P. B. M. W. M. Timmermans

Characterization of α-adrenoceptors participating in the central hypotensive and sedative effects of clonidine using yohimbine, rauwolscine and corynanthine☆

The central alpha-adrenoceptors responsible for mediating the clonidine-induced central hypotension in anaesthetized cats and sedation in mice have been characterized according to their sensitivities to the alpha-adrenoceptor antagonist yohimbine and its two diastereomeric congeners rauwolscine and corynanthine. Yohimbine and rauwolscine (1-10 microgram/kg) dose-dependently antagonized the central hypotensive response to clonidine (1 microgram/kg) applied 15 min later. Greater amounts of corynanthine (30-100 micrograms/kg) had to be administered to diminish the central depressor effect of clonidine. In these studies the drugs were infused via the left vertebral artery. The prolongation of the hexobarbitone-induced loss of the righting reflex in mice by clonidine (0.3 mg/kg, i.p.) was inhibited by previous treatment with yohimbine and rauwolscine (0.04-5 mg/kg, i.p.) in a dose-dependent manner, but not by corynanthine. Binding experiments with rat isolated cerebral membranes demonstrated the higher affinity of yohimbine and rauwolscine for the [3H] clonidine- than for the [3H]prazosin-specific binding sites. The reverse was found for corynanthine. The relative potencies of yohimbine, rauwolscine and corynanthine in inhibiting these central effects of clonidine are comparable to their order of efficacies in blocking peripheral alpha 2-adrenoceptors. Accordingly, clonidine-induced central hypotension and sedation are mediated by alpha 2-adrenoceptors.

Postsynaptic α1- and α2-adrenoceptors in the circulatory system of the pithed rat: Selective stimulation of the α2-type by B-HT 933

The antagonism by yohimbine (1 mg/kg, i.v.) of vasopressor responses in pithed rats was most pronounced towards B-HT 933 (dose ratio 18.3) and moderate towards clonidine (dose ratio 3.7) and especially L-phenylephrine (dose ratio 2.5). Prazosin (0.1 mg/kg, i.v.) had no effect on the pressor responses to B-HT 933, moderately affected those to clonidine (dose ratio 3.9), but strongly diminished those to L-phenylephrine (dose ratio 53). Phentolamine (1 mg/kg, i.v.) was devoid of a differential antagonism. The results obtained suggest a subclassification of postsynaptic alpha-adrenoceptors into alpha 1- and alpha 2-subtypes mediating pressor effects. B-HT 933 is a selective agonist and yohimbine an antagonist of postsynaptic alpha 2-adrenoceptors. L-Phenylephrine preferably stimulates and prazosin preferentially occupies the alpha 1-adrenoceptors. Clonidine is a potent agonist of both types and phentolamine behaves as a non-selective antagonist.

Possible subdivision of postsynaptic α-adrenoceptors mediating pressor responses in the pithed rat

SummaryAdditional evidence has been obtained indicating a possible subclassification of postsynaptic α-adrenoceptors into α1 and α2-subtypes. The pressor responses to the α-adrenoceptor agonists L-phenylephrine and guanfacine were quantified after i.v. administration to pithed rats. The α-sympatholytic drug yohimbine (1 mg/kg) displaced both dose-response curves to the right, but the effect was greatest for guanfacine. After prazosin (0.1 mg/kg) a 53-fold shift to the right was noticed for the dose-response characteristic of L-phenylephrine. Prazosin antagonized the effect of only the higher doses of guanfacine. The findings indicate that L-phenylephrine and prazosin preferentially interact with α1-adrenoceptors as agonist and antagonist, respectively. Yohimbine proved less selective than prazosin, but preferentially blocks postjunctional α2-adrenoceptors in the vascular wall. The results obtained with guanfacine may be interpreted to indicate that this drug acts on α2-adrenoceptors at lower doses and additionally stimulates α1-adrenoceptors at higher ones. Preliminary findings with corynanthine and rauwolscine support this interpretation.

VASCULAR SMOOTH-MUSCLE CONTRACTION INITIATED BY POSTSYNAPTIC ALPHA-2-ADRENOCEPTOR ACTIVATION IS INDUCED BY AN INFLUX OF EXTRACELLULAR CALCIUM

Vasoconstriction in pithed, normotensive rats elicited via stimulation of postsynaptic alpha 2-adrenoceptors by B-HT 920 was antagonized by EDTA and the calcium antagonists nifedipine, D 600 and verapamil, whereas pressor responses to the alpha 1-agonist methoxamine were unaffected. This indicates that vasoconstriction in vivo initiated via postsynaptic alpha 2-adrenoceptors requires an influx of extracellular calcium. Thus, the antihypertensive effect of calcium antagonists may be based upon a diminution of vascular tone maintained by postsynaptic alpha 2-adrenoceptors.

A lipophilic, selective α1 -adrenoceptor agonist: 2-(2-chloro-5-trifluoromethylphenylimino) imidazolidine (St 587)

Abstract A new compound (St 587) is described, which is a selective α 1 -adrenoceptor stimulating agent with lipophilic properties. This combination of characteristics is novel, since all α 1 -adrenoceptor agonists developed so far are hydrophilic. The α-adrenergic effects of 2-(2-chloro-5-trifluoromethylphenylimino) imidazolidine (St 587), a derivative of clonidine, were examined in several animal models. St 587 (1–10,000 μg/kg, i.v.) induced vasoconstriction in pithed, normotensive rats. This peripheral pressor activity was strongly antagonized by prazosin (0.1 mg/kg), but not affected by yohimbine (1 mg/kg). In intact, pentobarbitone-anaesthetized normotensive rats, St 587 (1–3,000 μg/kg, i.v.) evoked transient pressor responses, but a secondary fall in blood pressure and cardiac frequency was not observed. In pitched rats, St 587 (1–1,000 μg/kg) failed to modify the increase in heart rate produced by electrical stimulation of the cardioaccelerator sympathetic nerve fibres. St 587 (300 and 1,000 μg/kg) did not display central hypotensive activity, when injected into the left vertebral artery of anaesthetized cats. In addition, no hypotensive effect was observed when St 587 was administered i.v. to anaesthetized normotensive rats and cats. In mice, St 587 (10–10,000 μg/kg, i.p.) lacked sedative properties, since it did not prolong the hexobarbitone (75 mg/kg, i.p.)-induced loss of the righting reflex. The overall lipophilicity (log P′) of St 587 in the octanol/buffer (pH=7.4) reference system at 37°C amounted to 1.54. The experimental data suggest that St 587 is a lipophillic compound with selective α 1 - agonistic activity. The inability of St 587 to cause hypotension and sedation provides further evidence for the view that α 1 -adrenoceptors in the brain are not involved in the central hypotensive action and the sedation, caused by clonidine and related drugs. These effects are solely mediated by homogenous populations of α 2 -adrenoceptors.

Vasoconstriction mediated by postsynaptic α2-adrenoceptor stimulation

SummaryThe postsynaptic α-adrenoceptors involved in vasoconstriction brought about by B-HT 933 (2-amino-6-ethyl-4,5,7,8-tetrahydro-6H-oxazolo-[5,4-d]-azepin) administered i.v. to pithed, normotensive rats were characterized. The rate of onset of the hypertensive response to i.v. B-HT 933 is slower than that induced by (−)-phenylephrine, an agonist of α1-adrenoceptor. The antagonism of the α-adrenoceptor blocking drugs rauwolscine, yohimbine and corynanthine was quantified towards B-HT 933-induced increases in diastolic pressure. Rauwolscine (pA2=7.06) and yohimbine (pA2=6.83) were effective antagonists, whereas corynanthine proved much less potent (pA2=5.03). On the basis of the reported selectivity of yohimbine and its two diastereoisomers rauwolscine and corynanthine for α1- and α2-adrenoceptor, it is concluded that the postsynaptic α-adrenoceptors triggered by B-HT 933 are of the α2-type. B-HT 933 identifies a subclass of postsynaptic α2-adrenoceptor in vascular smooth muscle distinct from postsynaptic α1-adrenoceptor. Both types of α-adrenoceptors are likely to be involved in the mediation of vasoconstriction.

Participation of cardiac presynaptic α1-Adrenoceptors in the bradycardiac effects of clonidine and analogues

SummaryThe possible involvement of cardiac presynaptic α2-adrenoceptors in the acute negative chronotropic effect of intravenous clonidine and four selected analogues was tested in pentobarbitone-anaesthetized normotensive rats by establishing the following parameters: 1) the peripheral presynaptic activity (inhibition of tachycardia to stimulation of the spinal cord in pithed rats); 2) central hypotensive potency in intact rats (decrease in mean arterial pressure); 3) overall bradycardiac activity in intact rats (decrease in cardiac frequency); 4) sympathetic bradycardia in bilaterally vagotomized rat; and 5) vagal bradycardia in β-blocked rats. The lipophilicity of the imidazolidines (log P′) was measure in the octanol/buffer (pH=7.4) reference system at 37°C. Central hypotensive activity as well as vagal bradycardiac potency correlated with the ability to penetrate the central nervous system, as indicated by the overall lipophilic behavior (log P′) of the compounds. In contrast, brain penetration was not a prerequisite for the provocation of bradycardia in bilaterally vagotomized rats. This sympathetic bradycardia resembled peripheral presynaptic inhibition. The hydrophilic — poor brain-penetrating — substances already induced maximal sympathetic bradycardia at doses which did not induce hypotension or vagal bradycardia. The lipophilic drugs, however, only showed a preferentially sympathetic bradycardiac effect over vagal bradycardia at low doses. In intact pentobarbitone-anaesthetized rats, the log dose-bradycardiac response curves of the hydrophilic imidazolidines displayed a bisigmoidal character in contrast to their lipophlic analogues, i.e. at the lower dose-range of the hydrophilic drugs, bradycardia in intact rats resembled peripheral presynaptic inhibition, whereas a contribution of increased vagal activity only was significant after maximal sympathoinhibition was achieved. These data provide evidence for the view that the bradycardiac action of clonidine in pentobarbitone-anaesthetized normotensive rats is due, at least in part, to the functional role of cardiac presynaptic α2-adrenoceptors. The stimulation of these receptors will inhibit the sympathetic neurotransmission in the heart.

The interaction between prazosin and clonidine at α-adrenoceptors in rats and cats

Abstract In pithed rats prazosin (10μg/kg, i.v.) caused a prolonged antagonism of the hypertensive response to clonidine and (−)-noradrenaline, probably due to inhibition of vascular, postsynaptic α-adrenoceptors. The clonidine-induced reduction of the tachycardia evoked in pithed rats by electrical stimulation of cardiac sympathetic nerve fibres was antagonized by piperoxan and less effectively by prazosin, thus suggesting that prazosin displays a modest degree of cardiac presynaptic α-adrenoceptor blocking activity apart from its predominantly postsynaptic affinity. Prazosin (1 mg/kg, i.p.) significantly affected the hypotensive effect of clinidine (2 and 6 μg/kg, i.v.), but not the bradycardia induced by clonidine in pentobarbitone-anaesthetized, normotensive rats. Prazosin proved to be an effective hypotensive drug in anaesthetized cats. This action was peripheral as no central nervous origin could be demonstrated. Prazosin in low doses significantly reduced the central hypotensive effect of clonidine (1 μg/kg), injected into the left vertebral artery of chloralose-anaesthethized cats. Since the intravenous pretreatment with low doses of prazosin did not alter the central hypotensive response to clonidine, the interaction was likely to have occured within the brain-stem. Presumably, postsynaptic α-adrenoceptors in the brain, similarly to those in the periphery are inhibited by prazpsin, thereby preventing the central hypotensive effect of clonidine. It is submitted that clonidine and prazosin should not be combined in antihypertensive therapy in patients.

Lipophilicity and brain disposition of clonidine and structurally related imidazolidines.

SummaryThe apparent partition coefficients (P′) of clonidine and 27 of its structurally related imidazolidines were determined in the octanol/buffer (pH=7.4) system as a measure of lipophilic behaviour. Lipophilicity of the imidazolidines is limited to the free bases and the principle of additivity is valid for this series of structurally similar compounds.Brain concentrations of clonidine and a number of its derivatives, achieved at the moment of maximal decrease in blood pressure, were determined following intravenous administration to anaesthetized rats. These brain concentrations represent the maximally attainable values. The ratio of log brain concentration/ dose administered intravenously, log (Cbrain/Ci.v.), was employed as a measure of the penetration ability of the imidazolidines into the brain. The octanol/buffer (pH=7.4) system proved a satisfactory reference model in order to describe the transport process of the present imidazolidines from the blood to the brain. The penetration ability of these compounds into the brain could be expressed mathematically by a highly significant, parabolic relationship in log P′. Ideal lipophilic character for optimal brain concentrations is connected with a log P′ value of 2.16.

Identification of vascular postsynaptic alpha 1- and alpha 2-adrenoceptors in man.

We studied postsynaptic alpha-adrenoceptors in human blood vessels by measuring the influence on forearm blood flow induced by intra-arterial infusions of selective alpha 1- and alpha 2-adrenoceptor agonists (methoxamine, B-HT 933, clonidine and guanfacine) and antagonists (doxazosin and yohimbine). The studies were done in healthy volunteers, and forearm blood flow was measured by plethysmography. All agonists produced a significant and dose-dependent vasoconstriction. The effect of B-HT 933 was completely abolished by the concomitant infusion of yohimbine, whereas it was hardly influenced by doxazosin. The effect of methoxamine was prevented by doxazosin and little influenced by yohimbine. The vasoconstriction by clonidine and guanfacine was partially prevented by both doxazosin and yohimbine. The single intra-arterial infusion of yohimbine, as well as doxazosin, resulted in vasodilation. These findings provide strong evidence for the existence of postsynaptic alpha 1- as well as alpha 2-adrenoceptors, both mediating vasoconstriction and contributing to basal vascular tone. The (patho-)physiological significance of this subdivision of alpha-adrenoceptors remains to be elucidated.