Jaimie W. Polson

Central mechanisms underlying short- and long-term regulation of the cardiovascular system.

1. Sympathetic vasomotor nerves play a major role in determining the level of arterial blood pressure and the distribution of cardiac output. The present review will discuss briefly the central regulatory mechanisms that control the sympathetic outflow to the cardiovascular system in the short and long term.

Functional organization of brain pathways subserving the baroreceptor reflex: studies in conscious animals using immediate early gene expression.

Abstract1. This paper reviews studies carried out in our laboratory in which we have used the c-fos functional mapping method, in combination with other methods, to determine the functional organization of central baroreceptor pathways as they operate in the conscious rabbit.2. First, we showed that periods of induced hypertension or hypotension each result in a specific and reproducible pattern of activation of neurons in the brainstem and forebrain. In particular, hypotension (but not hypertension) results in the activation of catecholamine neurons in the medulla and pons and vasopressin-synthesizing neurons in the hypothalamus.3. The activation of medullary cell groups in response to induced hypertension or hypotension in the conscious rabbit is almost entirely dependent on inputs from arterial baroreceptors, while the activation of hypothalamic vasopressin-synthesising neurons in response to hypotension is largely dependent on baroreceptors, although an increase in circulating angiotensin also appears to contribute.4. Discrete groups of neurons in the rostral ventrolateral medulla (RVLM) and A5 area in the pons are the major groups of spinally projecting neurons activated by baroreceptor unloading. In contrast, spinally projecting neurons in the paraventricular nucleus in the hypothalamus appear to be largely unaffected by baroreceptor signals.5. Direct afferent inputs to RVLM neurons in response to increases or decreases in arterial pressure originate primarily from other medullary nuclei, particularly neurons located in the caudal and intermediate levels of the ventrolateral medulla (CVLM and IVLM), as well as in the nucleus tractus solitarius (NTS).6. There is also a direct projection from barosensory neurons in the NTS to the CVLM/IVLM region, which is activated by baroreceptor inputs.7. Collectively, the results of our studies in conscious animals indicate that baroreceptor signals reach all levels of the brain. With regard to the baroreceptor reflex control of sympathetic activity, our studies are consistent with previous studies in anesthetized animals, but in addition reveal other previously unrecognized pathways that also contribute to this reflex regulation.

Cardiovascular Responses Evoked by Leptin Acting on Neurons in the Ventromedial and Dorsomedial Hypothalamus

Abstract—Leptin, a circulating hormone produced by adipose tissue, is believed to act on the hypothalamus to increase sympathetic vasomotor activity, in addition to its well-known effects on appetite and energy expenditure. In this study, we determined the cardiovascular effects of direct application of leptin to specific cell groups within the hypothalamus that are known to be activated by circulating leptin. In rats anesthetized with urethane, microinjections of leptin (16 ng in 20 nL solution) were made into the ventromedial hypothalamic nucleus, dorsomedial hypothalamic nucleus, and paraventricular nucleus. Compared with vehicle solution, microinjections of leptin into the ventromedial hypothalamic nucleus evoked significant increases in arterial pressure and renal sympathetic nerve activity, but not heart rate. In contrast, microinjections of leptin into the dorsomedial hypothalamic nucleus evoked significant increases in arterial pressure and heart rate but not renal sympathetic nerve activity, whereas microinjections of leptin into the paraventricular nucleus had no significant effect on any of the measured cardiovascular variables. These results indicate that the ventromedial and dorsomedial hypothalamic regions might be important sites at which leptin activation leads to increases in sympathetic vasomotor activity and heart rate, as occurs in obesity-related hypertension.

Hypoxia-induced Fos expression in neurons projecting to the pressor region in the rostral ventrolateral medulla

Previous studies in anaesthetized animals have shown that the hypoxia-induced increase in sympathetic vasomotor activity is largely dependent on synaptic excitation of sympathoexcitatory pressor neurons in the rostral part of the ventrolateral medulla. The primary aim of this study was to determine, in conscious rabbits, the distribution of neurons within the brain that have properties characteristic of interneurons conveying excitatory inputs to the rostral ventrolateral medullary pressor region in response to systemic hypoxia. In a preliminary operation, a retrogradely-transported tracer, fluorescent-labelled microspheres, was injected into the physiologically-identified pressor region in the rostral ventrolateral medulla. After a waiting period of one to two weeks, the conscious rabbits were subjected to moderate hypoxia (induced by breathing 10% O2 in N2) for a period of 60 min. Control groups of animals were exposed to room air or to mild hypoxia (12% O2 in N2). Moderate hypoxia resulted in a modest hypertension of approximately 15 mmHg, and in the expression of Fos (a marker of neuronal activation) in many neurons in the nucleus tractus solitarius, the rostral, intermediate and caudal parts of the ventrolateral medulla, the Kölliker-Fuse nucleus, locus coeruleus, subcoeruleus and A5 area in the pons as well as in several midbrain and forebrain regions, including the periaqueductal grey in the midbrain and the paraventricular, supraoptic and arcuate nuclei in the hypothalamus. Fos expression was also observed in these regions in rabbits subjected to mild hypoxia or normoxia, but it was much reduced compared to rabbits subjected to moderate hypoxia. Approximately half of the neurons in the ventrolateral medulla, 27% of neurons in the nucleus tractus solitarius, and 49-81% of neurons in the locus coeruleus, sub-coeruleus and A5 area that expressed Fos following moderate hypoxia were also immunoreactive for tyrosine hydroxylase, and were therefore catecholamine cells. Approximately half of the neurons in the nucleus tractus solitarius and two-thirds of neurons in the Kölliker-Fuse nucleus that expressed Fos following moderate hypoxia were retrogradely labelled from the rostral ventrolateral medullary pressor region. Similarly, approximately one quarter of Fos-positive cells in the caudal and intermediate ventrolateral medulla were retrogradely labelled, but very few Fos-positive/retrogradely-labelled cells were found in other pontomedullary or suprapontine brain regions. The results indicate that systemic hypoxia results in activation of neurons in several discrete nuclei in the brainstem and forebrain, including neurons in all the major pontomedullary catecholamine cell groups. However, neurons that are activated by systemic hypoxia and that also project to the rostral ventrolateral medullary pressor region are virtually confined to the lower brainstem, primarily in the nucleus tractus solitarius and Kölliker-Fuse nucleus and to a lesser extent the caudal/intermediate ventrolateral medulla. In a previous study from our laboratory, we determined the distribution of neurons in the brainstem that are activated by hypertension and that also project to the rostral ventrolateral medullary pressor region. [Polson et al. (1995) Neuroscience 67, 107-123]. Comparison of the present results with those from this previous study indicates that the hypoxia-activated neurons in the nucleus tractus solitarius and Kölliker-Fuse nucleus that project to the rostral ventrolateral medulla are likely to be interneurons conveying excitatory chemoreceptor signals, while those in the caudal/intermediate ventrolateral medulla are likely to be mainly interneurons conveying inhibitory baroreceptor signals, activated by the rise in arterial blood pressure associated with the hypoxia-induced hypertension.

Pressor and sympathoexcitatory effects of nitric oxide in the rostral ventrolateral medulla.

Objective It has been shown that nitric oxide (NO) plays an important role in the central control of arterial pressure and sympathetic nerve activity. The aim of this study was to determine whether NO can regulate sympathetic nerve activity by an action on pressor neurons within the rostral part of the ventrolateral medulla (VLM). Design and methods Experiments were performed on anaesthetized rabbits with denervated arterial and cardiopulmonary baroreceptors. The mean arterial pressure (MAP), heart rate and renal sympathetic nerve activity were measured. Microinjections of the NO donors sodium nitroprusside (SNP, 4-50 nmol) and S-nitroso-glutathione (10 nmol), the NO precursor L-arginine (50 nmol) and the NO synthase inhibitor Nw-nitro-L-arginine methyl ester (LNAME, 50 nmol), were made into the functionally identified pressor region in the rostral VLM. The effects of SNP were also determined before and after injection of 5 nmol methylene blue into the same area. In control experiments, injections of D-arginine (50 nmol) and D-NAME (50 nmol), which are the inactive isomers of L-arginine and L-NAME, respectively, were also made into the functionally identified pressor region in the rostral VLM. Results Microinjections of SNP into the rostral VLM pressor region produced a dose-dependent increase in mean arterial pressure and renal sympathetic nerve activity. At the highest dose of 50 nmol, the increase in MAP was 26 ± 5 mmHg (P<0.001) and the sympathetic nerve activity was 53 ± 5% (P<0.001). These effects were abolished following methylene blue injection into the same region. Injection of 10 nmol S-nitroso-glutathion also produced increases in MAP (15 ± 2 mmHg, P<0.001) and in renal sympathetic nerve activity (28 ± 2%, P<0.001). Microinjections of L- or D-arginine resulted in very small depressor responses, but had no significant effect on renal sympathetic nerve activity. Microinjections of L-NAME, but not of D-NAME, caused significant decreases in MAP (19 ± 1 mmHg, P<0.001) and in sympathetic nerve activity (30 ± 3%, P<0.001). Conclusions The results indicate that, in the anaesthetized rabbit with denervated baroreceptors, NO has a pressor and sympathoexcitatory action in the rostral VLM, which is mediated by a cyclic GMP-dependent mechanism. Second, endogenous NO may modulate sympathetic activity tonically, by a direct or indirect action on sympathoexcitatory neurons within the rostral VLM.

Effects of sinoaortic denervation on Fos expression in the brain evoked by hypertension and hypotension in conscious rabbits.

We have previously shown [Li and Dampney (1994) Neuroscience 61, 613-634] that periods of sustained hypertension and hypotension each induces a distinctive and reproducible pattern of neuronal expression of Fos (a marker of neuronal activation) in specific regions of the brainstem and forebrain of conscious rabbits. The aim of this study was to determine the contribution of afferent inputs from arterial baroreceptors to the activation of neurons in these various brain regions that is caused by a sustained change in arterial pressure. Experiments were carried out on rabbits in which the carotid sinus and aortic depressor nerves were cut in a preliminary operation. Following a recovery period of seven to 10 days, a moderate hypertension or hypotension (increase or decrease in arterial pressure of 20-30 mmHg) was induced in conscious barodenervated rabbits for 60 min by the continuous infusion of phenylephrine or sodium nitroprusside, respectively. In control experiments, barodenervated rabbits were subjected to the identical procedures except that they were infused with the vehicle solution alone. Compared with the effects seen in barointact rabbits, [Li and Dampney (1994) Neuroscience 61, 613-634] the number of neurons that expressed Fos in response to hypertension was reduced by approximately 90% in the nucleus of the solitary tract and in the caudal and intermediate parts of the ventrolateral medulla. In supramedullary regions, baroreceptor denervation resulted in a reduction of approximately 60% in hypertension-induced Fos expression in the central nucleus of the amygdala and in the bed nucleus of the stria terminalis, but no significant reduction in the parabrachial complex in the pons. Following hypotension, the number of neurons that expressed Fos in barodenervated rabbits, compared with barointact rabbits, [Li and Dampney (1994) Neuroscience 61, 613-634] was reduced by approximately 90% in the nucleus of the solitary tract, area postrema, and caudal, intermediate and rostral parts of the ventrolateral medulla. Baroreceptor denervation also resulted in a similar large reduction in hypotension-induced Fos expression in many supramedullary regions (locus coeruleus, midbrain periaqueductal grey, hypothalamic paraventricular nucleus, and in the central nucleus of the amygdala and the bed nucleus of the stria terminalis in the basal forebrain). In the supraoptic nucleus, hypotension-induced Fos expression in barodenervated rabbits was reduced by 75% compared to barointact animals, but was still significantly greater than in control animals. There was also a high level of Fos expression, much greater than in control animals, in the circumventricular organs surrounding the third ventricle (subfornical organ and organum vasculosum lamina terminalis). The results indicate that in conscious rabbits the activation of neurons that occurs in several discrete regions at all levels of the brain following a sustained change in arterial pressure is largely dependent upon inputs from arterial baroreceptors, with the exception of neurons in the circumventricular organs surrounding the third ventricle that are activated by sustained hypotension. The latter group of neurons are known to project to vasopressin-secreting neurons in the supraoptic nucleus, and may therefore via this pathway trigger the hypotension-induced release of vasopressin that occurs in the absence of baroreceptor inputs.

Use of c-fos functional mapping to identify the central baroreceptor reflex pathway: advantages and limitations.

Prolonged stimulation of many neurons results in the expression of the immediate early gene c-fos, which in turn cause the production of the protein Fos, whose presence in a cell can be detected by immunocytochemistry. This method has been used in both conscious and anaesthetized animals to identify central neurons involved in the baroreceptor reflex. In this paper we review the factors that can influence c-fos expression, with particular emphasis on the effects of different anaesthetic agents. We conclude that the c-fos method of functional mapping, when applied carefully and critically, is a very useful method of identifying central neurons that are activated by cardiovascular stimuli in conscious animals. Anaesthetic agents can significantly alter c-fos expression, and this effect differs greatly according to the type of anaesthetic used.

Automation of analysis of cardiovascular autonomic function from chronic measurements of arterial pressure in conscious rats

At present, there is no single software package that provides a comprehensive power spectral analysis of pulse interval (PI) and arterial blood pressure (BP), spontaneous cardiac baroreceptor reflex gain (sBRG) and respiratory rate. Furthermore, scientific validation of the software that is currently commercially available and employed has not been published. We introduce ‘Hey‐Presto’ software, which fully evaluates cardiovascular autonomic function from the BP signal obtained from rats. The program performs power spectral analysis of HR and BP variability, respiratory rate and, based on a time‐series method, spontaneous cardiac baroreceptor (sBRG). We have validated Hey‐Presto with conventional pharmacological agents to block cardiac vagal and cardiac sympathetic transmission in conscious rats fitted with a radio‐telemetery BP transducer. Following administration of atropine (1 mg kg−1, i.v.), high‐frequency (HF) power of the PI decreased (P < 0.01) and was associated with the expected increase in HR. Subsequent cardiac sympathetic blockade (atenolol, 1 mg kg−1, i.v.) reduced the low frequency (LF) to HF ratio (LF:HF) of the PI (P < 0.01), which was consistent with the observed reduction in HR. We also found that alterations in sBRG after blockade of cardiac autonomic transmission were highly comparable to values computed manually using vasoactive drugs administered intravenously. The software also detected circadian rhythms in sBRG, HF component of the PI, LF:HF of the PI and LF component of the BP as well as BP and HR during continuous 24 h recording. By demonstrating its application to humans, we found appropriate changes in the power of PI and the LF power of the BP during postural changes. These results demonstrate that Hey‐Presto allows a fully automated, reliable, fast and comprehensive evaluation of cardiovascular autonomic function based on chronic measurements of BP in rats. Moreover, we have confirmed its versatility by demonstrating its application to man.

Fos expression in neurons projecting to the pressor region in the rostral ventrolateral medulla after sustained hypertension in conscious rabbits.

Previous studies in anaesthetized animals have shown that the baroreflex control of sympathetic vasomotor activity is mediated to a large extent by inhibitory inputs to sympathoexcitatory pressor neurons in the rostral part of the ventrolateral medulla. The aim of this study was to determine, in conscious rabbits, the distribution of neurons within the brain that have two properties characteristic of interneurons conveying baroreceptor signals to the rostral ventrolateral medulla: (i) they are activated by an increase in arterial pressure; and (ii) they project specifically to the rostral ventrolateral medulla pressor region. In a preliminary operation, an injection of the retrogradely transported tracer, fluorescent-labelled microspheres, was made into the physiologically identified pressor region in the rostral ventrolateral medulla. After a waiting period of one to eight weeks, hypertension was produced in the conscious rabbit by continuous intravenous infusion of phenylephrine at a rate sufficient to increase arterial pressure by approximately 20 mmHg, maintained for a period of 60 min. A control group of animals was infused with the vehicle solution alone. In confirmation of our previous study, hypertension produced by phenylephrine resulted in the neuronal expression of Fos (a marker of neuronal activation) in the nucleus of the solitary tract, area postrema, the intermediate and caudal parts of the ventrolateral medulla parabrachial complex, and in the central nucleus of the amygdala. Approximately 50% of the Fos-immunoreactive neurons in both the caudal and intermediate parts of the ventrolateral medulla were also retrogradely labelled from the rostral ventrolateral medulla pressor region; such double-labelled neurons were confined to a discrete longitudinal column located just ventrolateral to the nucleus ambiguus. Significant numbers of double-labelled neurons were also found in the nucleus of the solitary tract and area postrema, although these represented a much lower proportion (13-16%) of the total number of Fos-immunoreactive neurons in these regions. In the parabrachial complex, Fos-immunoreactive and retrogradely labelled neurons were largely separate populations, while in the amygdala they were entirely separate populations. In the control group of rabbits, virtually no double-labelled neurons were found in any of these regions. The results indicate that putative baroreceptor interneurons that project to the pressor region of the rostral ventrolateral medulla are virtually confined to the lower brainstem. In particular, they support the results of previous studies in anaesthetized animals indicating that neurons in the intermediate and caudal ventrolateral medulla convey baroreceptor signals to the rostral ventrolateral medulla pressor region, and extend them by demonstrating the precise anatomical distribution of these neurons.(ABSTRACT TRUNCATED AT 400 WORDS)

Signalling across the blood brain barrier by angiotensin II: novel implications for neurogenic hypertension

Angiotensin II (AngII) is a major culprit in essential hypertension. Based on a genetic rodent model of hypertension, we review here evidence that AngII may signal across the blood brain barrier to affect neuronal circuits within the nucleus tractus solitarii (NTS) of the brainstem, a pivotal region regulating both the baroreceptor reflex and set point control of arterial pressure. We have termed this form of signalling as vascular–neuronal signalling. We describe that the depressant action of AngII in NTS on the baroreceptor reflex is mediated via activation of endothelial nitric oxide synthase (eNOS) releasing NO that promotes release of the inhibitory transmitter—GABA. This could shunt the incoming excitatory baroreceptor afferent traffic impinging on NTS neurones. Chronic studies recording arterial pressure in conscious unrestrained rats using radio-telemetry have revealed that eNOS in NTS plays an endogenous physiological role in the homeostatic regulation of the gain of the cardiac baroreceptor reflex. However, in the spontaneously hypertensive rat, eNOS mRNA was higher (compared to normotensive rats), and its chronic blockade in NTS restored the abnormally depressed cardiac baroreceptor reflex to levels akin to normotensive rats, improved heart rate variability and lowered arterial pressure. Hence, it seems that excessive eNOS activity in NTS of the SHR contributes to the pathological state of this animal model’s cardiovascular autonomic nervous system. We speculate on why eNOS activity may be up regulated in the NTS of the SHR and propose that it is a consequence of high cerebral vascular resistance and inadequate blood perfusion of the brainstem.