Martin Appalsamy

Endothelin as a neuropeptide. Cardiovascular effects in the brainstem of normotensive rats.

The relevance of endothelin in central cardiovascular function was studied in urethane-anesthetized Sprague-Dawley rats. Blood pressure (BP) was monitored intra-arterially, and cerebrospinal fluid (CSF) was collected through an intracisternal catheter for radioimmunoassay of endothelin-1 (ET-1). Endothelin levels in the CSF were significantly higher (39 +/- 3 pg/ml) than in plasma (10 +/- 3 pg/ml, n = 11). ET-1 in CSF or plasma was not affected by systemic infusion of saline, but its levels significantly decreased when a sustained increase in BP was elicited with phenylephrine (14 +/- 7 pg/ml in the CSF and 6 +/- 4 pg/ml in plasma, n = 5). In sinoaortic-denervated animals, phenylephrine failed to reduce CSF endothelin levels. In different experiments, intracisternal administration of ET-1 (10 pmol) evoked an initial decrease in BP and heart rate (HR), followed by pronounced hypertension, bradycardia, and, in 70% of the animals, death from cardiorespiratory failure. Intracisternal administration of endothelin-3 (ET-3, 80 pmol, n = 11) evoked only a modest hypotensive and bradycardic response without cardiorespiratory impairment. Microinjection of ET-1 (0.5, 1, 2, 4, and 6 pmol/60 nl) into the nucleus of the solitary tract or area postrema produced a decrease in BP and HR. On the other hand, injection of low concentrations of ET-3 into the nucleus of the solitary tract increased BP and HR (at 2 pmol, 17 +/- 3 mm Hg, 14 +/- 6 beats per minute, n = 7), whereas ET-3 in the area postrema produced a prominent dose-related decrease in BP and HR. In the rostroventrolateral medulla, the lowest doses of ET-1 first modestly increased BP and renal sympathetic nerve activity. These effects were followed by hypotension, bradycardia, increase in respiratory frequency, and further enhancement of sympathetic nerve traffic. In 29% of the animals, these effects were followed by cardiorespiratory arrest. The specificity of the cardiovascular response to endothelin was demonstrated by the inhibitory effects of the receptor antagonist BQ-123. These results demonstrate that endothelin has specific cardiovascular effects in the brainstem of the rat and support a role for endothelin in cardiovascular regulation.

Cardiovascular effects of neuropeptide Y in rat brainstem nuclei.

Central catecholaminergic neurons are involved in cardiovascular regulation. Neuropeptide Y (NPY) coexists with adrenaline and noradrenaline in the rat brain, and interactions among these substances have been studied. The purpose of this study was to investigate the possible role of NPY in central cardiovascular control. Male Sprague-Dawley rats were anesthetized with urethane, and blood pressure was monitored intra-arterially. Intramedullary microinjection (60 nl) of NPY (0, 46.5 fmol, 465 fmol, 1.5 pmol, and 4.65 pmol) was made into the nucleus tractus solitarii (NTS), into the area postrema, and into the C1 area in the rostroventrolateral medulla. Injection site was identified by L-glutamate administration and confirmed histologically. Unilateral injection of NPY into the NTS produced a prominent dose-related decrease in heart rate and systolic and diastolic blood pressure (-106±8 beats/min, -56±2 mm Hg, and -33±2 mm Hg, respectively after 4.65 pmol NPY, n=7, p<0.001). Maximal changes occurred at 30 seconds and recovered in 10 minutes for blood pressure and 20 minutes for heart rate. Injection into the area postrema produced an initial increase in heart rate and mean blood pressure (+23 ±2 beats/min and +18 ±2 mm Hg) followed by a prolonged decrease in heart rate and mean blood pressure (-14±4 beats/min and -15±2 nun Hg, respectively, n=7, /?<0.01). However, injection of NPY into the Cl area produced a dose-related increase in blood pressure and a decrease in heart rate (+17±1 nun Hg and -23±8 beats/min, n=6, p<0.01). To test the specific effect of NPY, rabbit anti-NPY antiserum was used. Prior administration of the antiserum inhibited the effects of subsequent administration of NPY, whereas inactivated antiserum was not able to prevent these effects. These studies demonstrate that NPY has powerful cardiovascular effects in the NTS, area postrema, and Cl area of the medulla oblongata of the rat. This finding suggests that NPY may be important in cardiovascular regulation.

Portal Osmopressor Mechanism Linked to Transient Receptor Potential Vanilloid 4 and Blood Pressure Control

Human subjects with impaired baroreflex function cannot buffer rises or falls in blood pressure (BP), thus allowing BP effects of endogenous or environmental stimuli that previously escaped detection to emerge dramatically. Studies in these patients led us to discover that water ingestion induced a robust increase in BP and vascular resistance. Here, using a mouse model of baroreflex impairment, we show that the increase in blood pressure after water ingestion is mediated through sympathetic nervous system activation and that the osmosensitive transient receptor potential vanilloid 4 channel (Trpv4) is an essential component of the response. Although portal osmolality decreases after water ingestion in both wild-type and Trpv4−/− mice, only the wild-type animals show a pressor response. The same volume of physiological saline does not elicit an increase in BP, suggesting osmolality as the stimulus. The osmopressor response to water, and Trpv4 thus represent new factors now implicated in the physiology of BP regulation.

Modulatory effects of adenosine on baroreflex activation in the brainstem of normotensive rats

The effects of the adenosine antagonists, 1,3-dipropyl-8-p-sulphenylxanthine (DPSPX) and caffeine, on baroreflex activity were tested in normotensive Sprague-Dawley rats. The microinjection of DPSPX (0.92 nmol) into the nucleus tractus solitarii (NTS) of urethane-anesthetized animals did not modify basal blood pressure or heart rate but inhibited the reflex bradycardia elicited by phenylephrine. Similar inhibitory effects on baroreflex activation were observed after intracisternal administration of caffeine to conscious or anesthetized animals. These results suggest that central endogenous adenosine is involved in the medullary regulation of blood pressure and that adenosine antagonists such as caffeine can inhibit baroreflex activation.

Smoking and mechanisms of cardiovascular control

In humans short-term administration of nicotine, whether by smoking or intravenous injection, will typically raise blood pressure by 5 to 10 mm Hg and heart rate by 10 to 25 bpm. Smoking causes reduced myocardial contractility and left ventricular function in patients with angina pectoris or heart failure. Nicotines mechanism of action is more complex than the classic concept of nicotinic ganglionic stimulation can account for. Nicotine exerts a potent pressor effect in the ventral lateral medulla (C-1 area). Little current data are available documenting the efficacy of centrally acting antihypertensive agents and converting-enzyme inhibitors with regard to preventing nicotines acute cardiovascular effects.

Norepinephrine Transporter–Deficient Mice Exhibit Excessive Tachycardia and Elevated Blood Pressure With Wakefulness and Activity

Background—Norepinephrine (NE) is a primary neurotransmitter of central autonomic regulation and sympathetic nerve conduction, and the norepinephrine transporter (NET) is crucial in limiting catecholaminergic signaling. NET is sensitive to antidepressants, cocaine, and amphetamine. NET blockade often is associated with cardiovascular side effects, and NET deficiency is linked to tachycardia in familial orthostatic intolerance. Methods and Results—We telemetrically monitored NET-deficient (NET−/−) mice to determine the cardiovascular effects of reduced NE reuptake. Mean arterial pressure was elevated in resting NET−/− mice compared with NET+/+ controls (103±0.6 versus 99±0.4 mm Hg; P<0.01), and corresponding pressures increased to 122±0.3 and 116±0.3 mm Hg (P<0.0001) with activity. Heart rate was also greater in resting NET−/− mice (565±5 versus 551±3 bpm; P<0.05), and genotypic differences were highly significant during the active phase (640±5 versus 607±3 bpm; P<0.0001). Conversely, the respiratory rate of resting NET−/− mice was dramatically reduced, whereas increases after the day/night shift surpassed those of controls. Plasma catecholamines in NET−/− and NET+/+ mice were as follows: NE, 69±8 and 32±7; dihydroxyphenylglycol, 2+0.4 and 17±3; epinephrine, 15±3 and 4±0.6; and dopamine, 13±4 and 4±1 pmol/mL. Catechols in urine, brain, and heart also were determined. Conclusions—Resting mean arterial pressure and heart rate are maintained at nearly normal levels in NET-deficient mice, most likely as a result of increased central sympathoinhibition. However, sympathetic activation with wakefulness and activity apparently overwhelms central modulation, amplifying peripheral catecholaminergic signaling, particularly in the heart.

Norepinephrine transporter-deficient mice respond to anxiety producing and fearful environments with bradycardia and hypotension

The study of anxiety and fear involves complex interrelationships between psychiatry and the autonomic nervous system. Altered noradrenergic signaling is linked to certain types of depression and anxiety disorders, and treatment often includes specific transporter blockade. The norepinephrine transporter is crucial in limiting catecholaminergic signaling. Norepinephrine transporter-deficient mice have increased circulating catecholamines and elevated heart rate and blood pressure. We hypothesized, therefore, that reduced norepinephrine clearance would heighten the autonomic cardiovascular response to anxiety and fear. In separate experiments, norepinephrine transporter-deficient (norepinephrine transporter-/-) mice underwent tactile startle and trace fear conditioning to measure hemodynamic responses. A dramatic tachycardia was observed in norepinephrine transporter-/- mice compared with controls following both airpuff or footshock stimuli, and pressure changes were also greater. Interestingly, in contrast to normally elevated home cage levels in norepinephrine transporter-deficient mice, prestimulus heart rate and blood pressure were actually higher in norepinephrine transporter+/+ animals throughout behavioral testing. Upon placement in the behavioral chamber, norepinephrine transporter-deficient mice demonstrated a notable bradycardia and depressor effect that was more pronounced in females. Power spectral analysis indicated an increase in low frequency oscillations of heart rate variability; in mice, suggesting increased parasympathetic tone. Finally, norepinephrine transporter-/- mice exhibited sexual dimorphism in freeze behavior, which was greatest in females. Therefore, while reduced catecholamine clearance amplifies immediate cardiovascular responses to anxiety- or fear-inducing stimuli in norepinephrine transporter-/- mice, norepinephrine transporter deficiency apparently prevents protracted hemodynamic escalation in a fearful environment. Conceivably, chronic norepinephrine transporter blockade with transporter-specific drugs might attenuate recognition of autonomic and somatic distress signals in individuals with anxiety disorders, possibly lessening their behavioral reactivity, and reducing the cardiovascular risk factors associated with persistent emotional arousal.

Tachycardia, reduced vagal capacity, and age-dependent ventricular dysfunction arising from diminished expression of the presynaptic choline transporter

Healthy cardiovascular function relies on a balanced and responsive integration of noradrenergic and cholinergic innervation of the heart. High-affinity choline uptake by cholinergic terminals is pivotal for efficient ACh production and release. To date, the cardiovascular impact of diminished choline transporter (CHT) expression has not been directly examined, largely due to the transporters inaccessibility in vivo. Here, we describe findings from cardiovascular experiments using transgenic mice that bear a CHT genetic deficiency. Whereas CHT knockout (CHT(-/-)) mice exhibit early postnatal lethality, CHT heterozygous (CHT(+/-)) mice survive, grow, and reproduce normally and exhibit normal spontaneous behaviors. However, the CHT(+/-) mouse heart displays significantly reduced levels of high-affinity choline uptake accompanied by significantly reduced levels of ACh. Telemeterized recordings of cardiovascular function in these mice revealed tachycardia and hypertension at rest. After treadmill exercise, CHT(+/-) mice exhibited slower heart rate recovery, consistent with a diminished cholinergic reserve, a contention validated through direct vagal nerve stimulation. Echocardiographic and histological experiments revealed an age-dependent decrease in fractional shortening, increased left ventricular dimensions, and increased ventricular fibrosis, consistent with ventricular dysfunction. These cardiovascular phenotypes of CHT(+/-) mice encourage an evaluation of humans bearing reduced CHT expression for their resiliency in maintaining proper heart function as well as risk for cardiovascular disease.

Modulatory effects of endothelin on baroreflex activation in the nucleus of the solitary tract

In this study, we determine the effects of endogenous endothelin on baroreflex activation. After control baroreflex slopes were obtained, the animals received bilateral intra-nucleus tractus solitarii microinjections of saline, or equimolar doses (4 pmol/60 nl) of the endothelin ETA receptor antagonist cyclo (D-Trp-D-Asp-Pro-Val-Leu (BQ-123), Homopiperinidinyl-CO-Leu-D-Trp(CHO)-D-Trp-OH (BQ-610), or the endothelin ETB receptor antagonist N-cis-2,6-dimethylpiperidinocarbonyl-L-gamma-MeLeu-D-Trp( COOCH3)-D-Nle (BQ-788). Intra-nucleus tractus solitarii administration of BQ-123 resulted in a brief initial pressor effect followed by hypotension which resolved by 15 min. The baroreflex slope was significantly enhanced when tested 15 min after BQ-123 treatment (from 2.4 +/- 0.5 ms/mmHg to 3.5 +/- 0.4 ms/mmHg). Similar effects were observed with the other endothelin ETA receptor antagonist, except that the hypertensive and hypotensive responses were more pronounced while the baroreflex slope was similarly increased. In contrast, the endothelin ETB receptor antagonist did not evoke appreciable changes in hemodynamics or in baroreflex slopes. Our results support the concept that endothelin prominently affects reflex cardiovascular function through the endothelin ETA receptor subtype.

Animal Model of Neuropathic Tachycardia Syndrome

Abstract—Clinically relevant autonomic dysfunction can result from either complete or partial loss of sympathetic outflow to effector organs. Reported animal models of autonomic neuropathy have aimed to achieve complete lesions of sympathetic nerves, but incomplete lesions might be more relevant to certain clinical entities. We hypothesized that loss of sympathetic innervation would result in a predicted decrease in arterial pressure and a compensatory increase in heart rate. Increased heart rate due to loss of sympathetic innervation is seemingly paradoxical, but it provides a mechanistic explanation for clinical autonomic syndromes such as neuropathic postural tachycardia syndrome. Partially dysautonomic animals were generated by selectively lesioning postganglionic sympathetic neurons with 150 mg/kg 6-hydroxydopamine hydrobromide in male Sprague-Dawley rats. Blood pressure and heart rate were monitored using radiotelemetry. Systolic blood pressure decreased within hours postlesion (&Dgr;>20 mm Hg). Within 4 days postlesion, heart rate rose and remained elevated above control levels. The severity of the lesion was determined functionally and pharmacologically by spectral analysis and responsiveness to tyramine. Low-frequency spectral power of systolic blood pressure was reduced postlesion and correlated with the diminished tyramine responsiveness (r =0.9572, P =0.0053). The tachycardia was abolished by treatment with the &bgr;-antagonist propranolol, demonstrating that it was mediated by catecholamines acting on cardiac &bgr;-receptors. Partial lesions of the autonomic nervous system have been hypothesized to underlie many disorders, including neuropathic postural tachycardia syndrome. This animal model may help us better understand the pathophysiology of autonomic dysfunction and lead to development of therapeutic interventions.