Effects of vasopressin during a pulmonary hypertensive crisis induced by acute hypoxia in a rat model of pulmonary hypertension.

BACKGROUND: A pulmonary hypertensive crisis (PHC) can be a life-threatening condition. We established a PHC model by exposing rats with monocrotaline (MCT)-induced pulmonary hypertension to acute hypoxia, and investigated the effects of vasopressin, phenylephrine, and norepinephrine on the PHC. METHODS: Four weeks after MCT 60 mg kg-1 administration i.v., right ventricular systolic pressure (RVSP), systolic BP (SBP), mean BP (MBP), cardiac index (CI), and pulmonary vascular resistance index (PVRI) were measured. PHC defined as an RVSP exceeding or equal to SBP was induced by changing the fraction of inspiratory oxygen to 0.1. Rats were subsequently treated by vasopressin, phenylephrine, or norepinephrine, followed by assessment of systemic haemodynamics, isometric tension of femoral and pulmonary arteries, cardiac function, blood gas composition, and survival. RESULTS: PHC was associated with increased RV dilatation and paradoxical septal motion. Vasopressin increased MBP [mean (standard error)] from 52.6 (3.8) to 125.0 (8.9) mm Hg and CI from 25.4 (2.3) to 40.6 (1.8) ml min-1 100 g-1 while decreasing PVRI. Vasopressin also improved RV dilatation, oxygenation, and survival in PHC. In contrast, phenylephrine increased MBP from 54.8 (2.3) to 96.8 (3.2) mm Hg without improving cardiac pump function. Norepinephrine did not alter MBP. Vasopressin contracted femoral but not pulmonary arteries, whereas phenylephrine contracted both arterial beds. Hence, improvements with vasopressin in PHC might be associated with decreased PVRI and selective systemic vasoconstriction. CONCLUSIONS: In this rat model of a PHC, vasopressin, but not phenylephrine or norepinephrine, resulted in better haemodynamic and vascular recovery.

PML-Nuclear Bodies Regulate the Stability of the Fusion Protein Dendra2-Nrf2 in the Nucleus.

BACKGROUND/AIMS: Nuclear factor erythroid 2-related factor 2 (Nrf2) is a basic leucine-zipper transcription factor essential for cellular responses to oxidative stress. Degradation of Nrf2 in the cytoplasm, mediated by Keap1-Cullin3/RING box1 (Cul3-Rbx1) E3 ubiquitin ligase and the proteasome, is considered the primary pathway controlling the cellular abundance of Nrf2. Although the nucleus has been implicated in the degradation of Nrf2, little information is available on how this compartment participates in degrading Nrf2. METHODS: Here, we fused the photoconvertible fluorescent protein Dendra2 to Nrf2 and capitalized on the irreversible change in color (green to red) that occurs when Dendra2 undergoes photoconversion to study degradation of Dendra2-Nrf2 in single live cells. RESULTS: Using this approach, we show that the half-life (t1/2) of Dendra2-Nrf2 in the whole cell, under homeostatic conditions, is 35 min. Inhibition of the proteasome with MG-132 or induction of oxidative stress with tert-butylhydroquinone (tBHQ) extended the half-life of Dendra2-Nrf2 by 6- and 28-fold, respectively. By inhibiting nuclear export using Leptomycin B, we provide direct evidence that degradation of Nrf2 also occurs in the nucleus and involves PML-NBs (Promyelocytic Leukemia-nuclear bodies). We further demonstrate that co-expression of Dendra2-Nrf2 and Crimson-PML-I lacking two PML-I sumoylation sites (K65R and K490R) changed the decay rate of Dendra2-Nrf2 in the nucleus and stabilized the nuclear derived Nrf2 levels in whole cells. CONCLUSION: Altogether, our findings provide direct evidence for degradation of Nrf2 in the nucleus and suggest that modification of Nrf2 in PML nuclear bodies contributes to its degradation in intact cells.

Limited recovery of pineal function after regeneration of preganglionic sympathetic axons: evidence for loss of ganglionic synaptic specificity.

The cervical sympathetic trunks (CSTs) contain axons of preganglionic neurons that innervate the superior cervical ganglia (SCGs). Because regeneration of CST fibers can be extensive and can reestablish certain specific patterns of SCG connections, restoration of end organ function would be expected. This expectation was examined with respect to the pineal gland, an organ innervated by the two SCGs. The activity of pineal serotonin N-acetyltransferase (NAT) exhibits a large circadian rhythm that is dependent on the sympathetic input of the gland, with high activity at night. Thirty-six hours after the CSTs were crushed bilaterally, nocturnal NAT was decreased by 99%. Three months later, enzyme activity had recovered only to 15% of control values, a recovery dependent on regeneration of CST fibers. Nevertheless, a small day/night rhythm was present in lesioned animals. Neither the density of the adrenergic innervation of the gland nor the ability of an adrenergic agonist to stimulate NAT activity was reduced in rats with regenerated CSTs. In addition, stimulation of the regenerated CST at a variety of frequencies was at least as effective in increasing NAT activity as seen with control nerves. These data suggest that the failure of pineal function to recover is not attributable to a quantitative deficit in the extent of reinnervation or synaptic efficacy. Rather, we suggest that there is some loss of specificity in the synaptic connections made in the SCG during reinnervation, resulting in a loss of the central neuronal information necessary for directing a normal NAT rhythm and thus normal pineal function.

[Effects of various neuromediators and regulatory peptides on the impulse activity of neurons in the lateral vestibular nucleus].

The myoelectrode technique and microiontophoresis of physiologically active substances were applied to cats immobilized with neuromuscular relaxant to show that the classic neuromediators (acetylcholine, norepinephrine, GABA etc.) and regulatory peptides (enkephalins, TRHs, vasoactive intestinal peptide (VIP), somatostatin (SS) and others) can influence directly most neurons (58 to 100%) in the lateral vestibular nucleus (LVN). Enkephalins, VIP and SS retained largely their inhibitory effect on the neuron impulse activity in the presence of L-glutamate. Also, enkephalins, VIP and SS are able to stimulate or suppress the inhibitory effect of GABA and glycine. Consequently, the substances under study may act as LVN neuromediators and/or neuromodulators.

Involvement of central alpha1-adrenoceptors on renal responses to central moxonidine and alpha-methylnoradrenaline.

Moxonidine (alpha2-adrenoceptor/imidazoline receptor agonist) injected into the lateral ventricle induces diuresis, natriuresis and renal vasodilation. Moxonidine-induced diuresis and natriuresis depend on central imidazoline receptors, while central alpha1-adrenoceptors are involved in renal vasodilation. However, the involvement of central alpha1-adrenoceptors on diuresis and natriuresis to central moxonidine was not investigated yet. In the present study, the effects of moxonidine, alpha-methylnoradrenaline (alpha2-adrenoceptor agonist) or phenylephrine (alpha1-adrenoceptor agonist) alone or combined with previous injections of prazosin (alpha1-adrenoceptor antagonist), yohimbine or RX 821002 (alpha2-adrenoceptor antagonists) intracerebroventricularly (i.c.v.) on urinary sodium, potassium and volume were investigated. Male Holtzman rats (n = 5-18/group) with stainless steel cannula implanted into the lateral ventricle and submitted to gastric water load (10% of body weight) were used. Injections of moxonidine (20 nmol) or alpha-methylnoradrenaline (80 nmol) i.c.v. induced natriuresis (196 +/- 25 and 171 +/- 30, respectively, vs. vehicle: 101 +/- 9 microEq/2 h) and diuresis (9.0 +/- 0.4 and 12.3 +/- 1.6, respectively, vs. vehicle: 5.2 +/- 0.5 ml/2 h). Pre-treatment with prazosin (320 nmol) i.c.v. abolished the natriuresis (23 +/- 4 and 76 +/- 11 microEq/2 h, respectively) and diuresis (5 +/- 1 and 7.6 +/- 0.8 ml/2 h, respectively) produced by i.c.v. moxonidine or alpha-methylnoradrenaline. RX 821002 (320 nmol) i.c.v. abolished the natriuretic effect of alpha-methylnoradrenaline, however, yohimbine (320 nmol) did not change renal responses to moxonidine. Phenylephrine (80 nmol) i.c.v. induced natriuresis and kaliuresis that were blocked by prazosin. Therefore, the present data suggest that moxonidine and alpha-methylnoradrenaline acting on central imidazoline receptors and alpha2-adrenoceptors, respectively, activate central alpha1-adrenergic mechanisms to increase renal excretion.

Direct evidence for imidazoline (I1) receptor modulation of ethanol action on norepinephrine-containing neurons in the rostral ventrolateral medulla in conscious spontaneously hypertensive rats.

BACKGROUND: Enhancement of the rostral ventrolateral medulla (RVLM) presympathetic (norepinephrine, NE) neuronal activity represents a neurochemical mechanism for the pressor effect of ethanol. In this study, we tested the hypothesis that ethanol action on RVLM presympathetic neurons is selectively influenced by the signaling of the local imidazoline (I1) receptor. To support a neuroanatomical and an I1-signaling selectivity of ethanol, and to circumvent the confounding effects of anesthesia, the dose-related neurochemical and blood pressure effects of ethanol were investigated in the presence of selective pharmacological interventions that cause reduction in the activity of RVLM or nucleus tractus solitarius (NTS) NE neurons via local activation of the I1 or the alpha2-adrenergic receptor in conscious spontaneously hypertensive rats. RESULTS: Local activation of the I1 receptor by rilmenidine (40 nmol) or by the I1/alpha2 receptor mixed agonist clonidine (1 nmol), and local activation of the alpha2-adrenergic receptor (alpha2AR) by the pure alpha2AR agonist alpha-methylnorepinephrine (alpha-MNE, 10 nmol) caused reductions in RVLM NE, and blood pressure. Intra-RVLM ethanol (1, 5, or 10 microg), microinjected at the nadir of the neurochemical and hypotensive responses, elicited dose-dependent increments in RVLM NE and blood pressure in the presence of local I1--but not alpha2-receptor activation. Only intra-NTS alpha-MNE, but not rilmenidine or clonidine, elicited reductions in local NE and blood pressure; ethanol failed to elicit any neurochemical or blood pressure responses in the presence of local activation of the alpha2AR within the NTS. CONCLUSION: The findings support the neuroanatomical selectivity of ethanol, and support the hypothesis that the neurochemical (RVLM NE), and the subsequent cardiovascular, effects of ethanol are selectively modulated by I1 receptor signaling in the RVLM.

Neuronal norepinephrine responses of the rostral ventrolateral medulla and nucleus tractus solitarius neurons distinguish the I1- from the alpha2-receptor-mediated hypotension in conscious SHRs.

We tested the hypothesis that the I1 receptor mediates the reduction in rostral ventrolateral medulla (RVLM) neuronal norepinephrine (NE; index of sympathetic activity) that leads to hypotension independent of other brainstem areas or the alpha2-adrenergic receptor. To this end, we developed a model that permitted measurement of real-time changes in neuronal NE in the RVLM or nucleus tractus solitarius (NTS) along with blood pressure and heart rate in the conscious SHR in response to localized microinjections of selective I1 (rilmenidine) or alpha2-adrenergic (alpha-methylnorepinephrine; alpha-MNE) agonist versus the mixed I1/alpha2 agonist clonidine. To further support the hypothesis, we investigated the effects of localized selective alpha2- (SK&F86466) or I1 (efaroxan) blockade on the reductions in neuronal NE and blood pressure elicited by intra-RVLM rilmenidine. In the latter experiment, changes in RVLM neuronal c-Fos (another marker of sympathetic neural activity) were also investigated. Intra-RVLM rilmenidine (40 nmol) or clonidine (1 nmol) similarly reduced RVLM NE and blood pressure; these responses were approximately 2-fold greater than those elicited by the pure alpha2-adrenergic agonist alpha-MNE (10 nmol). By contrast, intra-NTS rilmenidine or clonidine had no effect on NTS NE or blood pressure versus significant reductions in both parameters by alpha-MNE. Intra-RVLM rilmenidine decreased c-Fos expression, and these responses were abolished by efaroxan but not by SK&F 86466. These findings suggest: (1) in the RVLM, I1-receptor signaling suppresses cardiovascular neuron activity, which leads to lowering of blood pressure; (2) although the alpha2-adrenergic receptor in the RVLM serves a similar role, it does not exert a tonic neuronal inhibitory effect and is not essential, as a downstream signaling entity, for the I1-evoked neurobiological effects in the brainstem. The potential confounding effects of anesthetics on the I1 and/or alpha2 receptor-mediated neuronal and cardiovascular responses were circumvented in the present study.

Site-dependent inhibition of neuronal c-jun in the brainstem elicited by imidazoline I1 receptor activation: role in rilmenidine-evoked hypotension.

Clonidine (a mixed alpha2-adrenoceptor and imidazoline I1 receptor agonist)-evoked hypotension was associated with dissimilar reductions in c-jun gene expression in the rostral ventrolateral medulla (RVLM) and the nucleus tractus solitarius (NTS) in normotensive rats. In the present study, we investigated the relative contribution of the alpha2-adrenoceptor vs. the imidazoline I1 receptor to the reduction in c-jun gene expression in these two brainstem areas. In conscious spontaneously hypertensive rats (SHRs), equihypotensive doses of three centrally acting hypotensive drugs with different selectivity for the two receptors were administered intracisternally (4 microl) to limit their actions to the brain. As a control, a similar hypotensive response was elicited by i.v. hydralazine. Clonidine (0.5 microg), or alpha-methylnorepinephrine (alpha-MNE, 4 microg), a highly selective alpha2-adrenoceptor agonist, similarly reduced c-jun mRNA expression in the NTS and rostral ventrolateral medulla. In contrast, a similar hypotensive response (-37+/-3.5 mm Hg) caused by the selective imidazoline I1 receptor agonist rilmenidine (25 microg) was associated with reduction in c-jun mRNA expression in the rostral ventrolateral medulla, but not in the NTS. Further, intra-rostral ventrolateral medulla rilmenidine (40 nmol) reduced c-Jun protein expression in rostral ventrolateral medulla and blood pressure and both responses were antagonized by selective imidazoline I1 receptor (efaroxan, 4 nmol), but not alpha2-adrenoceptor (SK&F 86466, 10 nmol) blockade. These results suggest: (1) the c-jun containing neurons in the brainstem are involved in the centrally mediated hypotension elicited by centrally acting antihypertensive agents, and (2) the alpha2-adrenoceptor modulates c-jun gene expression in the NTS and rostral ventrolateral medulla implicated in centrally mediated hypotension, and (3) the imidazoline I1 receptor mediated inhibition of c-jun gene expression in the rostral ventrolateral medulla, but not in the NTS, contributes to the centrally mediated hypotension by the second generation drugs.

The Leu7Pro polymorphism of preproNPY is associated with decreased insulin secretion, delayed ghrelin suppression, and increased cardiovascular responsiveness to norepinephrine during oral glucose tolerance test.

CONTEXT: Neuropeptide Y (NPY) plays a role in angiogenesis, cardiovascular regulation, and hormone secretion. The leucine7 to proline7 (Leu7Pro) polymorphism of preproNPY is associated with vascular diseases and has an impact on hormone levels in healthy subjects. OBJECTIVE: The current study investigated the role of the Leu7Pro polymorphism in metabolic and cardiovascular autonomic regulation. DESIGN AND SUBJECTS: A 5-h oral glucose tolerance test was performed on 27 healthy volunteers representing two preproNPY genotypes (Leu7/Pro7 and Leu7/Leu7) matched for age, sex, body mass index and physical activity. MAIN OUTCOME MEASURES: Simultaneously we performed cardiovascular autonomic function tests and plasma measurements of sympathetic transmitters, glucose, insulin, and ghrelin. RESULTS: The subjects with Leu7/Pro7 genotype had decreased plasma NPY, norepinephrine (NE), and insulin concentrations and insulin to glucose ratios. The suppression of ghrelin concentrations after glucose ingestion was delayed in these subjects. They also had increased heart rate variability indices and baroreflex sensitivity. However, they displayed significant negative association of NE concentration with variability of low-frequency R-R-intervals and with baroreflex sensitivity. CONCLUSIONS: The Leu7Pro polymorphism of preproNPY is related to decreased level of basal sympathetic activity, decreased insulin secretion, and delayed ghrelin suppression during oral glucose tolerance test. The increased responsiveness of autonomic functions to NE associated with the polymorphism may be connected to increased cardiovascular vulnerability.

Left ventricular ejection time: a potential determinant of pulse wave velocity in young, healthy males.

OBJECTIVE: Pulse wave velocity (PWV) is a classic marker of vascular stiffness. Recent studies showed that heart rate is an important determinant of PWV. The purpose of this study was to evaluate the role of myocardial function in determining PWV under resting conditions and under adrenergic stimulation. DESIGN AND METHODS: Hemodynamic parameters were investigated under resting conditions in 102 young, healthy males and under stimulation of either beta- or alpha(2)-adrenoceptors in six young, healthy males. PWV was determined from pressure tracing over the carotid and femoral artery. Central hemodynamics were assessed by impedance cardiography and systolic time intervals. Simple (r) and multiple (beta) regression analyses were used to assess the relationships between PWV and hemodynamic parameters. RESULTS: Under resting conditions, PWV was correlated to age (beta = 0.259, P = 0.0052), diastolic blood pressure (beta = 0.279, P = 0.0072) and left ventricular ejection time (beta = -0.314, P = 0.0277). Under alpha(2)-adrenergic stimulation PWV was only correlated to diastolic blood pressure (DBP) (beta = 0.806, P = 0.0020). Under beta-adrenergic stimulation PWV was only correlated to left ventricular ejection time index (r = -0.52, P = 0.0325). CONCLUSIONS: Left ventricular ejection time may be an important determinant of pulse wave velocity under resting and adrenergic conditions in young, healthy males. Further studies are needed to evaluate this relationship in other populations including females and patients with cardiovascular disease.

A comparison of alpha-methylnorepinephrine, vasopressin and epinephrine for cardiac resuscitation.

The objective of this research was to compare the effects of an alpha- and beta-adrenergic agonist, epinephrine, a selective alpha(2)-adrenergic agonist, alpha-methylnorepinephrine (alpha-MNE), and a non-adrenergic vasopressin on post-resuscitation myocardial function and duration of survival. Epinephrine continues to be the primary adrenergic agent for advanced cardiac life support. However, its major inotropic actions and especially its beta-adrenergic and, to a lesser extent, its alpha(1)-actions increase the severity of global ischemia during cardiac arrest and adversely affect post-resuscitation myocardial function and survival. We had previously observed significantly better outcomes with a selective alpha(2)-adrenergic agonist when compared with epinephrine. Non-adrenergic vasopressin also has promise of more favorable actions. The present study was, therefore, undertaken to compare a selective alpha(2)-adrenergic vasopressor drug with vasopressin, epinephrine, and saline placebo. Ventricular fibrillation (VF) was induced in 20 Sprague-Dawley rats. Mechanical ventilation and precordial compression were initiated after 8 min of untreated VF. About 2 min later, alpha-MNE in a dose of 100 microgram/kg, vasopressin in a dose of 0.4 U/kg, epinephrine in a dose of 30 microgram/kg, or saline control was administered. Defibrillation was attempted after 6 min of CPR. Left ventricular pressure, dP/dt(40), -dP/dt, and cardiac index were measured for an interval of 240 min after resuscitation. Except for saline controls, comparable increases in coronary perfusion pressure (CPP) were observed after each drug intervention. All animals were successfully resuscitated. Post-resuscitation myocardial function and survival were significantly better in animals treated with alpha-MNE. Both post-resuscitation myocardial function and survival were most improved after administration of the selective alpha(2)-adrenergic agonist, intermediate after vasopressin and least after epinephrine and saline placebo.

Myocardial dysfunction and male mortality in peroxisome proliferator-activated receptor alpha knockout mice overexpressing lipoprotein lipase in muscle.

Free fatty acids (FFA) are liberated from triglyceride-rich lipoproteins by lipoprotein lipase (LPL) and are considered to be a principal energy source for the heart. The peroxisome proliferator-activated receptor alpha (PPARalpha) is a key regulator of FFA catabolism. To investigate its role in cardiac muscle metabolism, transgenic mice overexpressing LPL in skeletal and cardiac muscle were bred on a PPARalpha knockout background. Fifty-five percent of male animals lacking PPARalpha and overexpressing LPL died within 4 months after birth. In contrast, females of this genotype stayed alive. Deceased animals exhibited cardiopulmonary congestion but had no increase of neutral lipids in the heart. Changes in plasma glucose, FFA, lactate, and triglycerides did not clearly account for gender-specific differences in mortality; however, they indicated a critical role for PPARalpha during fasting. Analysis of cardiac function revealed that in isolated perfused hearts, left ventricular developed pressure (a measure of contractility) was markedly lower in PPARalpha knockout mice overexpressing LPL compared with controls. Glucose uptake of isolated perfused hearts was significantly higher in PPARalpha knockout mice with both normal or increased LPL expression. However, uptake of FFA was not different among genotypes. In contrast, fasted FFA levels were significantly lower in cardiac muscle of PPARalpha knockout mice with normal LPL expression (-26%) and PPARalpha knockout mice overexpressing LPL (-38%) compared with controls. Our results indicate a critical role for PPARalpha in myocardial pump function and suggest that mouse models combining different genetic effects such as PPARalpha knockout mice overexpressing muscle LPL may be useful to study cardiomyopathies.

Short exposure of maturing, bone marrow-derived dendritic cells to norepinephrine: impact on kinetics of cytokine production and Th development.

The information gathered by dendritic cells (DC) during the innate immune response to a pathogen is determinant for the type of adaptive response. Here we show that short-term (3 h) exposure of bone marrow-derived DC to norepinephrine (NE), at the beginning of lipopolysaccharide (LPS) or keyhole limpet hemocyanin (KLH) stimulation hampers IL-12 production and increases IL-10 release. The NE effect was mediated by both beta- and alpha2-adrenergic receptors. The capacity of NE-exposed DC to produce IL-12 upon CD40 cross-linking as well as to stimulate allogeneic T-helper (Th) lymphocytes was reduced. Adoptive transfer of NE-exposed DC induced a Th2 slanted response in vivo. Thus, a brief NE exposure of antigen-stimulated DC seems to limit their Th1 polarizing properties. Noteworthy, the ganglionic blocker pentolinium administered in mice before skin sensitization with fluoroscein isothiocyanate (FITC) could increase the Th1-type response in the draining lymph nodes. Our results suggest that the extent of Th differentiation in the response to an antigen might be influenced by the local sympathetic nervous activity in the early phase of dendritic cell stimulation.

A selective alpha(2)-adrenergic agonist for cardiac resuscitation.

The effects of selective alpha(2)-adrenergic agonist alpha-methylnorepinephrine on the initial success of resuscitation and postresuscitation myocardial function were compared with nonselective alpha- and beta-adrenergic epinephrine in a swine model of cardiac arrest. Epinephrine, the primary pharmacological intervention in the treatment of cardiac arrest, improves immediate outcome. However, epinephrine increases the severity of myocardial dysfunction after cardiac resuscitation. Both inotropic and chronotropic actions provoke disproportionate increases in myocardial oxygen consumption by the ischemic heart, prompting this study, in which we hypothesized that a selective alpha(2)-adrenergic agonist, alpha-methylnorepinephrine (alpha-MNE), would moderate these adverse effects of epinephrine and minimize postresuscitation myocardial dysfunction. After 7 minutes of untreated ventricular fibrillation (VF) in 14 anesthetized male domestic pigs, precordial compression at a fixed rate of 80 compressions/min was begun, along with mechanical ventilation. Either alpha-MNE (100 microg/kg) or epinephrine (20 microg/kg) was administered as a bolus after 2 minutes of precordial compression. After an additional 4 minutes of precordial compression, defibrillation was attempted. Left ventricular systolic and diastolic function was quantitated with the use of transesophageal echo-Doppler imaging. Comparable increases in coronary perfusion pressure to 15 mm Hg were observed after the administration of both drugs. All animals were successfully resuscitated; epinephrine and alpha-MNE were equally quick in restoring spontaneous circulation after 7 minutes of untreated VF. Ejection fraction was reduced by 35% and 14% by epinephrine and alpha-MNE, respectively, after resuscitation. Epinephrine and alpha-MNE increased postresuscitation heart rate by 38% and 15%, respectively. Accordingly, significantly less postresuscitation impairment followed the administration of alpha-MNE. alpha-MNE, a selective alpha-adrenergic agonist, was as effective as epinephrine in restoring spontaneous circulation after 7 minutes of untreated VF in a porcine model for CPR and demonstrated lesser postresuscitation myocardial injury.

Evidence for synergy between alpha(2)-adrenergic and nonadrenergic mechanisms in central blood pressure regulation.

BACKGROUND: Both alpha(2)-adrenergic and non--alpha(2)-adrenergic mechanisms seem to be involved in the hypotensive effect of imidazoline-like drugs. This study aimed at investigating how these 2 mechanisms work together to modify blood pressure (BP). METHODS AND RESULTS: LNP 509, which appeared in this study to be devoid of alpha(2A)-adrenergic activity, was administered to anesthetized rabbits and wild-type (WT) mice into the cisterna magna and into the fourth ventricle, respectively. Mean arterial pressure decreased by a maximum of 46 +/- 4% and 16 +/- 2%, respectively. In D79N mice, which lack functional alpha(2A)-adrenergic receptors, LNP 509 also reduced mean arterial pressure by 17 +/- 2%. The hypotension induced by LNP 509 (100 microg/kg intracisternally) was prevented by S23757 (1 mg/kg intracisternally), an antagonist highly selective for I(1)-imidazoline binding sites (I(1)BS). A synergy between LNP 509 and the alpha(2)-adrenergic agonist alpha-methylnoradrenaline (alpha-MNA) was observed in rabbits (cisterna magna injection) and in WT mice (fourth ventricle injection) but not, as expected, in D79N mice. Similar to LNP 509 alone, rilmenidine (fourth ventricle injection), which binds both to alpha(2)-adrenergic receptors and to I(1)BS, decreased BP in D79N mice. In WT animals, rilmenidine had a significantly greater effect. Microinjections performed in rabbits showed that the synergism occurred at least in part in the nucleus reticularis lateralis of the brain stem. CONCLUSIONS: These results demonstrate that a central imidazoline-sensitive, but non--alpha(2)-adrenergic, mechanism can modify BP by itself. This mechanism, which may involve I(1)BS, interacts synergistically with an alpha(2)-adrenergic mechanism to decrease BP.

Nitric oxide discriminates the sites and mechanisms of action of centrally acting anti-hypertensive drugs in rabbits.

The aim of the present study was to further investigate the mechanisms of the central hypotensive action of catecholamines and imidazolines, in particular the role of nitric oxide (NO). Microinjections into the nucleus reticularis lateralis (NRL/RVLM) located in the rostroventrolateral part of the medulla (RVLM) and/or into the nucleus tractus solitarii (NTS) were performed in pentobarbital-anesthetized rabbits. Microinjections of brimonidine (1 ng/kg), which binds both alpha(2)-adrenergic receptors (alpha(2)-ARs) and I(1) imidazoline receptors (I(1)Rs), into the NRL/RVLM induced hypotension (69+/-2 vs. 88+/-2 mm Hg) (p<0.05). Microinjections of S23757 (1 microg/kg), an antagonist highly selective for I(1)Rs, into the same site, prevented the hypotensive effect of brimonidine. These data show that the hypotensive effects of low doses of brimonidine involve the I(1)Rs of the NRL/RVLM. Alpha-methylnoradrenaline (alpha-MNA) (0.5 microg/kg) microinjected into the NTS induced hypotension (76+/-4 vs. 91+/-4 mm Hg) (p<0.05). Microinjections of a low dose of brimonidine (1 ng/kg) into the NTS had no blood pressure (BP) effect at all. In contrast, a higher dose (10 ng/kg) acting on alpha(2)-ARs induced hypotension (72+/-3 vs. 96+/-2 mm Hg) (p<0.05). Nomega-Nitro-L-arginine (L-NNA) (1.5 microg/kg) injected into the NRL/RVLM prevented the hypotensive effect of both alpha-MNA and the higher dose of brimonidine injected into the NTS. Bicuculline (1.5 microg/kg) injected into the NRL/RVLM prevented the hypotensive effect of alpha-MNA injected into the NTS. It is demonstrated that (i) the activation of alpha(2)-ARs of NTS triggers a neuronal GABAergic pathway projecting to the NRL/RVLM region which is NO dependent (ii) both alpha(2)-adrenergic (NTS) and non-adrenergic I(1)R (NRL/RVLM) mechanisms account for the very powerful hypotensive effect of brimonidine, a compound with high affinities at both types of receptors.

Effects of central imidazolinergic and alpha2-adrenergic activation on water intake

Non-adrenergic ligands that bind to imidazoline receptors (I-R), a selective ligand that binds to alpha2-adrenoceptors (alpha2-AR) and mixed ligands that bind to both receptors were tested for their action on water intake behavior of 24-h water-deprived rats. All drugs were injected into the third cerebral ventricle. Except for agmatine (80 nmol), mixed ligands binding to I-R/alpha2-AR such as guanabenz (40 nmol) and UK 14304 (20 nmol) inhibited water intake by 65 percent and up to 95 percent, respectively. The selective non-imidazoline alpha2-AR agonist, alpha-methylnoradrenaline, produced inhibition of water intake similar to that obtained with guanabenz, but at higher doses (80 nmol). The non-adrenergic I-R ligands histamine (160 nmol, mixed histaminergic and imidazoline ligand) and imidazole-4-acetic acid (80 nmol, imidazoline ligand) did not alter water intake. The results show that selective, non-imidazoline alpha2-AR activation suppresses water intake, and suggest that the action on imidazoline sites by non-adrenergic ligands is not sufficient to inhibit water intake

Effects of central imidazolinergic and alpha2-adrenergic activation on water intake.

Non-adrenergic ligands that bind to imidazoline receptors (I-R), a selective ligand that binds to alpha2-adrenoceptors (alpha2-AR) and mixed ligands that bind to both receptors were tested for their action on water intake behavior of 24-h water-deprived rats. All drugs were injected into the third cerebral ventricle. Except for agmatine (80 nmol), mixed ligands binding to I-R/alpha2-AR such as guanabenz (40 nmol) and UK 14304 (20 nmol) inhibited water intake by 65% and up to 95%, respectively. The selective non-imidazoline alpha2-AR agonist, alpha-methylnoradrenaline, produced inhibition of water intake similar to that obtained with guanabenz, but at higher doses (80 nmol). The non-adrenergic I-R ligands histamine (160 nmol, mixed histaminergic and imidazoline ligand) and imidazole-4-acetic acid (80 nmol, imidazoline ligand) did not alter water intake. The results show that selective, non-imidazoline alpha2-AR activation suppresses water intake, and suggest that the action on imidazoline sites by non-adrenergic ligands is not sufficient to inhibit water intake.

Haemodynamic characterization of young normotensive men carrying the 825T-allele of the G-protein beta3 subunit.

A C825T polymorphism was recently identified in the gene for the G-protein beta3 subunit, the T-allele being associated with hypertension. To better understand the underlying pathophysiological mechanisms, we compared the haemodynamics of young healthy males with and without the T-allele. In three studies, subjects were investigated with regard to cardiac and vascular function at rest and following intravenous administration of the beta-adrenoceptor antagonist, propranolol, and the alpha2-adrenoceptor agonist, alpha-methylnoradrenaline, and with regard to local venous vasoconstriction in the dorsal hand vein in situ following infusion of the alpha2-adrenoceptor agonist, azepexol. alpha2-Adrenoceptor agonists were chosen as vasoconstrictor drugs since alpha2-adrenoceptors couple to pertussis toxin (PTX)-sensitive G-proteins and since in-vitro studies have demonstrated enhanced signal transduction of PTX-dependent pathways in the presence of the T-allele. Total peripheral resistance was determined as a parameter of vasoconstrictor tone and heart rate, stroke volume and systolic time intervals for cardiac function. T-allele carriers had a significantly elevated stroke volume and lower total peripheral resistance at baseline. After propranolol, their fall in stroke volume was significantly greater. During alpha-methylnoradrenaline infusion, elevation of total peripheral resistance was not increased relative to controls. Similarly, the constriction response of the dorsal hand vein to azepexol was not different. Our study does not support the idea of increased vasoconstrictor tone in T-allele carriers either at rest or during stimulation of alpha2-adrenoceptors. However, this allele may be associated with elevated cardiac stroke volume.