sensitivity of baroreceptor afferents in health and disease.

Abstract: Baroreceptors sense and signal the central nervous system of changes in arterial pressure through a series of sensory processes. An increase in arterial pressure causes vascular distension and baroreceptor deformation, the magnitude of which depends on the mechanical viscoelastic properties of the vessel wall. Classic methods (e.g., isolated carotid sinus preparation) and new approaches, including studies of isolated baroreceptor neurons in culture, gene transfer using viral vectors, and genetically modified mice have been used to define the cellular and molecular mechanisms that determine baroreceptor sensitivity. Deformation depolarizes the nerve endings by opening a new class of mechanosensitive ion channel. This depolarization triggers action potential discharge through opening of voltage‐dependent sodium (Na+) and potassium (K+) channels at the “spike initiating zone” (SIZ) near the sensory terminals. The resulting baroreceptor activity and its sensitivity to changes in pressure are modulated through a variety of mechanisms that influence these sensory processes. Modulation of voltage‐dependent Na+ and K+ channels and the Na+ pump at the SIZ by membrance potential, action potential discharge, and chemical autocrine and paracrine factors are important mechanisms contributing to changes in baroreceptor sensitivity during sustained increases in arterial pressure and in pathological states associated with endothelial dysfunction, oxidative stress, and platelet activation.

Endothelial dysfunction in the apolipoprotein E-deficient mouse: insights into the influence of diet, gender and aging.

Since the early 1990s, several strains of genetically modified mice have been developed as models for experimental atherosclerosis. Among the available models, the apolipoprotein E-deficient (apoE-/-) mouse is of particular relevance because of its propensity to spontaneously develop hypercholesterolemia and atherosclerotic lesions that are similar to those found in humans, even when the mice are fed a chow diet. The main purpose of this review is to highlight the key achievements that have contributed to elucidating the mechanisms pertaining to vascular dysfunction in the apoE-/- mouse. First, we summarize lipoproteins and atherosclerosis phenotypes in the apoE-/- mouse, and then we briefly discuss controversial evidence relative to the influence of gender on the development of atherosclerosis in this murine model. Second, we discuss the main mechanisms underlying the endothelial dysfunction of conducting vessels and resistance vessels and examine how this vascular defect can be influenced by diet, aging and gender in the apoE-/- mouse.

Cardiac and vascular changes in elderly atherosclerotic mice: the influence of gender

BackgroundAlthough advanced age is considered a risk factor for several diseases, the impact of gender on age-associated cardiovascular diseases, such as atherosclerotic processes and valvular diseases, remains not completely clarified. The present study was designed to assess aortic valve morphology and function and vascular damage in elderly using the apolipoprotein E knockout (ApoE KO) mouse. Our hypothesis was that advanced age-related cardiovascular changes are aggravated in atherosclerotic male mice.MethodsThe grade (0 to 4) of aortic regurgitation was evaluated through angiography. In addition, vascular lipid deposition and senescence were evaluated through histochemical analyses in aged male and female ApoE KO mice, and the results were compared to wild-type C57BL/6J (C57) mice.ResultsAortic regurgitation was observed in 92% of the male ApoE KO mice and 100% of the male C57 mice. Comparatively, in age-matched female ApoE KO and C57 mice, aortic regurgitation was observed in a proportion of 58% and 53%, respectively. Histological analysis of the aorta showed an outward (positive) remodeling in ApoE KO mice (female: 1.86 ± 0.15; male: 1.89 ± 0.68) using C57 groups as reference values. Histochemical evaluation of the aorta showed lipid deposition and vascular senescence only in the ApoE KO group, which were more pronounced in male mice.ConclusionThe data show that male gender contributes to the progression of aortic regurgitation and that hypercholesterolemia and male gender additively contribute to the occurrence of lipid deposition and vascular senescence in elderly mice.

Evaluation of vascular function in apolipoprotein E knockout mice with angiotensin-dependent renovascular hypertension.

It is known that the endothelial function is compromised in atherosclerosis and arterial hypertension and that angiotensin is an important factor contributing to both pathophysiologies. The aim of this study was to evaluate the vascular function in a hypercholesterolemia/atherosclerosis model, in the angiotensin II–dependent 2-kidney 1-clip (2K1C) hypertension model and when both conditions coexist. Eight-week-old apolipoprotein E knockout (apoE; n=20) and C57BL/6 (C57; n=20) mice underwent a 2K1C or sham operation and were studied 28 days later. Mean arterial pressure was higher in apoE—2K1C and C57–2K1C (126±3 and 128±3 mm Hg) when compared with the apoE–Sham and C57–Sham (103±2 and 104±2 mm Hg, respectively; P<0.05). The vascular reactivity to norepinephrine (NE; 10−9 to 2×10−3 mol/L), acetylcholine (ACh), and sodium nitroprusside (SNP; 10−10 to 10−3 mol/L) was evaluated in the mesenteric arteriolar bed through concentration–effect curves. NE caused vascular hyper-reactivity in apoE–Sham, apoE–2K1C, and C57–2K1C (maximal response 146±5, 144±5, and 159±4 mm Hg, respectively) compared with C57–Sham (122±7 mm Hg; P<0.05). The ACh-induced relaxation was smaller (P<0.05) in apoE–2K1C and C57–2K1C (maximal response 53±3% and 46±3%) than in apoE–Sham and C57–Sham mice (78±5% and 73±4%). SNP-induced vascular relaxation showed similar concentration–effect curves in all groups. We conclude that in C57–2K1C mice, the increased reactivity to NE and the decreased endothelium-dependent relaxation contribute to the maintenance of hypertension. The apoE mouse, at early stages of atherosclerosis, shows hyper-reactivity to NE but does not have endothelium dysfunction yet. However, the concurrence of both pathophysiologies does not result in additive effects on the vascular function.

Cardiac and vascular phenotypes in the apolipoprotein E-deficient mouse

Cardiovascular death is frequently associated with atherosclerosis, a chronic multifactorial disease and a leading cause of death worldwide. Genetically engineered mouse models have proven useful for the study of the mechanisms underlying cardiovascular diseases. The apolipoprotein E-deficient mouse has been the most widely used animal model of atherosclerosis because it rapidly develops severe hypercholesterolemia and spontaneous atherosclerotic lesions similar to those observed in humans. In this review, we provide an overview of the cardiac and vascular phenotypes and discuss the interplay among nitric oxide, reactive oxygen species, aging and diet in the impairment of cardiovascular function in this mouse model.

Neural reflex regulation of arterial pressure in pathophysiological conditions: interplay among the baroreflex, the cardiopulmonary reflexes and the chemoreflex

The maintenance of arterial pressure at levels adequate to perfuse the tissues is a basic requirement for the constancy of the internal environment and survival. The objective of the present review was to provide information about the basic reflex mechanisms that are responsible for the moment-to-moment regulation of the cardiovascular system. We demonstrate that this control is largely provided by the action of arterial and non-arterial reflexes that detect and correct changes in arterial pressure (baroreflex), blood volume or chemical composition (mechano- and chemosensitive cardiopulmonary reflexes), and changes in blood-gas composition (chemoreceptor reflex). The importance of the integration of these cardiovascular reflexes is well understood and it is clear that processing mainly occurs in the nucleus tractus solitarii, although the mechanism is poorly understood. There are several indications that the interactions of baroreflex, chemoreflex and Bezold-Jarisch reflex inputs, and the central nervous system control the activity of autonomic preganglionic neurons through parallel afferent and efferent pathways to achieve cardiovascular homeostasis. It is surprising that so little appears in the literature about the integration of these neural reflexes in cardiovascular function. Thus, our purpose was to review the interplay between peripheral neural reflex mechanisms of arterial blood pressure and blood volume regulation in physiological and pathophysiological states. Special emphasis is placed on the experimental model of arterial hypertension induced by N-nitro-L-arginine methyl ester (L-NAME) in which the interplay of these three reflexes is demonstrable.

Cardiovascular Neural Reflexes in L-NAME–Induced Hypertension in Mice

The mouse is the most used animal for studying the genetic basis of cardiovascular diseases. However, the mechanisms of regulation of cardiovascular function in this animal are not yet well understood. The goal of this study was to evaluate the baroreflex, the Bezold-Jarisch cardiopulmonary reflex (BJR), and the chemoreflex in mice with hypertension induced by inhibition of NO using N&ohgr;-nitro-l-arginine-methyl ester (L-NAME). Basal mean arterial pressure (MAP) measured under anesthesia (urethane, 1 mg/g IP) was significantly higher in L-NAME (400 &mgr;g/g IP for 7 days)–treated (HT) mice (n=7) compared with vehicle-treated (NT; n=10) animals (126±9 versus 79±2 mm Hg) without differences in heart rate (HR). Baroreflex sensitivity, evaluated using phenylephrine (1 &mgr;g/g IV) was enhanced in HT mice compared with NT mice (−9.8±1.4 versus −4.9±0.5 bpm/mm Hg). The BJR, induced by phenylbiguanide (40 ng/g IV), was significantly attenuated in HT animals (MAP, −13±5%; HR, −39±6%) compared with NT animals (MAP, −38±5%; HR, −66±2%). The chemoreflex, induced by potassium cyanide (0.26 &mgr;g/g IV), was significantly attenuated in HT animals (MAP, +14±4%; HR, −8±2%) compared with NT animals (MAP, +29±4%; HR, −15±4%). As has been observed in rats, chronic inhibition of NO synthase in mice results in arterial hypertension. Enhancement of baroreflex sensitivity and attenuation of BJR and chemoreflex seem to be mainly caused by inhibition of NO synthesis because individual analyses did not show positive correlation between changes in these reflexes and MAP levels in the HT group.

Sildenafil restores endothelial function in the apolipoprotein E knockout mouse

BackgroundAtherosclerosis is an inflammatory process of the arterial walls and is initiated by endothelial dysfunction accompanied by an imbalance in the production of reactive oxygen species (ROS) and nitric oxide (NO). Sildenafil, a selective phosphodiesterase-5 (PDE5) inhibitor used for erectile dysfunction, exerts its cardiovascular effects by enhancing the effects of NO. The aim of this study was to investigate the influence of sildenafil on endothelial function and atherosclerosis progression in apolipoprotein E knockout (apoE−/−) mice.MethodsApoE−/− mice treated with sildenafil (Viagra®, 40 mg/kg/day, for 3 weeks, by oral gavage) were compared to the untreated apoE−/− and the wild-type (WT) mice.Aortic rings were used to evaluate the relaxation responses to acetylcholine (ACh) in all of the groups. In a separate set of experiments, the roles of NO and ROS in the relaxation response to ACh were evaluated by incubating the aortic rings with L-NAME (NO synthase inhibitor) or apocynin (NADPH oxidase inhibitor). In addition, the atherosclerotic lesions were quantified and superoxide production was assessed.ResultsSildenafil restored the vasodilator response to acetylcholine (ACh) in the aortic rings of the apoE−/− mice. Treatment with L-NAME abolished the vasodilator responses to ACh in all three groups of mice and revealed an augmented participation of NO in the endothelium-dependent vasodilation in the sildenafil-treated animals. The normalized endothelial function in sildenafil-treated apoE−/− mice was unaffected by apocynin highlighting the low levels of ROS production in these animals. Moreover, morphological analysis showed that sildenafil treatment caused approximately a 40% decrease in plaque deposition in the aorta.ConclusionThis is the first study demonstrating the beneficial effects of chronic treatment with sildenafil on endothelial dysfunction and atherosclerosis in a model of spontaneous hypercholesterolemia. These data indicate that the main mechanism of the beneficial effect of sildenafil on the endothelial function appears to involve an enhancement of the NO pathway along with a reduction in oxidative stress.

Experimental-Induced Periodontitis is Exacerbated in Spontaneously Hypertensive Rats

Studies show that systemic diseases such as diabetes, hyperthyroidism, osteoporosis, and dyslipidemia may influence periodontal inflammation. However, few studies relate the influence of arterial hypertension on periodontitis. The present study was undertaken to assess the severity of the experimental ligature-induced periodontitis in an experimental model of genetic arterial hypertension. The experimental periodontitis model was induced in 6 spontaneously hypertensive rats (SHR) and 6 Wistar normotensive rats (NT) by cotton ligature, which was placed subgingivaly around the neck of the first left inferior molar tooth. In the same animal, the first right molar tooth was sham-ligated and used as a control. After 7 days, the mean arterial pressure (MAP) and heart rate (HR) were recorded in conscious animals. As expected, MAP was significantly higher in SHR (151 ± 6 mmHg) than in NT (105 ± 3 mmHg), without significant differences in HR. The histopathologic examination of the periodontal structure showed alveolar integrity and lack of neutrophils and osteoclasts in the control side of both SHR and NT. In contrast, examination of the ligated side in all animals showed collagen degradation in the alveolar process from a moderate (50%) to severe (50%) level in SHR and mild in NT (100%). These data show that experimental periodontitis, characterized by the spreading of the inflammatory process from the gingiva deep into periodontium tissues, is greatly exacerbated in SHR.

Adenovirus-mediated gene transfer to cultured nodose sensory neurons.

Recent advances have enabled transfer of genes to various types of cells and tissues. The goals of the present study were to transfer genes to nodose sensory neurons using replication-deficient adenovirus vectors and to define the conditions needed to optimize the gene transfer. Neurons were dissociated from rat nodose ganglia and maintained in culture. Cultures were exposed for 30 min to vectors containing the beta-galactosidase gene lacZ driven by either the Rous sarcoma virus (RSV) or the cytomegalovirus (CMV) promoter. Cultures were fixed and treated with X-gal to evaluate lacZ expression 1-7 days after exposure to virus. Increasing concentrations of virus led to dose-related increases in the number of neurons expressing lacZ. LacZ was expressed in 8 +/- 2, 39 +/- 6, and 82 +/- 3% of neurons 1 day after exposure to 10(7), 10(8), and 10(9) pfu/ml of AdRSVlacZ, respectively (P < 0.05). The same doses of AdCMVlacZ led to expression in 41 +/- 9, 60 +/- 10, and 86 +/- 4% of neurons. Expression driven by the CMV promoter was essentially maximal within 1 day and remained stable for at least 7 days. In contrast, expression driven by the RSV promoter was less on day 1 but increased over time (1-7 days). There was no lacZ expression in vehicle-treated cultures and exposure to the adenovirus vectors did not adversely influence cell viability. Exposure of the neuronal cultures to an adenovirus vector containing the gene for green fluorescent protein (AdRSVgfp, 10(9) pfu/ml) enabled visualization of successful gene transfer in living neurons. The results indicate that gene transfer to cultured nodose neurons can be accomplished using adenovirus vectors. The expression of the transferred gene persists for at least 7 days, occurs more rapidly when expression is driven by the CMV compared with the RSV promoter, and occurs without adversely affecting cell viability.