Helio Cesar Salgado

Heart Rate Variability and Baroreceptor Function in Chronic Diabetic Rats

In conscious chronic (12 to 18 weeks) streptozotocin diabetic rats, we examined the changes in basal heart rate, with particular attention to heart rate variability assessed by evaluating the standard deviation (bpm) of the lengths of adjacent pulse pressure. We also investigated in anesthetized rats the ability of the aortic baroreceptors to acutely (30 minutes) reset to hypertensive levels. For this purpose, pressure-nerve activity curves for the baroreceptors were obtained, and gain (slope of the curve) and mean arterial pressure at 50% of maximal baroreceptor activity were calculated. The shift of the pressure-nerve activity curve was used as an index of resetting. Conscious diabetic rats (n=6) exhibited lower mean arterial pressure (93+/-6 versus 109+/-4 mm Hg), heart rate (272+/-25 versus 359+/-11 bpm), and heart rate variability (18+/-7 versus 36+/-6 bpm) than control rats (n=7). Under anesthesia, diabetic rats (n=7) and control rats (n=8) exhibited similar mean arterial pressure (113+/-6 versus 109+/-7 mm Hg in control rats ), mean arterial pressure at 50% of maximal baroreceptor activity (117+/-5 versus 107+/-6 bpm), gain (1.66+/-0.08 versus 1.81+/-0.05%/mm Hg), and extent of resetting (44+/-12 versus 49+/-9%) to hypertensive levels. The present study demonstrated that conscious chronic diabetic rats presented lower heart rate variability than control rats. On the other hand, chronic diabetes was not associated with alterations in baroreceptor function or its ability to rapidly reset to hypertensive levels.

A Descriptive and Quantitative Light and Electron Microscopy Study of the Aortic Depressor Nerve in Normotensive Rats

There is no literature report of a detailed morphologic study of the aortic depressor nerve. The aim of this study was to describe the general morphological aspects and to obtain morphometric parameters for the aortic depressor nerve of normotensive Wistar rats (n=12). Before the morphologic studies, nerves were isolated and pressure-nerve activity curves were obtained. Basal mean arterial pressure was 117+/-5 mm Hg, the systolic pressure threshold was 100+/-7 mm Hg, and mean arterial pressure at 50% of maximal activity was 115+/-5 mm Hg and the baroreceptor gain 1.99+/-0.09%/mm Hg. Semithin and thin sections of proximal and distal nerve segments were then examined by light and electron microscopy, respectively. The main nerve components were (1) unmyelinated and myelinated axons; (2) Schwann cells; (3) capillary wall endothelial cells and pericytes; (4) collagen fibers in the epineurium and endoneurium and between perineurial cell layers; and (5) fibroblasts and mast cells. The depressor nerves were found to contain 204-996 axons per nerve, 80% of which, on average, were unmyelinated, with a 4:1 unmyelinated/myelinated axon ratio. The unmyelinated axon histogram was unimodal, with a mean diameter of 0.5+/-0.02 microm. Myelinated fibers had axons averaging 1.3+/-0.06 microm in diameter and representing 53% of the total fiber diameter. The ratio between axonal and total fiber diameter of myelinated fiber ranged from 0.4 to 0.8 and tended to increase with axon size. Proximal and distal segments were morphologically similar. In conclusion, the morphologic description of the depressor nerve provides important data for further investigations of the structural basis of altered baroreflex responses in conditions such as arterial hypertension, aging, atherosclerosis, and peripheral neuropathies.

Myocardial performance in conscious streptozotocin diabetic rats

BackgroundIn spite of a large amount of studies in anesthetized animals, isolated hearts, and in vitro cardiomyocytes, to our knowledge, myocardial function was never studied in conscious diabetic rats. Myocardial performance and the response to stress caused by dobutamine were examined in conscious rats, fifteen days after the onset of diabetes caused by streptozotocin (STZ). The protective effect of insulin was also investigated in STZ-diabetic rats.MethodsCardiac contractility and relaxation were evaluated by means of maximum positive (+dP/dtmax) and negative (-dP/dtmax) values of first derivative of left ventricular pressure over time. In addition, it was examined the myocardial response to stress caused by two dosages (1 and 15 μg/kg) of dobutamine. One-way analysis of variance (ANOVA) was used to compare differences among groups, and two-way ANOVA for repeated measure, followed by Tukey post hoc test, to compare the responses to dobutamine. Differences were considered significant if P < 0.05.ResultsBasal mean arterial pressure, heart rate, +dP/dtmax and -dP/dtmax were found decreased in STZ-diabetic rats, but unaltered in control rats treated with vehicle and STZ-diabetic rats treated with insulin. Therefore, insulin prevented the hemodynamic and myocardial function alterations observed in STZ-diabetic rats. Lower dosage of dobutamine increased heart rate, +dP/dtmax and -dP/dtmax only in STZ-diabetic rats, while the higher dosage promoted greater, but similar, responses in the three groups. In conclusion, the results indicate that myocardial function was remarkably attenuated in conscious STZ-diabetic rats. In addition, the lower dosage of dobutamine uncovered a greater responsiveness of the myocardium of STZ-diabetic rats. Insulin preserved myocardial function and the integrity of the response to dobutamine of STZ-diabetic rats.ConclusionThe present study provides new data from conscious rats showing that the cardiomyopathy of this pathophysiological condition was expressed by low indices of contractility and relaxation. In addition, it was also demonstrated that these pathophysiological features were prevented by the treatment with insulin.

Increase in parasympathetic tone by pyridostigmine prevents ventricular dysfunction during the onset of heart failure

Heart failure (HF) is characterized by elevated sympathetic activity and reduced parasympathetic control of the heart. Experimental evidence suggests that the increase in parasympathetic function can be a therapeutic alternative to slow HF evolution. The parasympathetic neurotransmission can be improved by acetylcholinesterase inhibition. We investigated the long-term (4 wk) effects of the acetylcholinesterase inhibitor pyridostigmine on sympathovagal balance, cardiac remodeling, and cardiac function in the onset of HF following myocardial infarction. Myocardial infarction was elicited in adult male Wistar rats. After 4 wk of pyridostigmine administration, per os, methylatropine and propranolol were used to evaluate the cardiac sympathovagal balance. The tachycardic response caused by methylatropine was considered to be the vagal tone, whereas the bradycardic response caused by propranolol was considered to be the sympathetic tone. In conscious HF rats, pyridostigmine reduced the basal heart rate, increased vagal, and reduced sympathetic control of heart rate. Pyridostigmine reduced the myocyte diameter and collagen density of the surviving left ventricle. Pyridostigmine also increased vascular endothelial growth factor protein in the left ventricle, suggesting myocardial angiogenesis. Cardiac function was assessed by means of the pressure-volume conductance catheter system. HF rats treated with pyridostigmine exhibited a higher stroke volume, ejection fraction, cardiac output, and contractility of the left ventricle. It was demonstrated that the long-term administration of pyridostigmine started right after coronary artery ligation augmented cardiac vagal and reduced sympathetic tone, attenuating cardiac remodeling and left ventricular dysfunction during the progression of HF in rats.

Reflex control of arterial pressure and heart rate in short-term streptozotocin diabetic rats

Impaired baroreflex sensitivity in diabetes is well described and has been attributed to autonomic diabetic neuropathy. In the present study conducted on acute (10-20 days) streptozotocin (STZ)-induced diabetic rats we examined: 1) cardiac baroreflex sensitivity, assessed by the slope of the linear regression between phenylephrine- or sodium nitroprusside-induced changes in arterial pressure and reflex changes in heart rate (HR) in conscious rats; 2) aortic baroreceptor function by means of the relationship between systolic arterial pressure and aortic depressor nerve (ADN) activity, in anesthetized rats, and 3) bradycardia produced by electrical stimulation of the vagus nerve or by the iv injection of methacholine in anesthetized animals. Reflex bradycardia (-1.4 +/- 0.1 vs -1.7 +/- 0.1 bpm/mmHg) and tachycardia (-2.1 +/- 0.3 vs -3.0 +/- 0.2 bpm/mmHg) were reduced in the diabetic group. The gain of the ADN activity relationship was similar in control (1.7 +/- 0.1% max/mmHg) and diabetic (1.5 +/- 0.1% max/mmHg) animals. The HR response to vagal nerve stimulation with 16, 32 and 64 Hz was 13, 16 and 14% higher, respectively, than the response of STZ-treated rats. The HR response to increasing doses of methacholine was also higher in the diabetic group compared to control animals. Our results confirm the baroreflex dysfunction detected in previous studies on short-term diabetic rats. Moreover, the normal baroreceptor function and the altered HR responses to vagal stimulation or methacholine injection suggest that the efferent limb of the baroreflex is mainly responsible for baroreflex dysfunction in this model of diabetes.

Morphology of aortic depressor nerve myelinated fibers in normotensive Wistar-Kyoto and spontaneously hypertensive rats

Reports on the morphology of the baroreceptor terminal of spontaneously hypertensive rats (SHR) did not demonstrate any difference when compared to the axonal terminal of normotensive rats. Although several studies reporting baroreceptor terminal and blood vessel wall morphology have been carried out to better understand the baroreceptor function and resetting to hypertensive levels, there are no reports examining the morphology of the fibers of the aortic depressor nerve (ADN) in hypertensive models. Therefore, the objective of the present study was to investigate the morphological aspects of SHR ADN compared to Wistar-Kyoto (WKY) rats. Before the morphologic study, the nerves were isolated and the pressure-nerve activity curve was determined for each ADN. SHR exhibited an increase in the threshold pressure for baroreceptor activation, a rightward shift in the pressure-nerve activity curve with decreases in slope and maximum activity. Semithin (0.3 to 0.5 microm thick) sections of the proximal (close to the nodose ganglion) and distal (close to the aortic arch) segments of the ADN were analyzed by light microscopy. A morphometric study of the nerve fascicles and myelinated fibers was performed. Comparison between proximal and distal segments of the two strains revealed that the ADN of WKY rats were consistently larger. All morphometric parameters studied in myelinated fibers and their respective axons were smaller in SHR. The area of the myelin sheath was comparatively larger in WKY rats. These data show morphologic differences between the ADN of SHR and WKY rats, which may explain, at least in part, the decreased slope and maximum activity of the pressure-nerve activity curve observed with the baroreceptor resetting in SHR.

Hemodynamic responses to electrical stimulation of the aortic depressor nerve in awake rats

Changes in mean arterial pressure (MAP), heart rate (HR), and vascular resistance (hindquarter and mesenteric territories) in response to electrical stimulation (ES) of the aortic depressor nerve (ADN) were evaluated in conscious freely moving rats. Platinum electrodes were implanted into the ADN of all rats studied, and some of these animals were also implanted with miniaturized Doppler probes around the superior mesenteric artery and inferior abdominal aorta (hindquarter). In both groups, the femoral artery and vein were catheterized one day before the experiments. In the first group of rats ( n = 7), the control ES of the ADN in the range from 0.5 to 3.0 V (50 Hz, 10 ms) produced bradycardia and hypotension in an intensity-dependent manner, and treatment with methylatropine (intravenously) blocked the bradycardia but produced no significant changes in the hypotensive response. In a second group ( n = 6), ES of the ADN was performed with the intensity fixed at 3 V and the frequency of the stimuli varying from 10 to 50 Hz. In this group, the hypotensive response was frequency dependent, whereas the bradycardic response was not. In a third group of rats ( n = 6), ES of the ADN (2.5 V) produced hypotension (-35 ± 4 mmHg), minor changes in the mesenteric (+5 ± 14%), and vasodilation in hindquarter (-32 ± 6%) vascular beds. The data show that 1) ES of the ADN produces a fall in pressure, bradycardia, vasodilation in the hindquarter, and no changes in the mesenteric vascular resistance, 2) methylatropine blocked the bradycardia and produced no effect on the hypotensive response to ES of the ADN, and 3) the baroreceptor afferent fibers involved in the hypotensive response to ES of ADN are sensitive to the variation of the frequency of the stimuli, whereas the fibers involved in the bradycardic response are not.Changes in mean arterial pressure (MAP), heart rate (HR), and vascular resistance (hindquarter and mesenteric territories) in response to electrical stimulation (ES) of the aortic depressor nerve (ADN) were evaluated in conscious freely moving rats. Platinum electrodes were implanted into the ADN of all rats studied, and some of these animals were also implanted with miniaturized Doppler probes around the superior mesenteric artery and inferior abdominal aorta (hindquarter). In both groups, the femoral artery and vein were catheterized one day before the experiments. In the first group of rats (n = 7), the control ES of the ADN in the range from 0.5 to 3.0 V (50 Hz, 10 ms) produced bradycardia and hypotension in an intensity-dependent manner, and treatment with methylatropine (intravenously) blocked the bradycardia but produced no significant changes in the hypotensive response. In a second group (n = 6), ES of the ADN was performed with the intensity fixed at 3 V and the frequency of the stimuli varying from 10 to 50 Hz. In this group, the hypotensive response was frequency dependent, whereas the bradycardic response was not. In a third group of rats (n = 6), ES of the ADN (2.5 V) produced hypotension (-35 +/- 4 mmHg), minor changes in the mesenteric (+5 +/- 14%), and vasodilation in hindquarter (-32 +/- 6%) vascular beds. The data show that 1) ES of the ADN produces a fall in pressure, bradycardia, vasodilation in the hindquarter, and no changes in the mesenteric vascular resistance, 2) methylatropine blocked the bradycardia and produced no effect on the hypotensive response to ES of the ADN, and 3) the baroreceptor afferent fibers involved in the hypotensive response to ES of ADN are sensitive to the variation of the frequency of the stimuli, whereas the fibers involved in the bradycardic response are not.

Arterial Baroreceptors and Experimental Diabetes

Abstract: Alterations of the autonomic reflex control of the cardiovascular system have been demonstrated in clinical and animal models of insulin‐dependent diabetes mellitus. Established neuroaxonal dystrophy is considered the neuropathological hallmark of chronic experimental diabetes. However, the afferent arm of the arterial baroreflex, that is, the carotid sinus nerve and the aortic depressor nerve, has received much less attention in studies dealing with this physiopathological model. The attenuation of the pressure response to bilateral carotid occlusion in conscious rats indicates a derangement of the baroreflex, probably involving an alteration of the carotid sinus nerve. There is histological evidence obtained from adult spontaneous insulin‐dependent diabetic rats (strain BB/S) of a carotid sinus nerve with signs of axonal swelling and intramyelinic edema, suggesting diabetic neuropathy. The study of aortic baroreceptor activity in anesthetized rats with short‐ and long‐term streptozotocin diabetes by means of cross‐spectral analysis of baroreceptor activity versus arterial pressure revealed a dysfunction in the afferent arm of the baroreflex even during a short period of diabetes. The morphology of the aortic depressor nerve of streptozotocin‐diabetic rats indicated axonal atrophy by visual analysis remarkably at the distal segments of the nerves. This finding was confirmed by morphometric study of the myelinated fibers. In conclusion, although studies of the arterial baroreceptors related to experimental diabetes are scanty in the literature, there is electrophysiological and histological evidence demonstrating that the carotid sinus and the aortic depressor nerves are abnormal in this experimental model.

Morphology and morphometry of the vagus nerve in male and female spontaneously hypertensive rats

The vagus nerve is an important component of the efferent arm of the baroreflex. Blood pressure levels as well as baroreflex control of circulation are significantly different in male and female spontaneously hypertensive rats (SHR). We proposed to investigate the morphometric differences between genders using the vagus nerve of SHR. Adult animals (20 weeks old) were anesthetized and had their arterial pressure (AP) and heart rate (HR) recorded by a computerized system. The rats were then systemically perfused with a fixative solution and had their cervical vagi nerves prepared for light microscopy. Proximal and distal segments of the left and right vagi nerves were evaluated for morphometric parameters including fascicle area and diameter, myelinated fiber number, density, area and diameter. Comparisons were made between sides and segments on the same gender as well as between genders. Differences were considered significant when p<0.05. Male SHR had significantly higher AP and HR. Morphometric data showed no differences between the same levels of both sides and between segments on the same side for male and female rats. In addition, no significant morphometric differences were observed when genders were compared. This is the first description of vagus nerve morphometry in SHR indicating that gender differences in AP and HR cannot be attributed to dissimilarities in vagal innervation of the heart. These data provide a morphological basis for further studies involving functional investigations of the efferent arm of the baroreflex in hypertension.

The treatment with pyridostigmine improves the cardiocirculatory function in rats with chronic heart failure

Sympathetic hyperactivity and its outcome in heart failure have been thoroughly investigated to determine the focus of pharmacologic approaches targeting the sympathetic nervous system in the treatment of this pathophysiological condition. On the other hand, therapeutic approaches aiming to protect the reduced cardiac parasympathetic function have not received much attention. The present study evaluated rats with chronic heart failure (six to seven weeks after coronary artery ligation) and the effects of an increased parasympathetic function by pyridostigmine (an acetylcholinesterase inhibitor) on the following aspects: arterial pressure (AP), heart rate (HR), baroreceptor and Bezold-Jarisch reflex, pulse interval (PI) and AP variability, cardiac sympathetic and parasympathetic tonus, intrinsic heart rate (i-HR) and cardiac function. Conscious rats with heart failure exhibited no change in HR, Bezold-Jarisch reflex, PI variability and cardiac sympathetic tonus. On the other hand, these animals presented hypotension and reduced baroreflex sensitivity, power in the low frequency (LF) band of the systolic AP spectrum, cardiac parasympathetic tonus and i-HR, while anesthetized rats exhibited reduced cardiac performance. Pyridostigmine prevented the attenuation of all the parameters examined, except basal AP and cardiac performance. In conclusion, the blockade of acetylcholinesterase with pyridostigmine was revealed to be an important pharmacological approach, which could be used to increase parasympathetic function and to improve a number of cardiocirculatory parameters in rats with heart failure.