A polymorphism in the norepinephrine transporter gene is associated with affective and cardiovascular disease through a microRNA mechanism.

Norepinephrine released from sympathetic nerves is removed from the neuroeffector junction via the action of the norepinephrine transporter (NET). NET impairment is evident in several clinically important conditions including major depressive disorder (MDD), panic disorder (PD), essential hypertension and the postural orthostatic tachycardia syndrome (POTS). We aimed to determine whether a single nucleotide polymorphism (SNP) in the 3' untranslated region (UTR) of the NET gene is associated with NET impairment and to elucidate the mechanisms involved. The analyses were carried out in two cohorts of European ancestry, which included healthy controls and MDD, PD, hypertensive and POTS patients. Compared with controls, cases had significantly higher prevalence of the T allele of rs7194256 (C/T), arterial norepinephrine, depression and anxiety scores, larger left ventricular mass index, higher systolic and diastolic blood pressures, and heart rate. Bioinformatic analysis identified that the microRNA miR-19a-3p could bind preferentially to the sequence created by the presence of the T allele. This was supported by results of luciferase assays. Compared with controls, cases had significantly lower circulating miR-19a-3p, which was associated with pathways related to blood pressure and regulation of neurotransmission. In vitro norepinephrine downregulated miR-19a-3p. In conclusion, the T allele of the rs7194256 SNP in the 3'UTR of the NET gene is more prevalent in diseases where NET impairment is evident. This might be explained by the creation of a binding site for the microRNA miR-19a-3p. A defect in NET function may potentiate the sympathetic neurochemical signal, predisposing individuals with affective diseases to increased risk of cardiovascular disease development.

Imidazoline receptors associated with noradrenergic terminals in the rostral ventrolateral medulla mediate the hypotensive responses of moxonidine but not clonidine.

We determined whether the cardiovascular actions of central anti-hypertensive agents clonidine and moxonidine are dependent on noradrenergic or serotonergic innervation of the rostral ventrolateral medulla (RVLM) in conscious rabbits. 6-Hydroxydopamine (6-OHDA) or 5,6-dihydroxytriptamine (5,6-DHT) was injected into the RVLM to deplete noradrenergic and serotonergic terminals respectively. One, 2 and 4 weeks later, responses to fourth ventricular (4V) clonidine (0.65 microg/kg) and moxonidine (0.44 microg/kg) were examined. Destruction of noradrenergic pathways in the RVLM by 6-OHDA reduced the hypotensive response to 4V moxonidine to 62%, 47% and 60% of that observed in vehicle treated rabbits at weeks 1, 2 and 4 respectively. The moxonidine induced bradycardia was similarly attenuated (to 46% of vehicle). Conversely, 6-OHDA had no effect on the hypotensive or bradycardic effects of 4V clonidine. Efaroxan (I(1)-imidazoline receptor/alpha(2)-adrenoceptor antagonist; 3.5, 11, 35 microg/kg) and 2-methoxyidazoxan (alpha(2)-adrenoceptor antagonist; 0.3, 0.9, 3 microg/kg) equally reversed the hypotension to 4V clonidine, suggesting a mainly alpha(2)-adrenoceptor mechanism. Efaroxan preferentially reversed responses to moxonidine in both vehicle and 5,6-DHT groups and in the 1st week after 6-OHDA, suggesting a mechanism involving mainly I(1)-imidazoline receptors. This selectivity was subsequently lost in the 2nd and 4th weeks when the remaining hypotension was mainly mediated by alpha(2)-adrenoceptors. Depletion of serotonergic terminals did not alter the responses to either agonist nor did it change the relative effectiveness of the antagonists. Western blots of RVLM tissues probed with imidazoline and alpha(2)-adrenoceptor antisera showed a pattern of bands close to that reported in other species. The main effect of 6-OHDA was an 18% lower level of the 42 kDa imidazoline protein (P<0.05). We conclude that the hypotensive and bradycardic actions of moxonidine but not clonidine are mediated through imidazoline receptors and are dependent on intact noradrenergic pathways within the RVLM. Furthermore, the noradrenergic innervation may be associated with a 42 kDa imidazoline receptor protein.

Baroreflex control of heart rate and cardiac hypertrophy in angiotensin II-induced hypertension in rabbits.

The cardiac hypertrophy observed in hypertension is thought to be responsible for the accompanying deficiency in the baroreflex control of heart rate. In this study, we assessed the baroreflex relationship between heart rate and arterial pressure on a group of seven rabbits during a normotensive period, during the early phase of angiotensin II (Ang II)-induced hypertension II week) (50 ng/kg per minute i.v. via osmotic minipumps), after 7 weeks of continuous hypertension, then 2 days after Ang II was stopped, and finally 7 days after Ang II. Left ventricles were weighed for measurement of left ventricular weight-body weight ratio. One week of intravenous Ang II infusion produced hypertension (mean arterial pressure from 80 +/- 2 up to 115 +/- 8 mm Hg), with significantly increased heart rate and hematocrit. The heart rate-arterial pressure baroreflex curve was shifted to the right, with a significant 45% reduction in the gain of the reflex (-6.4 +/- 1.5 to -3.5 +/- 0.2 beats per minute/mm Hg). After 7 weeks of Ang II, arterial pressure was still elevated (112 +/- 4 mm Hg) and the gain of the baroreflex curve still somewhat attenuated, although it was no longer markedly different from normotensive levels (gain, -5.09 +/- 0.95, 20% reduction from normotensive level). Two days after the Ang II infusion was stopped, arterial pressure had returned to normotensive levels, although hematocrit and heart rate remained elevated. At this time, the baroreflex curve was similar to prehypertensive control levels, with no further changes when measured again 7 days after Ang II. Cardiac hypertrophy was present when measured at 7 days after angiotensin (left ventricular weight-body weight ratio: 1.78 +/- 0.05 versus 1.35 +/- 0.04 g/kg, hypertensive versus normotensive, P < .05). Thus, although Ang II infusion produced an initial deficit in the baroreflex control of heart rate, this effect became less as the hypertension continued. Furthermore, although cardiac hypertrophy developed, its presence did not appear to be sufficient to produce a decrease in barosensitivity independent of raised arterial pressure.

ANP and bradycardic reflexes in hypertensive rats: influence of cardiac hypertrophy.

In previous studies we demonstrated that in normotensive rats, but not in spontaneously hypertensive rats (SHR), atrial natriuretic peptide (ANP) enhances bradycardic reflexes through an action on cardiac vagal afferent pathways. The present study aimed to determine whether cardiac hypertrophy, hypertension, or a nonreversible genetic factor accounted for the insensitivity of SHR to ANP action on cardiac reflex pathways. SHR were treated with the angiotensin-converting enzyme (ACE) inhibitor perindopril (3 mg/kg per day) for 6 weeks from 4 to 9 weeks of age (SHR-S, n=10) or for 9 weeks from 4 to 12 weeks of age (SHR-L, n=10) or were untreated (SHR, n=10) to produce differential effects on blood pressure and left ventricle/body weight ratio (LV/BW). Untreated normotensive Wistar-Kyoto rats (WKY, n=10) were also studied. At 13 weeks of age, all rats were instrumented with aortic and jugular catheters, and at 14 weeks we measured heart rate reflexes to rapid intravenous infusions of methoxamine (100 microg/kg, cardiac baroreflex) and serotonin (5 to 60 microg/kg, von Bezold-Jarisch cardiac chemosensitive reflex), with either alpha-rat ANP (150 ng/kg per minute IV) or saline vehicle (270 microL/h IV) infusion. Perindopril treatment for 6-week (SHR-S) and 9-week (SHR-L) durations maintained blood pressure at normotensive levels in both groups. SHR-S exhibited a small degree of cardiac hypertrophy (LV/BW was 8% higher than in WKY but 11% less than in untreated SHR), but LV/BW was normalized in SHR-L (to within 1% of WKY LV/BW). In WKY, ANP significantly (P<0.05) enhanced bradycardic responses to both the cardiac baroreflex (by 42+/-10%) and von Bezold-Jarisch chemosensitive reflex (by 17+/-5%) activation but had no effect in SHR. The cardiac reflex action of ANP was restored in SHR-L (ANP enhanced reflex bradycardia by 28+/-12% and 36+/-8%, baroreflex and von Bezold-Jarisch reflex, respectively; P<0.05), but SHR-S, which developed some cardiac hypertrophy, remained unresponsive to ANP. Our results suggest that the inability of ANP to sensitize cardiac vagal (nonarterial) afferents in SHR was not due to an inherited irreversible component, or the hypertension per se, but was associated with the presence of cardiac hypertrophy. A functional consequence of hypertension-induced cardiac hypertrophy may be the inhibition of the cardioprotective action of ANP through cardiac vagal reflexes.

ANP enhances bradycardic reflexes in normotensive but not spontaneously hypertensive rats.

Baroreflex control of heart rate in spontaneously hypertensive rats (SHR) is defective, largely because of a poor vagal contribution to the reflex. We have demonstrated previously that atrial natriuretic peptide (ANP) enhances reflex bradycardia in normotensive rats through an action on nonarterial vagal afferent pathways. In the present study, we investigated whether ANP could reverse the baroreflex abnormality in SHR. Heart rate reflexes were activated by three different methods in conscious, instrumented SHR and Wistar-Kyoto rats (WKY) in the presence of intravenous infusions of vehicle (saline) or rat ANP (150 ng/kg per minute). Heart rate responses were measured by (1) the steady-state changes in blood pressure after alternating slow infusions (over approximately 15 to 30 seconds) of a pressor (methoxamine) and depressor (nitroprusside) drug (stimulating predominantly arterial baroreceptors), (2) the ramp method of rapid infusion of methoxamine (over < 10 seconds; stimulating arterial and cardiopulmonary baroreceptors), and (3) the von Bezold-Jarisch method of activating chemically sensitive cardiac receptors through serotonin injections. ANP enhanced the heart rate range of the arterial baroreflex (steady-state method) by 13 +/- 3% in WKY but had no significant effect on the sensitivity or any other parameter of the steady-state baroreflex. When a very rapid rise in blood pressure was elicited by the ramp method in WKY, ANP significantly enhanced baroreflex bradycardia (sensitivity increased by 29 +/- 9%, P < .05). ANP also enhanced the bradycardia of the von Bezold-Jarisch reflex (by 33 +/- 16%, P < .05) in WKY. By contrast, ANP did not influence baroreceptor or chemoreceptor heart rate reflex responses in SHR. We conclude that in normotensive rats, ANP facilitates cardiopulmonary bradycardic reflexes. The lack of effect of ANP in SHR may be related to an underlying structural or genetic alteration in their cardiac sensors, perhaps associated with cardiac hypertrophy, that prevents the ANP-induced activation of cardiac sensory afferents, resulting in cardioinhibition.

Central and peripheral autonomic mechanisms involved in the circulatory actions of methyldopa.

Intracisternal (i.c.) and intravenous (i.v.) administration of methyldopa in conscious rabbits produced closely similar changes in hemodynamics, heart rate, and falls in plasma norepinephrine levels. Two weeks after giving i.c. 6-hydroxydopamine (6-OHDA), when there is widespread destruction of central noradrenergic neurons, the effects of i.c. methyldopa virtually were abolished. This suggests that noradrenergic neurons are the major central site of biotransformation into active metabolites. The circulatory and norepinephrine effects of i.v. methyldopa were attenuated but not completely abolished after giving i.c. 6-OHDA. Hence, in the rabbit about 70% of the action of methyldopa was central and about 30% was peripheral in the human therapeutic range of methyldopa concentrations. Preliminary lesion experiments suggest that the A5 nucleus plays an important role in the bradycardia. Two weeks after giving 5,6-dihydroxytryptamine (5,6-DHT) to destroy serotonergic (5HT) neurons the effects of i.c. methyldopa on mean arterial pressure (MAP) and heart rate were attenuated to approximately 50% of control effects. Therefore, some of the central effects of methyldopa apparently are mediated through 5HT pathways. We also compared the effects of i.c. methyldopa with those of i.c. clonidine (an alpha 2-adrenergic receptor agonist) and with the effects of transmitter release from the endings of noradrenergic and 5HT neurons during the first few hours after either 6-OHDA or 5,6-DHT administration. Our findings suggest that after biotransformation of methyldopa its active metabolites increase the activity of the bulbospinal noradrenergic neurons that control MAP and heart rate and reduce the activity of bulbospinal 5HT neurons.

Medullary neurons activated by angiotensin II in the conscious rabbit.

Previous studies have shown that angiotensin II (Ang II) can activate cardiovascular neurons within the medulla oblongata via an action on specific receptors. The purpose of this study was to determine the distribution of neurons within the medulla activated by infusion of Ang II into the fourth ventricle of conscious rabbits, using the expression of Fos, the protein product of the immediate early gene c-fos as a marker of neuronal activation. Experiments were done in both intact and barodenervated animals. In comparison with a control group infused with Ringer's solution alone, in both intact and barodenervated animals, fourth ventricular infusion of Ang II (4 to 8 pmol/min) induced a significant increase in the number of Fos-positive neurons in the nucleus of the solitary tract and in the rostral, intermediate, and caudal parts of the ventrolateral medulla. Double-labeling for Fos and tyrosine hydroxylase immunoreactivity showed that 50% to 75% of Fos-positive cells in the rostral, intermediate, and caudal ventrolateral medulla and 30% to 40% of Fos-positive cells in the nucleus of the solitary tract were also positive for tyrosine hydroxylase in both intact and barodenervated animals. The distribution of Fos-positive neurons corresponded very closely to the location of Ang II receptor binding sites as previously determined in the rabbit. The results indicate that medullary neurons activated by Ang II are located in discrete regions within the nucleus of the solitary tract and ventrolateral medulla and include, in all of these regions, both catecholamine and noncatecholamine neurons.

Importance of cardiac, but not vascular, hypertrophy in the cardiac baroreflex deficit in spontaneously hypertensive and stroke-prone rats.

In the present study, we examined whether antihypertensive treatment of young and adult hypertensive rats with the angiotensin-converting enzyme (ACE) inhibitor perindopril could restore the baroreflex vagal deficit and whether this was related to prevention of cardiac or vascular hypertrophy. Spontaneously hypertensive (SHR), stroke-prone spontaneously hypertensive (SHR-SP), and Wistar-Kyoto (WKY) rats were untreated or treated with perindopril (3 mg/kg/day) in the drinking water from 4-9 and from 14-20 weeks of age. Steady-state sigmoidal mean arterial pressure (MAP)-heart rate (HR) reflex curves were obtained in the conscious rats by the injection of pressor and depressor agents before and after atenolol (vagal component). Increased left ventricle to bodyweight ratio (LV/BW) indicated cardiac hypertrophy. After ganglion blockade, the minimum MAP produced by nitroprusside and the maximum produced by methoxamine were used as indications of vascular hypertrophy. Perindopril treatment reduced cardiac and vascular hypertrophy to different extents in SHR and SHR-SP. The 4-9 and 14-20 week treatments reduced MAP and both minimum and maximum blood pressure of the SHR to the levels of the untreated WKY. However, only in the older animals was LV/BW restored. In the SHR-SP, early treatment had a much greater effect on vascular hypertrophy than on LV/BW. The reverse occurred for the 14-20 week animals. In untreated hypertensive animals the baroreflex curves were shifted to the right with reduced vagal HR range. Perindopril treatment shifted the baroreflex curves back towards the WKY curves. Vagal HR range was strongly correlated with the LV/BW, whereas vagal HR range was less well related to the level of vascular hypertrophy or blood pressure. These results suggest that antihypertensive treatment can restore cardiac baroreflex function and that it is related to the reduction in cardiac hypertrophy. Although the mechanism of this relationship remains to be elucidated, these findings suggest that cardiac vagal afferents may be important.

Comparison of renal sympathetic baroreflex effects of rilmenidine and alpha-methylnoradrenaline in the ventrolateral medulla of the rabbit.

OBJECTIVE: To determine the influence of imidazoline receptors and alpha2-adrenoceptors in the rostral ventrolateral medulla (RVLM) on the renal sympathetic baroreflex. METHODS: The effects of rilmenidine (4 nmol) and alpha-methylnoradrenaline (alpha-MNA, 80 nmol) micro-injected into the RVLM of urethane-anaesthetized rabbits previously implanted with renal nerve recording electrodes were examined before and after micro-injection of the imidazoline receptor/alpha2-adrenoceptor antagonist idazoxan and the alpha2-adrenoceptor antagonist 2-methoxyidazoxan (2-MI). RESULTS: Rilmenidine and alpha-MNA both lowered mean arterial pressure (MAP) by 28% and renal sympathetic nerve activity (RSNA) by 35%, and reduced RSNA upper plateaus and ranges by 30-70%. Rilmenidine decreased both sympathetic burst frequency and amplitude while alpha-MNA reduced amplitude only. Rilmenidine shifted the RSNA baroreflex curve to the left while alpha-MNA shifted the curve to the right Idazoxan (13 nmol) reversed the hypotension and all RSNA effects of rilmenidine, while 2-MI (4 nmol) increased MAP 18% above the control and also reversed all RSNA parameters. By contrast, 2-MI reversed the alpha-MNA-induced hypotension and partially restored RSNA and the upper plateau of the RSNA baroreflex curve. Idazoxan treatment only partially reversed the hypotension after alpha-MNA and had no effect on any of the baroreflex curves. CONCLUSION: Both alpha-MNA and rilmenidine injected into the RVLM of rabbits produce renal sympathetic inhibition, but differences in the location of the baroreflex curve and the pattern of effects on burst amplitude and frequency suggest different mechanisms of action. The effects of idazoxan suggest that rilmenidine acts via imidazoline receptors. Since 2-MI reversed the actions of alpha-MNA and also rilmenidine, this suggests that alpha2-adrenoceptor hypotension can be produced in the rabbit RVLM and that rilmenidine may activate alpha2-adrenoceptors, possibly as a result of activating imidazoline receptors.

Relationship between cardiovascular hypertrophy and cardiac baroreflex function in spontaneously hypertensive and stroke-prone rats.

OBJECTIVE: To determine whether the reduced baroreceptor-heart rate reflex sensitivity in genetically hypertensive rats is related to the level of cardiac or vascular hypertrophy. DESIGN: Young spontaneously hypertensive rats (SHR), stroke-prone hypertensive rats (SHRSP) and Wistar-Kyoto (WKY) rats were treated chronically with the angiotensin converting enzyme (ACE) inhibitor perindopril in different regimens in order to produce a wide-ranging combination of cardiac and vascular hypertrophy. METHODS: All strains were treated with perindopril (0.1, 0.3, 1 or 3 mg/kg per day) in their drinking water from 4 until 9 weeks of age. Additional groups of SHR were treated with perindopril (3 mg/kg per day) from 4 until 12 weeks and from 4 until 14 weeks of age. At 13 weeks of age all animals were chronically instrumented with arterial and venous catheters. One week later, steady-state sigmoidal mean arterial pressure-heart rate reflex curves were obtained in the conscious rats by the injection of pressor and depressor agents before and after the administration of atenolol (1 mg/kg, intravenously) to determine the vagal component. The minimum and the maximum blood pressure produced by nitroprusside and methoxamine, respectively, after simultaneous ganglion and beta-adrenoceptor blockade were used as an index of whole body vascular hypertrophy. The left ventricular to body weight ratio was measured at the end of the experiment. RESULTS: At 14 weeks of age, mean arterial pressure, the maximum and minimum autonomically blocked blood pressure and the left ventricular to body weight ratio were 34, 20, 9 and 17% higher, respectively, in SHR, and 56, 35, 27 and 39% higher, respectively, in SHRSP than in WKY rats. Perindopril treatment dose-dependently reduced both cardiac and vascular hypertrophy but to different extents. The highest doses reduced mean arterial pressure and the autonomically blocked maximum and minimum blood pressure in both hypertensive strains to the levels of untreated WKY rats but approximately 50% of the cardiac hypertrophy was still present. The left ventricular to body weight ratio was normalized in SHR only with the longer term perindopril treatments. A comparison of the baroreflex function curves in untreated SHR and SHRSP showed that the vagal component of the heart rate range was markedly reduced compared to that in WKY rats. Treated SHRSP had a normal mean arterial pressure and a normal autonomically blocked maximum and minimum blood pressure, but their vagal heart rate range was only 63% of that in WKY rats. The heart rate range in SHR treated from 4 to 9 weeks of age was only marginally greater than that of untreated SHR, despite prevention of hypertension and vascular hypertrophy. In SHR treated from 4 until 12 weeks of age, which prevented cardiac hypertrophy, the vagal heart rate range was markedly greater. With perindopril from the age of 4 to 14 weeks, the vagal baroreflex heart rate range was similar to that of WKY rats. Thus the improvement in the vagal heart rate range was more closely related to the prevention of cardiac hypertrophy (r = 0.73, P < 0.001) than vascular hypertrophy or a blood pressure elevation. Since there were no further changes in the indices of hypertrophy, the presence of the ACE inhibitor was likely to have been responsible for restoring the remaining 35% of the vagal baroreflex not affected by structural factors. CONCLUSIONS: These results suggest that prevention of cardiac rather than vascular hypertrophy is the major requirement for normalizing the vagal component of the baroreceptor-heart rate reflex in hypertensive rats. However, structural changes accounted for only about two-thirds of the vagal deficit. The remainder was restored by the presence of an ACE inhibitor, suggesting that the vagal component of the baroreceptor-heart rate reflex is normally suppressed by the renin-angiotensin system.

Relative importance of central imidazoline receptors for the antihypertensive effects of moxonidine and rilmenidine.

OBJECTIVE: To determine the involvement of central imidazoline receptors in the cardiovascular actions of the antihypertensive agents moxonidine, rilmenidine and clonidine administered systemically. DESIGN AND METHODS: We determined the relative potency of these drugs with respect to their effects on mean arterial pressure and heart rate by performing cumulative intravenous dose-response relationship studies in six conscious rabbits. In another eight rabbits with implanted fourth-ventricular catheters, we investigated the central effects of three cumulative doses of an l1-imidazoline/ alpha 2-adrenoceptor antagonist, efaroxan, and of an alpha 2-adrenoceptor antagonist, 2-methoxyidazoxan (2-Ml), on the hypotension and bradycardia elicited by a single intravenous dose of the above agents. The doses of antagonists were matched for an equal reversal of the hypotension induced by fourth-ventricular alpha-methyldopa (an alpha 2-adrenoceptor agonist) and hence for similar alpha 2-adrenoceptor blockade. RESULTS: Moxonidine and rilmenidine were sevenfold and eightfold less potent, respectively, than was clonidine in eliciting hypotension. By comparison, moxonidine and clonidine were more potent than was rilmenidine in producing bradycardia. Efaroxan and 2-Ml reversed the hypotension and bradycardia induced by a single dose of all three agents dose-dependently. However, efaroxan was more effective than was 2-Ml at reversing the effects of rilmenidine and moxonidine. Complete reversal of their hypotensive effect was observed with the highest dose of efaroxan but the highest dose of 2-Ml reversed approximately 50% of that effect. In contrast, the two antagonists were equally effective at reversing the responses to clonidine. CONCLUSIONS: These results suggest that the hypotension and bradycardia induced by intravenous administration of moxonidine and rilmenidine were mediated mainly by actions on central imidazoline receptors whereas clonidine appears to act predominantly on central alpha 2-adrenoceptors.

Relative importance of medullary brain nuclei for the sympatho-inhibitory actions of rilmenidine in the anaesthetized rabbit.

OBJECTIVE: To determine the contribution of the rostral ventrolateral medulla and the nucleus of the solitary tract in mediating the attenuation of the renal sympathetic baroreflex produced by administration of rilmenidine to anaesthetized rabbits and to examine the relative contribution of alpha2-adrenoceptors and imidazoline receptors at these sites to the cardiovascular effects of rilmenidine. METHODS AND RESULTS: Rilmenidine micro-injected into the rostral ventrolateral medulla produced hypotension and inhibition of renal sympathetic nerve activity with doses an order of magnitude lower than those required in the nucleus tractus solitarius. Alpha-methylnoradrenaline, however, was similarly potent at producing hypotension when it was injected into the rostral ventrolateral medulla or nucleus tractus solitarius but, unlike rilmenidine, did not lower renal sympathetic nerve activity when it was injected into the nucleus tractus solitarius. The alpha2-adrenoceptor antagonist 2-methoxyidazoxan partially reversed the hypotension and renal sympathetic nerve activity inhibition due to alpha-methylnoradrenaline when it was administered into the rostral ventrolateral medulla, whereas the mixed alpha2-adrenoceptor/imidazoline receptor antagonists, idazoxan and efaroxan, did not. 2-Methoxyidazoxan, but not idazoxan, also reversed the hypotension when alpha-methylnoradrenaline was administered into the nucleus tractus solitarius. The hypotension induced by rilmenidine in the rostral ventrolateral medulla was completely reversed both by 2-methoxyidazoxan and by idazoxan, as was the sympathetic inhibition. To assess any interaction between the nucleus tractus solitarius and the rostral ventrolateral medulla in mediating the baroreflex effects of rilmenidine, we injected rilmenidine into the rostral ventrolateral medulla, the nucleus tractus solitarius or both nuclei and determined renal baroreflex responses of sympathetic nerve activity using drug-induced changes in blood pressure. Injection of 0.5 nmol rilmenidine into the rostral ventrolateral medulla reduced mean arterial pressure and basal renal sympathetic nerve activity as well as renal sympathetic baroreflex range (by 27%) and gain (by 35%). In contrast, injection of rilmenidine into the nucleus tractus solitarius had no effect on basal renal sympathetic nerve activity and renal sympathetic baroreflex parameters. The effect of combined injection was similar to that of administration into the rostral ventrolateral medulla alone. CONCLUSION: Our results show that the rostral ventrolateral medulla, rather than the nucleus tractus solitarius, is the major site involved in the hypotension and inhibition of the renal sympathetic baroreflex by rilmenidine. Comparison of the actions of alpha2-adrenoceptor and imidazoline receptor antagonists on the effects of rilmenidine and alpha-methylnoradrenaline suggests that these agents are acting at different receptors, presumably imidazoline and alpha2-adrenoceptors receptors, respectively, and that both are important in lowering sympathetic tone and blood pressure in the rostral ventrolateral medulla.

Pressor responsiveness of the sub-retrofacial nucleus and the midbrain reticular formation in the rat after 6-hydroxydopamine-induced lesions of ascending and descending catecholamine pathways.

We have recently shown that intracerebroventricular (icvt) administration of 6-hydroxydopamine (6-OHDA) inhibits centrally-evoked pressor activity. To see whether this effect is attributable to the disruption of descending bulbospinal or, alternatively, ascending suprabulbar catecholamine (CA) pathways, spontaneously hypertensive rats (SHR) were given localized intracerebral injections of 6-OHDA. One month later, pressor responses evoked by electrical or chemical stimulation in the rostral ventrolateral medulla or midbrain were examined under urethane anaesthesia. Injections of 6-OHDA into the medial forebrain bundle, which depleted noradrenaline and adrenaline in the hypothalamus, lowered basal blood pressure but potentiated the pressor responses to stimulation. In contrast, intraspinal injection of 6-OHDA raised basal blood pressure and attenuated pressor responses. This was accompanied by a partial depletion of adrenaline and the almost complete disappearance of noradrenaline in the spinal cord. Thus, the attenuation of pressor responses observed previously following icvt 6-OHDA can be attributed to an effect on spinal CA pathways. The effects on basal blood pressure suggest that, in SHR, ascending CA pathways are tonically pressor, while spinal CA pathways are depressor. Whilst it is unlikely, therefore, that spinal CAs mediate vasomotor outflow, the altered responses to stimulation after 6-OHDA suggest that central CA pathways can modulate the sensitivity of vasomotor neurones.

Limited baroreflex control of heart rate in young stroke-prone spontaneously hypertensive rats.

Controversy regarding possible differences of baroreflex gain in spontaneously hypertensive rats (SHR) and their relationship to the rise in blood pressure may be due in part to variations in the methods used to assess baroreflex function. In this study, we have compared the baroreflex control of heart rate in normotensive (Wistar-Kyoto, WKY) and stroke-prone spontaneously hypertensive (SHRSP) rats at 1 and at 7 months of age. Mean arterial pressure and heart rate were monitored in conscious rats following implantation of arterial and venous catheters. Phenylephrine and nitroprusside were given intravenously and the peak responses of mean arterial pressure and heart rate were recorded. In the young rats, these recordings were repeated under anaesthesia. Individual slopes for responses to phenylephrine or nitroprusside were obtained by linear regression. A single relationship covering both sets of responses was also obtained by fitting the data to a sigmoidal curve. The latter approach enabled the baroreflex to be represented as a single function which has a single determinant of gain, operates within defined limits and can be readily related to resting mean arterial pressure and heart rate. This approach demonstrated that: (1) in adult SHRSP, the baroreflex had reset to operate at higher resting levels of mean arterial pressure; (2) the range of heart rate control was smaller in both young and adult SHRSP compared with WKY; (3) average gain was slightly, but not significantly lower in adult SHRSP; (4) anaesthesia reduced heart rate range and average gain in both strains of rat.(ABSTRACT TRUNCATED AT 250 WORDS)

Similar baroreflex bradycardic actions of atrial natriuretic peptide and B and C types of natriuretic peptides in conscious rats.

OBJECTIVE: We have previously shown that atrial natriuretic peptide (ANP) modulates cardiac barosensitive afferent pathways to enhance reflex bradycardia in rats. The present study examined whether B-type natriuretic peptide (BNP) and C-type natriuretic peptide (CNP) also modulate heart rate reflex function. DESIGN: Baroreflex bradycardia was evoked by rapid (over 4-6 s) intravenous (i.v.) infusions of methoxamine (100 microg/kg; 'ramp' baroreflex technique) in the presence of infused i.v. natriuretic peptide and of vehicle (0.9% saline, 270 microl/h) in conscious adult Munich-Wistar rats. Initially a dose-response study to ANP (infused at 25, 50 and 100 pmol/kg per min i.v.) was performed in 10 rats to determine an appropriate dose for subsequent experiments with the other peptides. In a separate group of 11 animals, rat BNP-32 and rat CNP-22 were infused at 50 pmol/kg per min i.v. RESULTS: Reflex responses to ANP were dose-related, with a significant increase in baroreflex sensitivity of 50+/-15% at the 25 pmol dose, 102+/-10% at the 50 pmol dose and 117+/-11% at 100 pmol dose (all P<0.05). BNP and CNP (50 pmol/kg/min i.v.) substantially increased baroreflex bradycardia (by 115+/-17% and 62+/-15%, respectively; P<0.05) compared to vehicle infusion. CONCLUSIONS: Both BNP and CNP augmented baroreflex slowing of heart rate in response to rapid increases in blood pressure in rats. Whereas other reports have shown marked differences in cardiovascular responses between the natriuretic peptides, particularly with CNP, our findings demonstrate an important common action of ANP, BNP and CNP to facilitate vagal heart rate baroreflexes.

Involvement of imidazoline receptors in the baroreflex effects of rilmenidine in conscious rabbits.

OBJECTIVE: It has been suggested that imidazoline receptors rather than alpha2-adrenoceptors are involved in the sympathoinhibitory action of centrally acting antihypertensive drugs such as rilmenidine. In the present study, we examined the relative importance of alpha2-adrenoceptors and imidazoline receptors in modulating the renal sympathetic and heart rate (HR) baroreflex in response to central administration of rilmenidine in conscious normotensive rabbits. METHODS: In seven conscious rabbits, chronically instrumented with a fourth ventricular (4V) catheter, aortic and vena caval cuff occluders and a renal nerve electrode, we continuously recorded renal sympathetic nerve activity (RSNA), mean arterial pressure (MAP) and HR and assessed baroreflex MAP-RSNA and MAP-HR relationships with balloon-induced ramp rises and falls in MAP. Rabbits were treated with 4V rilmenidine (22 microg/kg) followed by 4V idazoxan (30 microg/kg; a mixed alpha2-adrenoceptor and imidazoline receptor antagonist) or 4V 2-methoxy-idazoxan (1 microg/kg; an alpha2-adrenoceptor antagonist with little affinity for imidazoline receptors). RESULTS: Rilmenidine lowered blood pressure by 24% and reduced both upper and lower plateaus of the renal sympathetic baroreflex curve, such that the RSNA range (difference between plateaus) was reduced by 40% (-32 +/- 10 normalized units). Curves were shifted to the left with the fall in MAP. Idazoxan restored MAP, maximum RSNA and the RSNA baroreflex range. By contrast the alpha2-adrenoceptor antagonist 2-methoxy-idazoxan caused only a partial recovery of MAP and RSNA baroreflex upper plateau and range (-9 +/- 2 mmHg, 29 and 33% lower than control). Both antagonists partially restored the HR baroreflex. CONCLUSION: These findings suggest that in conscious rabbits, both imidazoline receptors and alpha2-adrenoceptors are involved in the central antihypertensive and baroreflex actions of rilmenidine, but that activation of imidazoline receptors is more important for its renal sympathoinhibitory action.

Endothelin induces dopamine release from rat striatum via endothelin-B receptors.

The aim of the present study was to determine whether local administration of endothelin induces the release of dopamine in the rat striatum and to characterize and localize endothelin receptors in this brain region. Local injection of endothelin-1 (10 pmol) into the ventral striatum of urethane-anaesthetized rats caused an increase of 8 microM in the extracellular concentration of dopamine as measured by in vivo chronoamperometry. The peak increase in dopamine concentration occurred within 5 min of endothelin injection. Injection of the selective endothelin-B receptor agonist [Ala1.3,11.15]endothelin-1 (10 pmol) also caused an increase in extracellular dopamine concentration, suggesting that endothelin is acting at the endothelin-B receptor to elicit its effect. In rats with unilateral 6-hydroxydopamine lesions of the nigrostriatal pathway, the response to local injection of endothelin-1 (10 pmol) was significantly inhibited on the lesioned side as compared to the non-lesioned side. In contrast, pretreatment of the rats with the N-methyl-D-aspartate receptor antagonist dizocilpine maleate (5 mg/kg, i.p.) or the nitric oxide synthase inhibitor NG-nitro-L-arginine (3 mg/kg, i.p.) did not alter the endothelin-induced release of dopamine. In binding studies, addition of endothelin-1 displaced [125I]endothelin-1 with a Ki of 220 pM. The endothelin-B receptor antagonist BQ788 displaced [125I]endothelin-1 with a Ki of 120 nM, whereas the endothelin-A receptor antagonist BQ123 produced only a 25% displacement at 10 microM, suggesting that endothelin receptors in the striatum are of the endothelin-B subtype. In rats with unilateral 6-hydroxydopamine lesions of the nigrostriatal dopamine system, [125I]endothelin-1 binding was reduced by 53% in lesioned striatum compared to non-lesioned striatum, with no difference in the Kd. These data provide evidence that endothelin acts on a homogeneous population of endothelin-B receptors within the striatum to cause the release of dopamine and that a significant proportion of these receptors is located on dopaminergic neurons.

Impaired central stress-induced release of noradrenaline in rats with heart failure: a microdialysis study.

Sympathetic hyperactivity in rats with heart failure is associated with increased extracellular noradrenaline in the hypothalamic paraventricular nucleus at rest. However, it is unknown how this nucleus responds to stressful stimuli. In the present study we therefore examined the basal and stress-induced release of noradrenaline in the paraventricular nucleus of conscious Sprague-Dawley rats with heart failure measured by in vivo microdialysis. Basal noradrenaline concentration in the paraventricular nucleus of rats with heart failure was more than double that in sham-operated controls. Immobilization stress decreases noradrenaline levels in the paraventricular nucleus of rats with heart failure to 57% of baseline, while it increased in sham-operated controls to 228%. However, serum corticosterone was similarly elevated at 30 and 90 min post-stress in both experimental groups. We have shown that heart failure causes an impairment of the central noradrenergic system's response to acute sympatho-excitation but does not affect the hypothalamo-pituitary-adrenocortical response.

Involvement of imidazoline-preferring receptors in regulation of sympathetic tone.

We examined the contribution of imidazoline-preferring receptors (IPR) and alpha 2-adrenoceptors at different levels of the central nervous system in the antihypertensive and sympathoinhibitory actions of rilmenidine in 2 conscious animal models, the spontaneously hypertensive rat (SHR) and the normotensive rabbit. In conscious SHRs, we compared the potency of rilmenidine and clonidine administered intravenously into the lateral cerebral ventricle, the cisterna magna, and into the subarachnoidal space of the thoracolumbar spinal cord. In SHRs, we found that rilmenidine was more potent and more effective by the intrathecal than the intracisternal route. By contrast, clonidine was most effective after administration into the cisterna magna. Intravenous administration of rilmenidine or clonidine induced dose-dependent and prolonged decreases in blood pressure and heart rate. Neither rilmenidine nor clonidine altered mean arterial pressure or heart rate when given into the lateral cerebral ventricle. These data suggest that in SHRs the spinal cord may be an important site for the antihypertensive action of rilmenidine. We therefore characterized the receptor type involved. We observed in conscious SHRs that intrathecal post-treatment with idazoxan, a mixed alpha 2-adrenoceptor and IPR antagonist, abolished the antihypertensive effect of rilmenidine, whereas 2-methoxyidazoxan, a selective alpha 2-adrenoceptor antagonist, caused only a partial reversal of the blood pressure effects of rilmenidine. These results suggest that rilmenidine acts mainly through IPR rather than alpha 2-adrenoceptors in the spinal cord. In view of these findings, we compared the hypotensive actions of rilmenidine and clonidine, administered into the lateral cerebral ventricle, the cisterna magna, and the subarachnoid space of the thoracolumbar spinal cord in conscious normotensive rabbits. Both drugs were less potent and effective when administered intrathecally than intracisternally. These experiments suggest that the hypotensive action of rilmenidine and clonidine in the rabbit is mediated through receptors mainly located in the brainstem. Further, we found that idazoxan reversed the hypotensive action of rilmenidine more readily than 2-methoxyidazoxan. Surprisingly, both idazoxan and 2-methoxyidazoxan completely reversed the depressor effects of clonidine. Therefore, in the rabbit, rilmenidine acts through IPR located in the brainstem and clonidine acts predominantly through alpha 2-adrenoceptors. In conclusion, our studies demonstrate that IPR are involved in the vasodepressor action of rilmenidine in both conscious SHRs and rabbits. However, although the main site of action of rilmenidine in SHRs may be located in the thoracolumbar spinal cord, in the rabbit it appears to be in the brainstem.(ABSTRACT TRUNCATED AT 250 WORDS)

Cardiovascular functions of central noradrenergic and serotonergic neurons in conscious rabbits. Their contributions to the central actions of clonidine.

Synaptic release of transmitter from central noradrenergic (NA) and serotonergic (5HT) neurons was studied in intact and pontine decerebrate unanesthetized rabbits, following intracisternal injections of the selective neurotoxic drugs 6-hydroxydopamine (6-OHDA) and 5,6-dihydroxytryptamine (5,6-DHT). The NA and 5HT neurons both raise blood pressure through a suprapontine pathway, with 5HT neurons in series with NA neurons. Descending bulbospinal fibers have antagonistic effects on blood pressure, with NA release lowering blood pressure and 5HT release increasing it. The two transmitters also have antagonistic effects on the cardiac vagus, with NA neurons increasing vagal activity and 5HT neurons inhibiting it. Our results suggest that both NA and 5HT neurons contribute to the cardiovascular effects of clonidine. The actions of clonidine on blood pressure and heart rate mimic the effects of NA and are opposite those of 5HT released at synaptic sites in the bulb and spinal cord.