An analysis of central adrenoceptors for control of cardiovascular function

1. In dogs anaesthetized with pentobarbitone sodium, bradycardia with hypotension occurred on intracerebroventricular (i.c.v.) injection of noradrenaline (50-200 mug) or phenylephrine (100-400 mug), but tachycardia with hypotension occurred on i.c.v. injection of isoprenaline (100-200 mug).2. These cardiovascular responses were central effects, and from the results obtained after bilateral vagotomy, removal of both stellate ganglia and transection of the upper cervical cord, it was evident that the efferent nervous pathway for all these effects was the sympathetic nervous system.3. An i.c.v. injection of the alpha-adrenoceptor blocking agent phenoxybenzamine (10 mg) blocked the bradycardia and hypotension produced by noradrenaline or phenylephrine, and an i.c.v. injection of a beta-adrenoceptor blocking agent, either propranolol (2 mg) or N-isopropyl-p-nitrophenyl-ethanolamine (INPEA) (10 mg), blocked the tachycardia and hypotension produced by isoprenaline.4. The cardiovascular effects produced by i.c.v. injection of the three sympathomimetic amines could be reproduced in cross-circulation experiments in the recipient dog when the amines were injected into its head circulation, and the effects of noradrenaline and phenylephrine, but not those of isoprenaline, were abolished by the alpha-adrenoceptor blocking agent yohimbine (1 mg/kg) injected intravenously into the donor dog.5. It is concluded that the central alpha-adrenoceptors are concerned with bradycardia and the central beta-adrenoceptors with tachycardia, but that both receptors are concerned with hypotension.     

Regarding the unitary theory of agonist and antagonist action at presynaptic adrenoceptors

1. The linkage between potentiation of field stimulation-induced noradrenaline release and blockade of the presynaptic inhibitory effect of exogenous noradrenaline by a presynaptic antagonist was examined in superfused rabbit aorta preparations. 2. Rauwolscine clearly potentiated the release of noradrenaline in response to 100 pulses at 2 Hz but reduced the capacity of noradrenaline to inhibit transmitter release to a questionable extent, and then only when comparisons were made with untreated, rather then to rauwolscine-treated, controls. 3. Aortic preparations exposed for 60 min to rauwolscine followed by superfusion with antagonist-free Krebs for 60 min retained the potentiation of stimulation-induced transmitter release but no antagonism of the noradrenaline-induced inhibition could be detected at either of two noradrenaline concentrations when comparisons were made with rauwolscine treated controls. 4. Comparisons of the inhibitory effect of exogenous noradrenaline (1.8 x 10-6 M) on transmitter efflux in the presence and absence of rauwolscine pretreatment revealed that the antagonist enhanced rather than antagonized the presynaptic inhibition by noradrenaline. 5 It is concluded that the unitary hypothesis that asserts that antagonist enhancement of transmitter release and its blockade of noradrenaline induced inhibition are manifestations of a unitary event are not supportable.     

Evidence against a functional link between noradrenaline uptake mechanisms and presynaptic alpha-2 adrenoceptors

Summary Slices prepared from rat cerebral cortex were labelled with ³H-noradrenaline and superfused. Electrical field stimulation was carried out 15 min (S₁) and 45 min (S₂) after the start of collection of 5-min samples using 4 pulses delivered at 100 Hz. Drugs acting at ₂-adrenoceptors were added 20 min before S₂, and their effects were evaluated using the S₂/S₁-ratio. The ₂-adrenoceptor antagonists idazoxan (1 mol/l) and rauwolscine (1 mol/l) failed to increase stimulation-evoked overflow of radioactivity in the absence or presence of the noradrenaline reuptake inhibitor desipramine (1 gmol/l). This indicates that the duration of electrical stimulation was too short to allow development of ₂-adrenoceptor-mediated auto-inhibition by released noradrenaline. The effect of clonidine (3–1000 nmol/l) on stimulation-evoked overflow of radioactivity was tested in the absence and presence of three different reuptake inhibitors (desipramine, 1 ol/l; maprotiline, 1 ol/l; cocaine, 10 mol/l). The analysis yielded identical concentration-response curves under all conditions. These results argue against an action of inhibitors of neuronal reuptake of noradrenaline at the presynaptic ₂-adrenoceptor and against the concept of a functional link between uptake site and receptor. Send offprint requests to E. A. Singer     

Evidence against the unitary hypothesis of agonist and antagonist action at presynaptic adrenoceptors.

1 The concept that presynaptic receptors regulate noradrenergic transmitter release via a system of inhibitory receptors mediating negative feedback relies on a supposed association between increases in stimulation-induced efflux of [3H]-noradrenaline by antagonists and blockade by them of the inhibitory effects of exogenous noradrenaline. 2 It was shown in guinea-pig ureter, that yohimbine (3 X 10(-7)M), a presumed selective presynaptic antagonist, increased transmitter efflux substantially at 1 Hz and 5 Hz with 100 pulses, purportedly representing antagonism of the inhibitory effect of locally released noradrenaline but did not reduce the inhibitory effect of exogenous noradrenaline (1.8 X 10(-6)M or 1.8 X 10(-7)M) except in one case. 3 Additionally, the inhibitory effect of oxymetazoline (1.0 X 10(-7)M or 1.0 X 10(-8)M) on stimulation-induced efflux was in no way antagonized by yohimbine (3 X 10(-7)M). 4 It is concluded that the increased efflux of [3H]-noradrenaline produced by antagonists and the decreased efflux produced by exogenous agonists may represent actions at different loci and that the hypothesis of presynaptic feedback regulatory sites is still not substantiated.     

Modulation of noradrenaline release by presynaptic alpha-2 and beta adrenoceptors in rat atria

Summary The effect of pretreatment with the beta-2-selective adrenoceptor agonist, (+-)-clenbuterol (0.3 mg/kg, twice daily, 14 days) on prejunctional alpha-2- and beta-adrenoceptors was studied in rat atria. When atria from non-pre-treated rats had been preincubated with (³H)-noradrenaline, (-)-isoprenaline (0.02 to 4.0 M) did not affect tritium overflow evoked by stimulation of the cardioaccelerant nerves, but a higher concentration (40 M) decreased it. Blockade of prejunctional inhibitory alpha-2-adrenoceptors by yohimbine (0.03, 0.3 and 0.8 M) enhanced the overflow of tritium. In the presence of yohimbine, isoprenaline (1.2 M) significantly increased stimulation-induced transmitter overflow, suggesting that in rat atria the facilitatory effect of isoprenaline mediated via prejunctional beta-adrenoceptors, is masked by the dominant influence of inhibitory alpha-2-adrenoceptors. (-)-Propranolol (0.1 M) prevented the isoprenaline-induced increase in atrial rate and the isoprenaline-induced enhancement of transmitter release in the presence of yohimbine (0.3 M), but did not modify by itself the stimulation-induced efflux of tritium, suggesting that neuronally released noradrenaline failed to activate facilitatory prejunctional beta-adrenoceptors. When atria from clenbuterol-pretreated rats had been preincubated with ³H-noradrenaline, the facilitatory effect of yohimbine 0.03 and 0.3 M was markedly enhanced and, in this case, isoprenaline (1.2 and 12.0 M) failed to cause its facilitatory effect in the presence of the alpha-2-adrenoceptor antagonist. Propranolol did not modify the facilitatory effect of yohimbine. No changes in the isoprenaline-induced increase in atrial rate were observed in clenbuterol-treated rats. In addition, the treatment reduced the positive chronotropic effect of nerve stimulation without affecting the response to exogenous noradrenaline, suggesting a reduction in the transmitter release induced by nerve stimulation in clenbuterol-treated rats. These results suggest that chronic treatment with clenbuterol desensitizes facilitatory prejunctional beta-adrenoceptors, without affecting the postsynaptic beta-adrenoceptors, thus implying that prejunctional beta-adrenoceptors possess properties of the beta-2-subtype. Send offprint requests to M. A. Enero at the above address     

Rapid and reversible desensitisation of vascular and platelet alpha2 adrenoceptors

Summary 1. The effects of intravenous infusion with the alpha₂ adrenoceptor selective agonist alpha methylnoradrenaline on pressor responses to alpha adrenoceptor agonists, alpha₂ adrenoceptor mediated platelet aggregation and adenylate cyclase were examined in conscious rabbits. 2. Pressor responses to alpha methylnoradrenaline but not phenylephrine were decreased in a dose dependent manner during methylnoradrenaline infusion at all times examined. 3. Recovery of these responses after stopping infusion was dependent on both the dose infused and the duration of the infusion. 4. Alpha methylnoradrenaline infusion resulted in a dose and time dependent decrease in the pro-aggregatory response of platelet to adrenaline without any significant change in the response to ADP or in the number of [³H] yohimbine binding sites. 5. The ability of PGE₁ to stimulate adenylate cyclase was not influenced by alpha methylnoradrenaline infusions. However, reversal of this stimulation by adrenaline was decreased by relatively long (30 min) infusions of the highest dose of alpha methylnoradrenaline examined. 6. It is concluded that alpha methylnoradrenaline infusions resulted in desensitisation of all the alpha₂ adrenoceptor mediated responses examined. However the time course for the desensitisation apparently differed according to the response examined. Send offprint requests to C. A. Hamilton at the above address     

Streptozotocin-induced diabetes modulates presynaptic and postsynaptic function in the rat ileum

Altered gastrointestinal motility frequently occurs in diabetic patients and also in animal models of diabetes but the underlying causes are not clear. In the present study, contractile responses to agonists and electrical field stimulation (EFS) and the inhibitory actions of an adenosine A(1) receptor agonist were investigated on ilea from 8-week streptozotocin (STZ)-induced diabetic rats. Contractile responses to carbachol, prostaglandin F(2 alpha) (PGF(2 alpha)), the calcium ionophore A23187 and to EFS were increased in diabetic tissues compared to controls. In contrast, the inhibitory effects of a potent and selective adenosine A(1) receptor agonist N(6)-cyclopentyladenosine (CPA) on electrical field stimulation-evoked contractions were decreased in diabetic tissues compared to controls but its ability to relax carbachol-contracted tissues was unaltered. These results suggest that diabetes may cause alterations at both pre- and postsynaptic sites and this may lead in turn to the gastrointestinal complications seen in diabetic patients.     

Identification of adrenoceptors and dopamine receptors mediating vascular responses in the superior mesenteric arterial bed of the rat

The nature of the adrenoceptors and dopamine receptors mediating vascular responses in the in-situ blood perfused superior mesenteric arterial bed of the rat have been studied. alpha 1-Adrenoceptor agonists produced vasoconstriction but alpha 2-agonists had no significant effect on vascular resistance. The vasoconstrictor effects of noradrenaline were antagonized by low doses of prazosin (26 nmol kg-1 i.v.). Isoprenaline and salbutamol produced vasodilation when the vasculature was preconstricted with arginine vasopressin. The responses to isoprenaline were potently antagonized by propranolol (1.69 mumol kg-1 i.v.) and weakly but significantly reduced by practolol (3.75 mumol kg-1 i.v.) whereas the responses to salbutamol were unaffected by the same dose of practolol. After preconstriction of the vasculature and alpha-adrenoceptor blockade, dopamine and apomorphine produced dilator responses with both compounds producing the same maximal response and apomorphine being 1.8 times more potent than dopamine. The dopamine responses were present after the animals had been pithed and were resistant to spiperone (506 nmol kg-1 i.v.) but were antagonized by cis-alpha-flupenthixol (460 nmol kg-1 i.v.). These results suggest that this vascular bed possesses vasoconstrictor alpha 1- but not alpha 2-adrenoceptors, vasodilator beta 1- and beta 2-adrenoceptors and vasodilator dopamine receptors which appear similar to the D1-type found centrally.     

The vascular system. An overview of structure and function

It is the function of the vascular system, through a complex network of arteries, capillaries and veins, to maintain cellular homeostasis. As research scientists it is necessary to understand not only some of the basic properties of the blood vessel itself but also how these vessels differ in cellular and physiological function. This review provides an overview of the basic physiological and pharmacological tenets of blood vessels. It also briefly describes in vivo and in vitro methods used in the measurement of blood flow and blood vessel function. It is hoped that this review will provide readers of this focussed issue of the Journal of Pharmacological & Toxicological Methods with an appreciation of the many mechanical, electrical and biochemical methodologies described within this issue.     

Further investigation of the sites of vascular alpha 1 and alpha 2 adrenoceptors in the anaesthetised spontaneously hypertensive rat

We have looked at the role played by alpha 1 and alpha 2 adrenoceptors in the control of blood pressure in the anaesthetised spontaneously hypertensive rat (SHR) by examining the blood-pressure-lowering effects of the alpha 1-adrenoceptor antagonist prazosin and the alpha 2-adrenoceptor antagonist rauwolscine in 5 groups of animals: unoperated, adrenal demedullated, shamoperated, sympathectomised with 6-hydroxydopamine, and vehicle treated. Prazosin (1 mg/kg) produced significant falls in diastolic blood pressure (DBP) in all groups of rat but the fall was significantly less in sympathectomised animals. In the presence of prazosin (1 mg/kg), rauwolscine (0.1-1 mg/kg) lowered DBP in all groups of rat except the adrenal demedullated animals. In the absence of prazosin, rauwolscine (0.1 mg/kg) produced a significant fall in DBP only in sympathectomised animals. These results demonstrate that alpha 2 adrenoceptors are involved in blood pressure control in the intact or sympathectomised SHR but not following adrenal demedullation, suggesting that circulating catecholamines rather than neurally released noradrenaline (NA) are responsible for these alpha 2-adrenoceptor-mediated pressor effects. In contrast, only alpha 1-adrenoceptor-mediated pressor responses are reduced by sympathectomy, suggesting that neurally released NA acts mainly on alpha 1-adrenoceptors.     

Localization in the CNS of adrenoceptors which facilitate a cardioinhibitory reflex

Summary In dogs (pentobarbitone, 25 mg/kg) the brain was removed rostrally to the pons, leaving the cerebellum intact (decerebrate animals). In other animals the cerebellum was additionally removed (bulbar animals). In all animals -adrenoceptors were blocked by toliprolol (5 mg/kg s.c.). Angiotensin (0.025–0.3g/kg) was repeatedly injected i.v. and the resulting maximal reflex bradycardia was recorded. Intracisternal (i.ci.) injection of clonidine, 0.5 or 1 g/kg, in decerebrate or i.v. injection of 10 or 30 g/kg in bulbar animals significantly facilitated the reflex bradycardia. This effect was antagonised by a subsequent injection of piperoxan, 50 g/kg i.ci. in decerebrate or 1 mg/kg i.v. in bulbar animals. It is concluded that the facilitatory action of clonidine is mediated by -adrenoceptors within the medulla oblongata.Presented in part at the 15th Spring Meeting of the German Pharmacological Society, Mainz, March 1974.     

Effects of glucocorticoids on 5-HT1A presynaptic function in the mouse

8-OH-DPAT, a selective 5-HT_1A agonist, produced a hypothermic response in mice at a dosage of 0.5 mg/kg. Administration of corticosterone-21-acetate (0.5, 5 and 50 mg/kg, daily for 3 and 10 days) produced a dose-dependent attenuation of this hypothermic response in mice. When all controls and corticosterone treated mice were retested, 14 days after initial testing, they did not differ in the hypothermic responses induced by 8-OH-DPAT. Mice treated with aldosterone (50 mg/kg), dexamethasone (50 mg/kg) and the specific type 2 corticosteroid receptor agonist, 11b,17b-dihydroxy-21-methyl-17a-pregna-1,4,6-trien-20-yn-3-one (RU26988, 30 mg/kg) for 10 days, did not differ from vehicle treated controls in the hypothermic response to 8-OH-DPAT. Mice administered corticosterone-21-acetate (30 mg/kg, daily) for 10 days displayed a motor behavioural syndrome, which was not seen in controls, when injected with 5-hydroxytryptophan (5-HTP, 100 mg/kg) 15 min after the injection of carbidopa (25 mg/kg). This was significantly decreased by pretreatment with the 5-HT_1A receptor antagonist 1-(2-methoxyphenyl)-4-(4-phthalimidobutyl)-piperazine (NAN-190 5 mg/kg, 30 min prior to administration of carbidopa). Taken together, this evidence is compatible with a specific corticosterone induced facilitation of 5-HT release due to attenuation of inhibitory 5-HT_1A autoreceptor function.     

The effects of chronic administration of adrenaline on the function and number of adrenoceptors in the rabbit

Chronic (10-day) intravenous infusions of adrenaline (0.05 mumol/kg/h) were given to rabbits via osmotic minipumps implanted at the femoral vein. Blood pressure, heart rate, and plasma catecholamine concentrations were measured five times during the period of infusion. Tenfold elevations in circulating adrenaline levels were achieved within 24 h of commencing infusion and maintained throughout the study. This increase in plasma adrenaline was not accompanied by significant changes in blood pressure or heart rate. Rabbits were killed after 10 days: blood was withdrawn for platelet aggregation studies. Kidney, heart, and lung were also collected and alpha 2-adrenoceptor number on platelets and kidney measured using [3H]yohimbine. Beta adrenoceptors on platelets, lymphocytes, heart, and lung were quantified using [125I]iodocyanopindolol. Adrenaline infusion led to a significant reduction in platelet aggregation responses to adrenaline (0.001-100 microM), together with a decrease in alpha 2-adrenoceptor number on platelets, but no significant decrease in kidney alpha 2-adrenoceptors. A significant decrease in the density of beta adrenoceptors on heart and lung membranes was observed with no reduction in platelet and lymphocyte beta-adrenoceptor number. Thus adrenaline-induced down-regulation of adrenoceptors in the rabbit was dependent on the location and subtype of adrenoceptor.     

Heterogeneity of presynaptic muscarinic receptors located on different tissue preparations

Three different tissue preparations were used to demonstrate the heterogeneity of presynaptic muscarinic receptors that modulate neurotransmitter release. The presynaptic antimuscarinic potency of several muscarinic antagonists was characterized with the enhancement of the neurotransmitter release evoked by electrical stimulation on the guinea-pig ileum Auerbach plexus, the guinea-pig atrium and the rat brain cortex. Presynaptic muscarinic receptors located on the Auerbach plexus proved to be different of those present on the cortical cholinergic interneurons and on the sympathetic plexus of the guinea-pig atrium.