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Brain cortex slices were preincubated with 3H-noradrenaline and superfused with physiological salt solution containing desipramine. We studied the inhibition of the electrically evoked tritium overflow caused by histamine in the presence of α-adrenoceptor ligands (mouse and rat brain cortex), and the inhibition caused by talipexole (the former B-HT 920) in the presence of H3-receptor ligands (mouse brain cortex). In mouse brain cortex slices, the inhibitory effect of histamine on the tritium overflow evoked by 36 pulses, 0.3 Hz was not changed by the α1-adrenoceptor antagonist prazosin, but increased by the α2-adrenoceptor antagonist rauwolscine. When the current strength or the duration of electrical pulses was reduced to compensate for the increase in evoked tritium overflow produced by rauwolscine, the latter still. enhanced the effect of histamine. The histamine-induced inhibition of tritium overflow evoked by 360 pulses, 3 Hz was not affected by the α1-adrenoceptor agonist phenylephrine but attenuated by the α2-adrenoceptor- agonist talipexole. Finally, the inhibition by histamine of the tritium overflow evoked by 3 pulses, 100 Hz was attenuated by talipexole but not affected by rauwolscine. Conversely,. the inhibitory effect of talipexole on tritium overflow elicited by 360 pulses, 3 Hz was slightly attenuated by the H3-receptor agonist R-(−)-α-methylhistamine but not, affected by the H3- receptor antagonist thioperamide. In rat brain cortex slices, histamine only tended to inhibit tritium overflow evoked by 360 pulses, 3 Hz, both in the absence of α-adrenoceptor antagonists and in the presence of prazosin. However, histamine markedly inhibited the evoked overflow in the presence of rauwolscine. Again, enhancement of the histamine-induced inhibition also occurred when the current strength or the duration of pulses was reduced in order to compensate for the increase in evoked tritium overflow produced by rauwolscine. The results suggest that the α2-autoreceptors and the H3-heteroreceptors at the noradrenergic nerve endings in the brain of mouse and rat interact with each other. Activation of the α2-autoreceptors decreases, whereas blockade of the activated (but not of the non-activated) α2-autoreceptors increases, the inhibitory effect of histamine. Activation of the H3-heteroreceptors slightly decreases, whereas blockade of the H3-receptors fails to affect, the inhibitory effect of talipexole.

The aim of the present investigation was to study, taking into account both quantitative and qualitative differences, the influence of full and partial α-adrenoceptor agonists on spontaneous myogenic activity in the rat portal vein. We found that the α-adrenoceptor agonists cirazoline, adrenaline, noradrenaline, phenylephrine, St 587, Sgd 101/75, B-HT 920 and UK-14,304 could increase the amplitude of the phasic myogenic contractions in the rat portal vein with apparent differences in EC50and E(max) values. In addition to an increase in phasic myogenic activity, the α-adrenoceptor agonists cirazoline, adrenaline, noradrenaline, and phenylephrine were also able (in higher concentrations) to increase the basal tone of the rat portal vein preparation, again with apparent differences in EC50and E(max) values. Changing the extracellular Ca2+concentration from 0.9 mmol/l to 2.5 mmol/l had no influence on the phasic character and the concentration range in which St 587 and UK-14,304 increased spontaneous myogenic activity, although changes in amplitude and frequency of the spontaneous myogenic contractions were less pronounced at a higher extracellular Ca2+concentration (2.5 mmol/l). By the use of Schild analysis with the competitive α-adrenoceptor antagonists prazosin (pA2= 8.74) and 5-methyl-urapidil (pA2= 8.37), it was established that the contractile responses to St 587 were mediated by the same α1-adrenoceptor subtype as the phasic and tonic type of contraction elicited by phenylephrine as described in a previous study. The concentration-response curve of UK-14,304 was significantly shifted to the right by low concentrations of prazosin (3 nmol/l-30 nmol/l), indicating stimulation of α1-adrenoceptors by UK-14,304 in the rat portal vein. The α-adrenoceptor antagonists phenoxybenzamine and chloroethylclonidine irreversibly blocked the contractile responses to St 587. Based on the method of receptor alkylation with phenoxybenzamine an affinity constant was calculated for St 587 (pK(a) = 5.91). Phenoxybenzamine was approximately 1000-fold more potent in inactivating α1-adrenoceptors than chloroethylclonidine. In conclusion there appeared to be a divergence in the excitation-contraction coupling of α1-adrenoceptors in the rat portal vein, which is reflected by two types of contraction (phasic versus tonic). The extent to which both the phasic and tonic types of contraction are stimulated by agonists depends on the affinity and intrinsic efficacy for each of the receptor-coupled effector pathways. Thus, partial and full agonism can only meaningfully be discussed if confined to one particular effector pathway.

Variations in the measurement of ligand's intrinsic activity between receptor subtypes is a common consequence of unequal receptor:G protein density ratios. We have investigated ligand activation at the alpha2-adrenoceptor (alpha2-AR) subtypes under defined expression conditions of one receptor molecule for one Galpha protein molecule using fusion proteins. Fusion between either a wt alpha2C AR or a mutant Thr382Lys alpha2C AR and a chimeric Galphaq/il protein displayed robust, transient (-)-adrenaline-mediated Ca2+ responses with similar potencies (pEC50: 7.78 and 7.66) and kinetic properties. A comparison of the intrinsic activities of alpha2 AR agonists found d-medetomidine to be the only compound with an efficacy similar to that of (-)-adrenaline. The Ca2+ responses as mediated by UK 14304, oxymetazoline and clonidine became more potent and efficacious at the Thr381Lys alpha2C AR, whereas the response as mediated by talipexole displayed a higher potency with an unaltered maximal response. Whereas only small differences in ligand's intrinsic activities between the wt alpha2A, alpha2B and alpha2C AR fusion proteins were observed with most ligands, oxymetazoline was virtually silent at the alpha2A AR while active as a partial and apparently full agonist at the alpha2C AR and alpha2B AR, respectively. The mutant alpha2 AR subtypes could be differentiated using the apparent positive efficacy of ligands that used to be defined as antagonists. The following rank order of maximal responses was observed for the Thr381Lys alpha2C AR: idazoxan approximately equals SKF 86466atipamezoledexefaroxan; Thr373Lys alpha2A AR: SKF 86466idazoxan = atipamezoledexefaroxan; and Thr370Lys alpha2B AR: atipamezoleidazoxan dexefaroxan. RX 811059 (10 microM) was the only compound to be completely silent at both the wt and mutant alpha2 AR subtypes. In conclusion, silent alpha2 AR ligands are probably rare in these specified alpha2 AR systems. Most antagonists may actually possess partial agonist properties at the alpha2 AR subtypes, which are facilitated by the same mutation in the distal portion of their third intracellular loop.

Previous study has shown that alpha2D-adrenoceptors are involved in modulation of peristalsis in the rat ileum. The aim of the present study was to determine the tissue location of alpha-adrenoceptors in the rat ileum by using a recently devised method. The pre-synaptic alpha-adrenoceptors were characterised by measuring the potencies of agonists to inhibit transmurally-evoked (1 ms pulses, 10 Hz, 8-10 s trains) contractions of the longitudinal and circular muscles and the affinities of antagonists. Post synaptic alpha-adrenoceptors were identified by screening agonists and antagonists in carbachol-contracted tissues. In the circular muscle the order of potencies for inhibiting transmurally-induced contraction was: clonidineor = oxymetazolineor = UK 14,304or = guanfacinetalipexolephenylephrineazepexole. The potency ratios relative to clonidine correlated to those previously derived using the rat ileum peristaltic reflex preparation. Most of the alpha-adrenoceptor agonists, however, caused only small inhibitions of the longitudinal muscle contraction in response to transmural stimulation, except phenylephrine and azepexole. RX 821002, yohimbine, rauwolscine, BRL 44408, phentolamine, idazoxan, ARC 239, and prazosin inhibited the effect of clonidine on the circular muscle response with apparent pK(B) values best correlated with pK(B) or pKi values derived from the rat ileum peristaltic reflex preparation and other tissues known to have the alpha2D-subtype. The rank order of potencies at inhibiting carbachol-induced responses of both muscle layers was: phenylephrineor = oxymetazolineclonidineor = talipexoleazepexoleguanfacine. UK 14,304 was inactive up to 10 microM. The EC50 value of each agonist on the longitudinal muscle was not significantly different to the corresponding value on the circular muscle. Prazosin was more potent than yohimbine at inhibiting the relaxant effect of phenylephrine in both muscle layers of carbachol-contracted tissues. It is concluded that the recently identified alpha2D-adrenoceptors of the rat ileum are located on cholinergic neurons controlling circular muscle contraction. The study also demonstrated the presence of postsynaptic alpha1-adrenoceptors involved in mediating relaxation in both muscle layers.

Mouse or rat brain cortex slices were preincubated with 3H-noradrenaline and superfused with physiological salt solution containing desipramine. We studied the effects of prostaglandin E2 (PGE2), prostaglandin D2 (PGD2) and related drugs on the electrically evoked (50 mA, 2 ms, 0.3 Hz) tritium overflow. PGE2 inhibited the electrically evoked tritium overflow from mouse brain cortex slices; the maximum effect of PGE2 (79010) was attenuated by the α2-adrenoceptor agonist talipexole (to 52010) and enhanced by the α2-adrenoceptor antagonist rauwolscine (to 92%). Rauwolscine was added to the superfusion medium in all subsequent experiments. The effect of PGE2 was readily reversible upon withdrawal from the medium and remained constant upon prolonged exposure of the tissue to the prostanoid. Studies with EP receptor agonists, mimicking the inhibitory effect of PGE2, showed the following potencies (pIC50): sulprostone (8.22); misoprostol (8.00); PGE2 (7.74); PGEZ (7.61); iloprost (5.86). The concentration-response curve of PGE2 was marginally shifted to the right by the EP1 receptor antagonist AH 6809 (6-isopropoxy-9-oxoxanthene-2-carboxylic acid; apparent pA2 3.97) and by the TP receptor antagonist vapiprost (4.50). AH 6809, by itself, did not affect the evoked overflow whereas vapiprost increased it. PGD2 inhibited the evoked overflow at high concentrations (pIC50 4.90); this effect was not altered by the DP receptor antagonist BW A868C (3-benzyl-5-(6-carboxyhexyl)-1-(2-cyclohexyl-2hydroxyethylamino)hydantoin), which, by itself, did not affect the evoked overflow. Indometacin slightly increased the evoked overflow and tended to increase the inhibitory effect of PGE2. PGE2 inhibited the electrically evoked tritium overflow also in rat brain cortex slices. The maximum effect (obtained in the presence of rauwolscine) was 61%; the pIC30 value was 7.67. The present study suggests that PGE2 inhibits noradrenaline release from mouse brain cortex via EP3 receptors and that its maximum effect is more marked in the mouse than in the rat. The inhibitory effect of PGD2 (in the mouse brain) does not involve DP receptors and may also be related to EP3 receptors. The EP3 receptors interact with a2-adrenoceptors and may be activated by endogenous prostanoids.

The interactions of talipexole (B-HT 920) and clonidine with selective alpha-adrenoceptor antagonists, yohimbine (alpha 2) and prazosin (alpha 1), as well as with dopamine receptor antagonists, metoclopramide (D2), domperidone (D2) and SCH23,390 (D1) were investigated in anaesthetized rabbits after i.v. administration. Both talipexole (0.03-0.1 mg/kg) and clonidine (0.01-0.03 mg/kg) dose-dependently induced hypotension and bradycardia. Talipexole had a shorter duration of action. The hypotensive effect of the alpha 2-adrenoceptor and D2 agonist talipexole (0.03 mg/kg) was antagonized by pretreatment with metoclopramide (3 mg/kg) or domperidone (0.3-3 mg/kg), but not with yohimbine (3 mg/kg), prazosin (0.1 mg/kg) or SCH23,390 (1 mg/kg). Its bradycardic effect was antagonized only by metoclopramide (3 mg/kg). The hypotensive and bradycardic effects of clonidine (0.03 mg/kg) were most effectively antagonized by yohimbine (0.3-3 mg/kg). These findings indicate that in anaesthetized rabbits after i.v. administration, talipexole may lower blood pressure by peripheral, and heart rate by central, dopamine D2 agonism.

The effect of pretreatment with a high dose of the alkylating agent N-ethoxycarbonyl-2-ethoxy-1,2-dihydroquinoline (EEDQ) (20 mg/kg, 24 h) on the intrinsic activity displayed by a series of full and partial dopamine D2 receptor agonists on prolactin regulating pituitary D2 receptors in male rats was studied. To increase baseline prolactin levels, gamma-butyrolactone in a dose inhibiting brain dopamine neurotransmission was given to all animals. In controls, i.e. rats not given EEDQ, supramaximal doses of all full and partial D2 receptor agonists tested decreased serum prolactin levels with greater than 80%. While the intrinsic activities of the dopamine precursor 1-DOPA and of the full agonists (+)-3-PPP, 5-OH-DPAT, B-HT 920 (talipexole), apomorphine, and NPA (R-(-)-N-n-propylnorapomorphine) were not affected by pretreatment with EEDQ, the effects of supramaximal doses of the partial agonists (-)-HW-165, TDHL (terguride), SDZ208-911, (-)-3-PP) (preclamol), and SDZ 208-912 were reduced to 66%, 74%, 59%, 100%, and 100%, respectively. The effect of EEDQ on the intrinsic activity displayed by the various agonists corresponds inversely to the intrinsic efficacy displayed by the drugs in other models of D2 receptor function with one exception only; thus, the prolactin suppressive effect of (-)-3-PPP was more effectively antagonized by EEDQ than would have been predicted from the intrinsic efficacy usually attributed to the drug. Since the dose of EEDQ used in the present study has previously been shown not to decrease D2 receptor density in the pituitary as measured using in vivo radioligand binding, it is suggested that alkylation of D2 receptors may change the conformation of the individual receptor complexes in a way that decreases the responsiveness to partial but not full agonists.

Previous studies using clonidine indicate that alpha 2-adrenoceptors are involved in suppressing opiate-withdrawal symptoms. However, clonidine may act as a partial agonist at alpha 2-adrenoceptors and it also possesses significant alpha 1-receptor agonist activity. The aim of this study was to determine the role of alpha 2-adrenoceptors in the expression of opiate withdrawal signs using morphine-dependent rats. A range of agonists were selected for study on the basis of their differential preferences for alpha-adrenoceptors. Hooded Wistar rats were made physically dependent on morphine (s.c. injection of an emulsion releasing a total of 250 mg/kg of morphine base over 48 h). Test drugs were injected s.c. followed by naloxone (10 mg/kg i.p.) 20 min later. The incidence of 5 selected withdrawal signs was recorded during the following 20 min. The alpha 2-adrenoceptor agonists displayed different profiles of activity. Azepexole (1-10 mg/kg) reduced all signs. Clonidine (80-800 micrograms/kg) reduced all signs except paw shakes while guanfacine (25-250 micrograms/kg) reduced all except jumping and diarrhoea. Talipexole (0.1-1 mg/kg) reduced all signs except diarrhoea which was not affected and jumping which was markedly enhanced. UK 14,304 (80-800 micrograms/kg) reduced jumps, potentiated paw shakes but did not affect body shakes, teeth chattering or diarrhoea. The results suggest that there are subpopulations of alpha 2-adrenoceptors that modulate the expression of opiate withdrawal signs and/or that some of the drugs used affect receptors other than alpha 2-adrenoceptors.

1. The azepine derivatives B-HT 920 and B-HT 933 did not increase the motor activity of mice pretreated with reserpine or reserpine plus apomorphine, indicating that they do not stimulate postsynaptic dopamine receptors or noradrenaline α1-receptors in the brain. 2. The motor activity of mice not pretreated with reserpine was reduced by a low dose of B-HT 920 and by B-HT 933. The α1-adrenoceptor antagonist yohimbine reversed the sedation induced by B-HT 933, but not that induced by B-HT 920. 3. B-HT 933 and a high dose of B-HT 920 retarded the α-methyltyrosine-induced disappearance of noradrenaline in the mouse brain by a yohimbine-sensitive mechanism. 4. The α-methyltyrosine-induced disappearance of dopamine in the mouse brain was decelerated by a low dose of B-HT 920 and to a smaller degree by B-HT 933. The effects were inhibited by the dopamine receptor antagonist haloperidol. The effect of B-HT 933, but not that of B-HT 920, was partly antagonized by yohimbine. 5. The enhanced synthesis of dopamine in the corpus striatum of mice following treatment with gammabutyrolactone was completely antagonized by B-HT 920, but not by B-HT 933, via a haloperidol-sensitive mechanism. The synthesis of noradrenaline in the brain stem and in the hemispheres was reduced by B-HT 933 via a yohimbine-sensitive mechanism. 6. The results indicate that B-HT 920 can selectively and potently stimulate dopamine autoreceptor and that B-HT 933 can preferentially stimulate noradrenaline autoreceptors (α2-adrenoreceptors). These actions might cause the decreases in motor activity observed in mice not pretreated with reserpine.

Interactions between the putative calcium entry promotor Bay k 8644 and both α1- and α2-adrenoceptor mediated increases in diastolic pressure were studied in the pithed normotensive rat. The α2-adrenoceptor mediated pressor responses elicited by B-HT920, TL-99, DP-6,7-ADTN, and B-HT958 were potentiated by Bay k 8644, reflected by a leftward shift and an increase in the maximum of the log dose-pressor response curves. The α1-adrenoceptor-mediated effects elicited by cirazoline, methoxamine, (-)-amidephrine, St 587, (-)-phenylephrine and Sgd 101/75 were less enhanced by Bay k 8644. Only a leftward shift of the dose-response curves was observed, which was most pronounced for (-)-phenylephrine and Sgd 101/75. The α1- and α2-adrenoceptor-mediated pressor components of (-)-noradrenaline were similarly distinguished by Bay k 8644 as observed for the selective α1- or α2-adrenoceptor agonists. Effects of Bay k 8644 on the increase in diastolic pressure mediated by B-HT 920, St 587 and cirazoline were also studied after pretreatment with the calcium entry blocker nifedipine. After additional pretreatment with nifedipine the potentiation by Bay k 8644 observed for B-HT 920 and St 587 was more pronounced. The presence of nifedipine had no effect on the interaction between Bay k 8644 and cirazoline. It is concluded that Bay k 8644 behaves as a mirror image of nifedipine. The observation that Bay k 8644 enhances α2-adrenoceptor mediated pressor effects more effectively than - α1-adrenoceptor-mediated increases in diastolic pressure is in accordance with the hypothesis of the more pronounced calcium dependency of α2-adrenoceptor mediated pressor responses. The data obtained for St 587 and (-)-phenylephrine are in apparent contradiction to the finding that the pressor responses to the former drug are more markedly inhibited by calcium entry blockade than those of the latter. It is suggested that St 587 employs calcium channels which are already maximally modulated and that (-)-phenylephrine makes use of calcium channels which are in a rather inactive state. The hypothesis is put forward that the intrinsic activity of α2-adrenoceptor agonists reflects their ability to bring calcium channels in an active state.