Vascular actions of MDMA involve alpha1 and alpha2-adrenoceptors in the anaesthetized rat

We have investigated the effects of methylenedioxymethamphetamine (MDMA, 'ecstasy'), i.v., on diastolic blood pressure (DBP) in pithed and pentobarbitone anaesthetized rats. In pithed rats, the non-selective 5-HT receptor antagonist methiothepin (0.1 mg kg(-1)) and the alpha2-adrenoceptor antagonists methoxyidazoxan and yohimbine (1 mg kg(-1)) showed significant alpha1-adrenoceptor antagonist potency, but methiothepin did not show alpha2-adrenoceptor antagonist potency. MDMA (1 and 5 mg kg(-1)) produced pressor responses which were significantly reduced by the alpha(1)-adrenoceptor antagonist prazosin (0.1 mg kg(-1)), yohimbine (1 mg kg(-1)) or methiothepin (0.1 mg kg(-1)), but not by the 5-HT2 receptor antagonist ritanserin (1 mg kg(-1)). In anaesthetized rats, antagonists revealed two phases with three components to the effects of MDMA (5 mg kg(-1)) on DBP: an initial pressor response, a later pressor component at 1 min, the sustained depressor response. Methoxyidazoxan, methiothepin or the combination ritanserin/prazosin significantly reduced the initial pressor response, although neither of the latter compounds alone had any effect. The pressor response to MDMA (5 mg kg(-1)) at 1 min was converted to a depressor response by prazosin and to a lesser extent methiothepin and methoxyidazoxan. The depressor response to MDMA (5 mg kg(-1)) was significantly reduced by methoxyidazoxan (0.1 mg kg(-1)), and by the noradrenaline re-uptake blocker cocaine 10 mg kg(-1) but not 1 mg kg(-1). However, the most marked reduction in the depressor response was produced by the combination of methoxyidazoxan and cocaine. It is concluded that the initial pressor response to MDMA (5 mg kg(-1)) in anaesthetized rats involves alpha2- and possibly alpha1-adrenoceptors and 5-HT2 receptors, the pressor component at 1 min is largely alpha1-adrenoceptor mediated, and the sustained depressor response involves alpha2-adrenoceptors.     

Contribution of alpha 2-adrenoceptors to the central cardiovascular effects of clonidine and S 8350 in anaesthetized rats.

1. The alpha 2-adrenoceptor agonist clonidine elicits centrally mediated effects through an interaction with both alpha 2-adrenoceptors and imidazoline binding sites. 2. We selected a new oxazoline derivative, S 8350, which competes with [3H]-yohimbine for binding to cerebral alpha 2-adrenoceptors (IC50, 67 +/= 17 nmol/L) and displays a higher affinity (35-fold) for alpha 2- than for alpha 1-adrenoceptors. 3. As observed for clonidine, intravenous (i.v.) administration of S 8350 resulted in a brief pressor effect followed by a prolonged hypotension. When S 8350 was administered i.v. to spinally pithed rats, only a rise in blood pressure was observed. 4. In order to discriminate the cardiovascular effects related to the central imidazoline receptor or alpha 2-adrenoceptor activation, the effects of intracisternal (i.c.) administration of clonidine and S 8350 were investigated in the rat. 5. In the anaesthetized rat, both clonidine and S 8350 displayed a profound central (i.c. route) hypotensive effect associated with a bradycardia. 6. The cardiovascular effects of S 8350 were abolished by the central administration of the selective alpha 2-adrenoceptor antagonist rauwolscine. Conversely, rauwolscine completely prevented bradycardia but it induced only a partial reversion of the hypotension elicited by clonidine. 7. These results suggest that central alpha 2-adrenoceptors are responsible for hypotension and bradycardia while imidazoline binding sites do not apparently contribute to heart rate control.     

Postjunctional alpha 2-adrenoceptors mediate venoconstriction in the hindquarters circulation of anaesthetized cats.

1 A study was made of the subtypes of postjunctional alpha-adrenoceptors which mediate arterial and venous constriction in the hindquarters circulation of anaesthetized cats, as measured by changes in perfusion pressure and vena cava blood flow, respectively. 2 It was found that, while noradrenaline caused constriction in both the arterial and venous compartments, methoxamine caused only arterial constriction. Clonidine and B-HT 920 also caused arterial and venous constriction although autodesensitization to both drugs occurred. 3 The ability of either prazosin or yohimbine to antagonize the constrictor effects of noradrenaline was also examined. It was found that the combination of both antagonist drugs abolished both the arterial and venous constrictor effects of noradrenaline. However, there was a greater prazosin-resistant response to noradrenaline in the venous compartment as compared with the arterial effects of noradrenaline. Yohimbine caused approximately equal reductions in the effect of noradrenaline in both arteries and veins, which was greater than that observed with prazosin. 4 These results suggest that, in the cat hindquarters, both alpha 1- and alpha 2-adrenoceptors are present in the arterial circulation, whereas there are mainly alpha 2-adrenoceptors in the venous circulation.     

Pharmacological evidence that alpha1-and alpha2-adrenoceptors mediate vasoconstriction of carotid arteriovenous anastomoses in anaesthetized pigs.

Vasoconstriction of carotid arteriovenous anastomoses may be involved in the therapeutic action of acutely acting anti-migraine agents, including the triptans and ergot alkaloids. While 5-HT1B/1D receptors mediate the effect of triptans, ergotamine and dihydroergotamine also interact with alpha-adrenoceptors. In the present study, we investigated the potential role of alpha1- and alpha2-adrenoceptors in mediating vasoconstriction of carotid arteriovenous anastomoses in anaesthetized pigs. Ten minute intracarotid infusions of phenylephrine (1, 3 and 10 microg kg(-1) min(-1)) or BHT 933 (3, 10 and 30 microg kg(-1) min(-1)) produced dose-dependent decreases in total carotid and arteriovenous anastomotic conductances; no changes were observed in the capillary fraction. The carotid vascular effects of phenylephrine and BHT 933 were selectively abolished by prazosin (100 microg kg(-1), i.v.) and rauwolscine (300 microg kg(-1), i.v.), respectively. The responses to phenylephrine and BHT 933 were not affected by the selective 5-HT1B/1D receptor antagonist GR127935 (500 microg kg(-1), i.v.). These results show that both alpha1- and alpha2-adrenoceptors can mediate vasoconstriction of carotid arteriovenous anastomoses in anaesthetized pigs. Since vasoconstrictor activity in this in vivo model is predictive of anti-migraine activity, an agonist activity at particularly the alpha2-adrenoceptor subtypes, in view of their less ubiquitous nature, could provide migraine abortive potential. Thus, the present results may aid further understanding of the mode of action of some current anti-migraine agents and may eventually be helpful in the development of future treatment in migraine.     

Functional evidence for heterogeneity of peripheral prejunctional alpha 2-adrenoceptors.

1. We have examined the potencies of a series of alpha 2-adrenoceptor antagonists in functional studies of prejunctional alpha 2-adrenoceptors in rat atrium and vas deferens, and compared potencies with affinities for the alpha 2A-ligand binding site of human platelet and the alpha 2B-site of rat kidney. 2. Antagonist potency in rat atrium was expressed as an EC30 (concentration producing 30% increase in the stimulation-evoked overflow of tritium in tissues pre-incubated with [3H]-noradrenaline). Antagonist potency in rat vas deferens was expressed as a pA2 or KB at antagonizing the inhibition by the alpha 2-adrenoceptor agonist xylazine of the isometric twitch to a single stimulus, or as an EC30. 3. In ligand binding studies, Ki values were obtained for the displacement by alpha-adrenoceptor antagonists of [3H]-yohimbine binding to human platelet or rat kidney membranes. 4. In functional studies, three antagonists (ARC 239, prazosin and chlorpromazine) distinguished between prejunctional alpha 2-adrenoceptors of rat atrium (EC30) and rat vas deferens (pA2) and showed 49, 12 and 7 times higher potency in rat atrium, respectively. ARC 239 was also 17 times more potent in rat atrium than rat vas deferens when EC30 values were compared. 5. The correlation of affinity for the alpha 2A-site of human platelet was better with prejunctional potency in rat vas deferens than rat atrium. 6. The correlation of affinity for the alpha 2B-site of rat kidney was better with prejunctional potency in rat atrium than rat vas deferens.(ABSTRACT TRUNCATED AT 250 WORDS)     

Comparison of central and peripheral alpha1-adrenoceptors

Summary A series of alpha-adrenergic agonists and antagonists having diverse chemical structure was examined for both central and peripheral alpha₁-adrenoceptor activity. The agonists tested included several novel aminotetralin derivatives which were potent and selective alpha₁-agonists. Peripheral alpha₁-activity was determined in the isolated rabbit ear artery; central alpha₁-receptor affinity was measured as the ability to inhibit ³H-WB 4101 binding to rat brain homogenates. In the agonist series, an excellent correlation between peripheral alpha₁-activity and central alpha₁-affinity was obtained, providing that partial agonists were excluded. Likewise, the receptor dissociation constant for blockade of the peripheral alpha₁-adrenoceptor correlated well with affinity for the central receptor for all of the alphaantagonists. These data support the conclusion that central and peripheral alpha₁-adrenoceptors are similar or identical.     

Dysfunctional presynaptic alpha 2-adrenoceptors expose facilitatory beta 2-adrenoceptors in the vasculature of spontaneously hypertensive rats.

Previous studies on spontaneously hypertensive rats (SHR) have yielded inconsistent information about functional aberrations of the presynaptic alpha 2- and beta 2-adrenoceptor-mediated modulation of sympathetic neurotransmitter release. In the present investigation we studied the capacity of presynaptic beta 2-adrenoceptors that enhance noradrenaline (NA) release in the portal vein of freely moving, unanesthetized SHR and normotensive Wistar rats (WR) using the beta 2-selective agonist fenoterol. The results show that the presynaptic beta 2-adrenoceptor population in SHR responds to significantly lower dosages of fenoterol than that in WR. The reason for this enhanced action, however, could not be attributed to the beta 2-adrenoceptor itself, nor to a diminished neuronal uptake of NA, but to a diminished responsiveness of the presynaptic alpha 2-adrenoceptor. Stimulation of presynaptic alpha 2-adrenoceptors with oxymetazoline (45 micrograms/min) decreased basal NA levels by 46% in WR and by 3% in SHR. Blockade of alpha 2-adrenoceptors, using 0.5 mg/kg yohimbine, induced a 4.86-fold rise in the basal NA level in WR but only a 1.89-fold rise in SHR. A subsequent dose of fenoterol, however, resulted in a further 2.5- and 2.6-fold rise in WR and SHR, respectively, indicating that there is a normal presynaptic beta 2-adrenoceptor population in the vasculature of SHR.     

Possible function of alpha 1-adrenoceptors in the CNS in anaesthetized and conscious animals

The influence of St 587 (2-(2-chloro-5-trifluoromethylphenylimino)imidazolidine), a selective alpha 1-adrenoceptor agonist which easily penetrates the blood-brain barrier, was tested on behavior and cardiovascular functions, respectively. The substance (up to 10 mg/kg subcutaneously (s.c.)) did not increase the exploratory activity of naive mice. The hexobarbitone 'sleeping' time in mice was reduced in a dose-dependent manner (St 587 ED50 = 14.4 mg/kg s.c.). Haloperidol 10 mg/kg s.c. induced catalepsy which was antagonized by St 587 in a dose-dependent manner (ED50 = 2.7 mg/kg i.p.). Conversely, the alpha 1-adrenoceptor-blocking agents prazosin and corynanthine elicited catalepsy in mice which had been treated with a subthreshold dose (2 mg/kg s.c.) of haloperidol; the ED50 values of the antagonists were 0.26 and 4.7 mg/kg i.p., respectively. In anaesthetized cats blood pressure and heart rate were not affected by 100 micrograms/kg St 587 injected into the left vertebral artery. In conscious dogs with beta-adrenoceptors blocked, the drug was without effect (100 micrograms/kg intracisternally) on vagally mediated reflex bradycardia, as evoked by intravenous noradrenaline injection. As a positive control the alpha 2-adrenoceptor agonist B-HT 920 which is equipotent to St 587 with respect to peripheral vasopressor effects in rats was injected with 10 micrograms/kg intracisternally and facilitated the reflex bradycardia. It is concluded that alpha 1-adrenoceptors within the brain mediate behavioral activation in states of CNS depression but remain without effect on cardiovascular centers.     

The antinociceptive effect of FR140423 in mice: involvement of spinal alpha(2)-adrenoceptors

We investigated the role of the spinal noradrenergic system in the antinociceptive effect of FR140423, 3-(difluoromethyl)-1-(4-methoxyphenyl)-5-[4-(methylsulfinyl)phenyl]py razole, by using the tail-pinch test in mice and various adrenoceptor antagonists. The antinociceptive effect of FR140423 injected i.t. was completely abolished by co-administration of the non-selective alpha-adrenoceptor antagonist phentolamine and the alpha(2)-adrenoceptor antagonist yohimbine but not by the alpha(1)-adrenoceptor antagonist prazosin or the beta-adrenoceptor antagonist propranolol. Oral administration of FR140423, at doses of 5-80 mg/kg, produced a dose-dependent antinociceptive effect with an ED(50) value of 19 mg/kg. This antinociception was abolished by i.t., but not i.c.v., injection of phentolamine and yohimbine (10 microg/mouse). These results suggest that alpha(2)-adrenoceptors in the spinal cord are involved in the antinociceptive effect of FR140423 against mechanical noxious stimulus as they are in the effect of morphine and clonidine.     

Involvement of alpha 2-adrenoceptors in the sacral micturition reflex in rats.

We have studied the effects of intrathecally-injected drugs that act on alpha-adrenoceptors in the urinary bladder reflex contractile activity evoked by continuous infusion of fluid into the bladder of anesthetized rats. Clonidine (10 and 30 micrograms) facilitated and yohimbine (100 micrograms) abolished the bladder contractile activity, and pretreatment with yohimbine (30 micrograms) inhibited the effect of clonidine (10 micrograms). Phenylephrine (60 micrograms) abolished the bladder contractile activity, but prazosin (40 micrograms) had no significant effect on it. The bladder contractions induced by electrical stimulation of the pontine micturition center were inhibited by yohimbine in a dose-dependent manner. These results suggest that transmission in the descending neurons from the pontine micturition center to the sacral parasympathetic neurons that control bladder motility is mediated by alpha 2-adrenoceptors in rats.     

Interaction between central alpha 1- and alpha 2-adrenoceptors on sympathetic tone in rats

The interaction between piperoxan and alpha 2-agonists on sympathetic tone was studied in rats. The sympatho-inhibitory effect of alpha 2-agonists (clonidine, guanfacine, B-HT 933) was assessed by recording heart rate in normotensive bilaterally-vagotomized rats. Clonidine (3 micrograms/kg, i.c.v.) and B-HT 933 (100 micrograms/kg, i.c.v.) induced a bradycardia which was fully reversed by piperoxan (30 micrograms/kg, i.c.v.). However, in rats treated with guanfacine, piperoxan induced a partial recovery of the bradycardic effect. The injection of a small dose of the specific alpha 1-adrenoceptor blocking drug, AR-C 239 (10 micrograms/kg, i.c.v.) which, by itself did not modify heart rate, completely inhibited the reversal effect of piperoxan in rats treated with clonidine, B-HT 933 or guanfacine. In rat brainstem membranes, B-HT 933 was found to bind to both alpha 1- and alpha 2-adrenoceptors and was as potent as clonidine in competing for alpha 1-sites bound by [3H]prazosin. On the other hand, in bilaterally vagotomized rats, piperoxan (30 micrograms/kg, i.c.v.) induced an increase in blood pressure and heart rate which was inhibited by previous administration of AR-C 239 (10 micrograms/kg, i.c.v.). These data suggest that, by inhibiting central alpha 2-adrenoceptors, piperoxan unmasks central alpha 1-adrenoceptor stimulation by endogenous catecholamines leading to an increase in the sympathetic tone, but a full recovery in heart rate could be observed only with the mixed alpha 1- and alpha 2-adrenoceptor agonists, clonidine and B-HT 933. In addition, these data further indicate that alpha 1-adrenoceptors are implicated in a tonic control of the sympathetic nerve activity in normotensive rats.     

Decreased density of alpha 2-adrenoceptors in medulla oblongata of spontaneously hypertensive rats

To study the role of medullary alpha-adrenoceptors in hypertension, we compared specific binding of [3H]prazosin and [3H]clonidine in different brain regions of spontaneously hypertensive rats (SHR), stroke-prone SHR (SHRSP), and normotensive Wistar-Kyoto rats (WKY). As compared with age-matched WKY, Bmax values for specific [3H]clonidine binding in the medulla oblongata were significantly lower in SHR and SHRSP at 16-24 weeks of age. In the SHRSP medulla oblongata, the decrease was more prominent in dorsomedial and ventrolateral regions than in the ventromedial region. Density of alpha 2-adrenoceptor binding sites was also decreased in the medulla oblongata of young (4-5-week-old) SHRSP. In contrast, there was no difference in Kd and Bmax values for medullary [3H]prazosin binding between WKY and SHRSP. The dorsomedial and ventrolateral regions of the SHRSP medulla oblongata showed significantly lower levels of norepinephrine (NE). Thus, the present study demonstrates that there is a specific loss of alpha 2-adrenoceptors in the medulla oblongata of SHR and SHRSP that may be partly involved in the pathogenesis of spontaneous hypertension.     

Neuronal responses to noradrenaline in the cerebral cortex: evidence against the involvement of alpha 2-adrenoceptors.

The technique of microelectrophoresis was used to test the hypothesis that alpha 2-adrenoceptors are involved in mediating the excitatory responses of single neurones to noradrenaline in the somatosensory cerebral cortex of the rat. In the first series of experiments the effects of two alpha 2-adrenoceptor antagonists, yohimbine and idazoxan (RX-781094), were compared on excitatory responses to noradrenaline, phenylephrine and acetylcholine. The response to noradrenaline was not more susceptible to antagonism by these drugs than the response to the alpha 1-adrenoceptor stimulant, phenylephrine. Yohimbine antagonized responses to all three agonists equally, while idazoxan antagonized responses to noradrenaline and phenylephrine equally with relative preservation of responses to acetylcholine. In the second series of experiments the effects of the selective alpha 2-adrenoceptor stimulant, UK-14304, were examined. UK-14304 produced weak and inconsistent excitations on a small number of cells; however, most of the cells did not respond to this drug. When applied continuously using low ejection currents, UK-14304 selectively and reversibly antagonized responses to noradrenaline and phenylephrine without affecting responses to acetylcholine. These results suggest that, in the somatosensory cortex of the rat, neuronal excitation to noradrenaline is unlikely to be mediated either wholly or partly by alpha 2-adrenoceptors. The antagonism of neuronal responses to noradrenaline and phenylephrine by idazoxan probably reflects the alpha 1-adrenoceptor antagonistic properties of the drug which is known to occur at higher concentrations.(ABSTRACT TRUNCATED AT 250 WORDS)     

3H-atipamezole binding sites in mouse cerebral cortex: possible involvement of alpha 2-adrenoceptors in sexual behavior.

Atipamezole is a new specific alpha 2-adrenoceptor antagonist. In this study, first, the presence of specific 3H-atipamezole binding sites in the sagittal and coronal sections of mouse brain was established using autoradiography. In vitro experiments with mouse cerebral cortex membranes indicated that d-medetomidine, a new alpha 2-adrenoceptor agonist structurally related to atipamezole, displaces labelled atipamezole more potently than noradrenaline. The saturation isotherm with d-medetomidine demonstrated high affinity binding with the apparent number of binding sites KD 1.36 nM and 760 fmol/mg, respectively. In the next series of experiments male mice were sacrificed immediately after copulation and cerebral cortex 3H-atipamezole and 3H-flumazenil binding was studied. Oxymetazoline and prazosin are known to label preferably alpha 2A and alpha 2B subtypes of alpha 2-adrenoceptors. Therefore, parallelly with noradrenaline both these compounds were used to determine non-specific binding of 3H-atipamezole. When noradrenaline or oxymetazoline were used as displacing agents copulation caused a significant increase of 3H-atipamezole binding sites. No significant changes were observed when prazosin was used. 3H-Flumazenil binding remained unchanged by copulation. The up-regulation of 3H-atipamezole binding sites indicates that not only alpha 2-adrenoceptors in the periphery but also in the CNS may participate in the regulation of sexual behavior. Moreover, in regulation of sexual behavior central alpha 2-adrenoceptors may be more important than benzodiazepine receptors.     

Diurnal changes in paraventricular hypothalamic alpha1 and alpha2-adrenoceptors and food intake in rats.

The prominent feeding rhythm evident in rats may reflect circadian variation in activity of feeding-relevant adrenoceptors within the hypothalamic paraventricular nucleus (PVN). In the present study, separate groups of rats were sacrificed at six time points (ZT0, ZT4, ZT8, ZT12, ZT16, ZT20) over a diurnal cycle. Food intakes were recorded during the 4-h period prior to sacrifice in each group. Brain sections were incubated with either an alpha1-adrenoceptor ligand (3H)-prazosin [(3H)-PRZ] or an alpha2-adrenoceptor ligand (3H) para-aminoclonidine [(3H)-PAC] prior to autoradiography analyses. Binding of (3H)-PRZ within the PVN varied as a function of the diurnal cycle, with significantly greater binding evident during the light phase of ZT0 (first 4 h of the light phase) and at ZT4, compared to nadir binding during the dark phase at ZT16 (first 4 h of the dark phase). Binding of (3H)-PAC within the PVN also varied as a function of the diurnal cycle, with significantly greater binding evident during the first 8 h of the dark phase (ZT16 and ZT20) than during the light phase. Food intake and alpha1-adrenergic binding were inversely related across the diurnal cycle. These results support the hypothesis that PVN adrenergic systems may be organized in an antagonistic fashion so as to modulate feeding in the rat.