Adrenergic signaling molecular complexes. / Complejos moleculares de la señalización adrenérgica.

Adrenaline and noradrenaline bind to membrane receptors of the superfamily of G protein-coupled receptors (GPCR) in target cells, where they modulate physiological responses such as metabolism, vasoconstriction, vasodilation and proliferation. Alteration in their function is associated with conditions such as hypertension, benign prostatic hyperplasia and cardiac hypertrophy. In response to adrenaline, receptors form signaling complexes, which enables adrenergic action to be specific, rapid and efficient. These signaling complexes or signalosomes are composed of kinases, phosphatases, and adapter and scaffold proteins, which together modulate the receptor function. Manipulation of each protein-protein interaction of the adrenergic signaling complex emerges as a promising therapeutic strategy for the design of drugs that modulate adrenergic action and help to define its pathophysiological significance. An important biological model to perform these investigations is the heart, since it expresses all adrenergic receptors; to date, several heart signalosomes have been described. Mass spectrometry (proteomics), genetic manipulation and biochemical assays, such as two-hybrid and co-immunoprecipitation assays, are tools that are used in these studies.

La adrenalina y la noradrenalina se unen a receptores membranales de la superfamilia de receptores acoplados a proteínas G (GPCR) en las células blanco, donde modulan respuestas fisiológicas tales como el metabolismo, vasoconstricción, vasodilatación y proliferación. La alteración en su función está asociada con hipertensión, hiperplasia prostática benigna e hipertrofia cardiaca. En respuesta a la adrenalina, los receptores forman complejos de señalización, lo que permite que la acción adrenérgica sea específica, rápida y eficiente. Estos complejos de señalización o signalosomas están integrados por cinasas, fosfatasas, proteínas adaptadoras y de andamio, que en conjunto modulan la función del receptor. La manipulación de cada interacción proteína-proteína del complejo de señalización adrenérgico emerge como una estrategia terapéutica prometedora para el diseño de fármacos que modulen la acción adrenérgica y ayuden a definir su significado fisiopatológico. Un modelo biológico importante para realizar estos estudios es el corazón, ya que expresa todos los receptores adrenérgicos; en la actualidad se han descrito varios signalosomas cardiacos. La espectrometría de masas (proteómica), manipulación genética y ensayos bioquímicos como el doble híbrido o la coinmunoprecipitación son herramientas que se emplean en estos estudios.

Neural control of the circulation.

The purpose of this brief review is to highlight key concepts about the neural control of the circulation that graduate and medical students should be expected to incorporate into their general knowledge of human physiology. The focus is largely on the sympathetic nerves, which have a dominant role in cardiovascular control due to their effects to increase cardiac rate and contractility, cause constriction of arteries and veins, cause release of adrenal catecholamines, and activate the renin-angiotensin-aldosterone system. These effects, as well as the control of sympathetic outflow by the vasomotor center in the medulla and the importance of sensory feedback in the form of peripheral reflexes, especially the baroreflexes, are discussed in the context of cardiovascular regulation.

Distinct role of adrenoceptor subtypes in cardiac adaptation to chronic pressure overload.

1. With the generation of gene knockout (KO) or transgenic overexpression (TG) mouse models targeting adrenoceptors (AR), recent studies in vivo have investigated the role of AR subtypes in pressure overload-induced left ventricular (LV) hypertrophy and remodelling. 2. Although subjecting alpha(1B)-KO mice to transverse aortic constriction (TAC) did not reveal significant phenotype differences compared with controls, mice deficient in both alpha(1A)- and alpha(1B)-AR responded to TAC with poor survival, increased cardiomyocyte apoptosis, more severe fibrosis and dysfunction, but a similar degree of LV hypertrophy, compared with wild-type littermates. Following TAC, alpha(1B)-TG mice developed more severe hypertrophy, interstitial fibrosis and LV dysfunction. In contrast, overexpression of alpha(1A)-AR preserved cardiac function and reduced death from heart failure without affecting the degree of LV hypertrophy. Thus, alpha(1A)- and alpha(1B)-adrenoceptor signalling impacts differently on myocardial adaptation to pressure overload. 3. The absence of both beta(1)- and beta(2)-AR significantly suppressed pressure overload-evoked hypertrophy, fibrosis and expression of inflammatory or fibrogenic genes. Conversely, studies on beta(2)-TG mice with TAC revealed adverse consequences, including accelerated development of heart failure, poor survival and more severe interstitial fibrosis, but a comparable degree of hypertrophy compared with wild-type littermates. 4. Collectively, these findings suggest that the effect of ARs on pressure overload-induced myocardial adaptation is subtype specific. Whereas activation of alpha(1B)-AR or beta(2)-AR contributes to maladaptation and the onset of heart failure, activation of alpha(1A)-AR or inactivation of beta(2)-AR is beneficial in the setting of chronic pressure overload.

Presynaptic metabotropic receptors for acetylcholine and adrenaline/noradrenaline.

Presynaptic metabotropic receptors for acetylcholine and adrenaline/noradrenaline were first described more than three decades ago. Molecular cloning has resulted in the identification of five G protein-coupled muscarinic receptors (M(1) - M(5)) which mediate the biological effects of acetylcholine. Nine adrenoceptors (alpha(1ABD),alpha(2ABC),beta(123)) transmit adrenaline/noradrenaline signals between cells. The lack of sufficiently subtype-selective ligands has prevented identification of the physiological role and therapeutic potential of these receptor subtypes for a long time. Recently, mouse lines with targeted deletions for all muscarinic and adrenoceptor genes have been generated. This review summarizes the results from these gene-targeting studies with particular emphasis on presynaptic auto- and heteroreceptor functions of muscarinic and adrenergic receptors. Specific knowledge about the function of receptor subtypes will enhance our understanding of the physiological role of the cholinergic and adrenergic nervous system and open new avenues for subtype-selective therapeutic strategies.

Probenecid Inhibits α-Adrenergic Receptor-Mediated Vasoconstriction in the Human Leg Vasculature.

Coordination of vascular smooth muscle cell tone in resistance arteries plays an essential role in the regulation of peripheral resistance and overall blood pressure. Recent observations in animals have provided evidence for a coupling between adrenoceptors and Panx1 (pannexin-1) channels in the regulation of sympathetic nervous control of peripheral vascular resistance and blood pressure; however, evidence for a functional coupling in humans is lacking. We determined Panx1 expression and effects of treatment with the pharmacological Panx1 channel inhibitor probenecid on the vasoconstrictor response to α1- and α2-adrenergic receptor stimulation in the human forearm and leg vasculature of young healthy male subjects (23±3 years). By use of immunolabeling and confocal microscopy, Panx1 channels were found to be expressed in vascular smooth muscle cells of arterioles in human leg skeletal muscle. Probenecid treatment increased (P<0.05) leg vascular conductance at baseline by ≈15% and attenuated (P<0.05) the leg vasoconstrictor response to arterial infusion of tyramine (α1- and α2-adrenergic receptor stimulation) by ≈15%, whereas the response to the α1-agonist phenylephrine was unchanged. Inhibition of α1-adrenoceptors prevented the probenecid-induced increase in baseline leg vascular conductance, but did not alter the effect of probenecid on the vascular response to tyramine. No differences with probenecid treatment were detected in the forearm. These observations provide the first line of evidence in humans for a functional role of Panx1 channels in setting resting tone via α1-adrenoceptors and in the constrictive effect of noradrenaline via α2-adrenoceptors, thereby contributing to the regulation of peripheral vascular resistance and blood pressure in humans.

Changes in adrenoreceptors in the prefrontal cortex of subjects with dementia: evidence of compensatory changes.

In Alzheimer's disease (AD) there is a significant loss of locus coeruleus (LC) noradrenergic neurons. However, recent work has shown the surviving noradrenergic neurons to display many compensatory changes, including axonal sprouting to the hippocampus. The prefrontal cortex (PFC) is a forebrain region that is affected in dementia, and receives innervation from the LC noradrenergic neurons. Reduced PFC function can reduce cognition and disrupt behavior. Because the PFC is an important area in AD, we determined if noradrenergic innervation from the LC noradrenergic neurons is maintained and if adrenoreceptors are altered postsynaptically. Presynaptic PFC alpha2-adrenoreceptor (AR) binding site density, as determined by 3H-RX821002, suggests that axons from surviving noradrenergic neurons in the LC are sprouting to the PFC of subjects with dementia. Changes in postsynaptic alpha1-AR in the PFC of subjects with dementia indicate normal to elevated levels of binding sites. Expression of alpha1-AR subtypes (alpha1A- and alpha1D-AR) and alpha2C-AR subtype mRNA in the PFC of subjects with dementia is similar to what was observed in the hippocampus with one exception, the expression of alpha1A-AR mRNA. The expression of the alpha1A-AR mRNA subtype is significantly reduced in specific layers of the PFC in subjects with dementia. The loss of alpha1A-, alpha1D- and alpha2C-AR mRNA subtype expression in the PFC may be attributed to neuronal loss observed in dementia. These changes in postsynaptic AR would suggest a reduced function of the PFC. Consequence of this reduced function of the PFC in dementia is still unknown but it may affect memory and behavior.

GPCRsclass: a web tool for the classification of amine type of G-protein-coupled receptors.

The receptors of amine subfamily are specifically major drug targets for therapy of nervous disorders and psychiatric diseases. The recognition of novel amine type of receptors and their cognate ligands is of paramount interest for pharmaceutical companies. In the past, Chou and co-workers have shown that different types of amine receptors are correlated with their amino acid composition and are predictable on its basis with considerable accuracy [Elrod and Chou (2002) Protein Eng., 15, 713-715]. This motivated us to develop a better method for the recognition of novel amine receptors and for their further classification. The method was developed on the basis of amino acid composition and dipeptide composition of proteins using support vector machine. The method was trained and tested on 167 proteins of amine subfamily of G-protein-coupled receptors (GPCRs). The method discriminated amine subfamily of GPCRs from globular proteins with Matthew's correlation coefficient of 0.98 and 0.99 using amino acid composition and dipeptide composition, respectively. In classifying different types of amine receptors using amino acid composition and dipeptide composition, the method achieved an accuracy of 89.8 and 96.4%, respectively. The performance of the method was evaluated using 5-fold cross-validation. The dipeptide composition based method predicted 67.6% of protein sequences with an accuracy of 100% with a reliability index > or =5. A web server GPCRsclass has been developed for predicting amine-binding receptors from its amino acid sequence [http://www.imtech.res.in/raghava/gpcrsclass/ and http://bioinformatics.uams.edu/raghava/gpersclass/ (mirror site)].

Quantitative mRNA analysis of adrenergic receptor subtypes in the intestines of healthy dairy cows and dairy cows with cecal dilatation-dislocation.

OBJECTIVE: To investigate the distribution of mRNA coding for 9 adrenoceptor subtypes in the intestines of healthy dairy cows and cows with cecal dilatationdislocation (CDD). SAMPLE POPULATION: Full-thickness specimens of the intestinal wall were obtained from the ileum, cecum, proximal loop of the ascending colon (PLAC), and external loop of the spiral colon (ELSC) of 15 cows with CDD (group 1) and 15 healthy (control) cows (group 2, specimens collected during laparotomy; group 3, specimens collected after slaughter). PROCEDURES: Concentrations of mRNA for 9 adrenoceptor subtypes (alpha(1A), alpha(1B), alpha(1D), alpha(2AD), alpha(2B), alpha(2C), beta(1), beta(2), and beta(3)) were measured by quantitative real-time reverse transcriptase-PCR assay. Results were expressed relative to mRNA expression of a housekeeping gene. RESULTS: Expression of mRNA for alpha(1B)-, alpha(2AD)-, alpha(2B)-, beta(1)-, and beta(2)-adrenoceptors was significantly lower in cows with CDD than in control cows. In the ileum, these receptors all had lower mRNA expression in cows with CDD than in control cows. The same effect was detected in the ELSC for mRNA for alpha(2AD)-, alpha(2B)-, beta(1)-, and beta(2)-adrenoceptors, and in the cecum and PLAC for alpha(2B)- and beta(2)-adrenoceptors. Groups did not differ significantly for alpha(1A)-adrenoceptors. The mRNA expression for alpha(1D)-, alpha(2C)-, and beta(3)-adrenoceptors was extremely low in all groups. CONCLUSIONS AND CLINICAL RELEVANCE: Differences in expression of mRNA coding for adrenoceptors, most pronounced in the ileum and spiral colon, between cows with CDD and control cows support the hypothesis of an implication of adrenergic mechanisms in the pathogenesis of CDD in dairy cows.

Molecular characterization of alpha 1- and alpha 2-adrenoceptors.

Three 'alpha 1-adrenoceptors' and three 'alpha 2-adrenoceptors' have now been cloned. How closely do these receptors match the native receptors that have been identified pharmacologically? What are the properties of these receptors, and how do they relate to other members of the cationic amine receptor family? Kevin Lynch and his colleagues discuss these questions in this review.

Adrenergic receptor characterization of CA1 hippocampal neurons using real time single cell RT-PCR.

The CA1 region of the rat hippocampus exhibits both alpha and beta adrenergic receptor (AR) responses, however, the specific AR subtypes involved and the neuronal expression patterns for these receptors are not well understood. We have employed single cell real time RT-PCR in conjunction with cell-specific immunohistochemical markers to determine the AR expression patterns for hippocampal neurons located in CA1, a region often implicated in learning and memory processes. Cytoplasmic samples were taken from 55 individual cells located in stratum oriens, pyramidale, or radiatum and reverse transcribed. All successfully amplified pyramidal neuron samples (n = 17) expressed mRNA for the beta2AR, with four cells additionally expressing mRNA for the beta1AR subtype. Positive interneurons from stratum oriens (n = 10) and stratum radiatum (n = 8) expressed mRNA for the alpha1A and/or alpha(1B)AR (n = 9/18) only when coexpressing transcripts for somatostatin. Interneurons containing neuropeptide Y or cholecystokinin (n = 9/18) were not positive for any of the nine AR subtypes, suggesting that CA1 interneuron AR expression is limited to a subset of somatostatin-positive cells. These findings suggest that only a select number of AR subtypes are transcriptionally expressed in CA1 and that these receptors are selective to specific neuronal cell types.

[Influence of adrenoreceptors on functions of the body]. / Adrenoreceptoriu poveikis organizmo funkcijoms.

The remarkably diverse effects of the catecholamines and similar sympathomimetic agents are directly related to an understanding of the classification and different types of adrenoreceptors. Characteristics and physiological regulatory mechanisms of the receptor result in variable response of organ systems to catecholamines stimulation. Different adrenoreceptors regulate distinct physiological processes by controlling the synthesis or release of a variety of second messengers. The goal of this review was to turn one's attention to the below mentioned aspects. There are three known subtypes of each alpha1-, alpha2- and beta-adrenoreceptor types. Structure of the adrenoreceptors, which belong to subtypes of the same receptor type, is similar and structure of the adrenoreceptors of the separate types is very different. Genetic peculiarities of the receptors may influence liability to some diseases. Acting on the adrenoreceptors may change function of many organs and may serve for the treatment of cardiovascular, respiratory tract diseases and allergic reactions. Selective acting on the adrenoreceptors of the separate subtypes may have the different effect on the organs. Great consideration is given for that property in the development of new drugs: substitution by different chemical radicals leads to increasing selectivity for the separate subtypes of the adrenoreceptors. The prolonged use of the adrenomimetics may lead to refractoriness.

Characterization of alpha- and beta-adrenergic agonist stimulation of adenylate cyclase activity in human epidermal keratinocytes in vitro.

Adrenergic receptors are responsible for selective recognition and binding of catecholamines and may in turn have an effect on epidermal cell growth and maturation via adenosine-3',5'-monophosphate (cAMP). Using endogenous catecholamines and drugs specific for alpha- and beta-receptor subtypes, we have characterized the adrenergic receptor coupled to adenylate cyclase in cultured human epidermal keratinocytes. The relative potency order of stimulation of adenylate cyclase was: isoproterenol greater than epinephrine much greater than norepinephrine. The predominant adrenergic receptor is of the beta 2-subtype, as also confirmed by use of the selective antagonists propranolol, butoxamine, and atenolol. No evidence of alpha-adrenergic receptor mediation of adenylate cyclase was noted with the alpha 2-specific agonist, clonidine. Phenylephrine, the alpha 1-specific agonist, affected cAMP formation but this response could not be totally inhibited with prazosin, suggesting an unknown mechanism of action. Human keratinocytes retained the same beta-adrenergic receptor potency order properties throughout growth and maturation.

How should one study brain adrenergic receptors in aging?

Contradictory findings and lack of definitive information regarding adrenergic receptors in aging results in part from problems related to the methodology that has been used to study this question. Limitations of available techniques and new biochemical, molecular biological, and physiological methods that may prove particularly helpful for future studies are discussed.

Astrocytic adrenoceptors: a major drug target in neurological and psychiatric disorders?

Considerable attention has recently been paid to astrocyte functions, which are briefly summarized. A large amount of data is available about adrenoceptor expression and function in astrocytes, some of it dating back to the 1970's and some of it very recent. This material is reviewed in the present paper. The brain is innervated by noradrenergic fibers extending from locus coeruleus in the brain stem, which in turn is connected to a network of adrenergic and noradrenergic nuclei in the medulla and pons, contributing to the control of (nor)adrenergic, serotonergic, dopaminergic and cholinergic function, both in the central nervous system (CNS) and in the periphery. In the CNS astrocytes constitute a major target for noradrenergic innervation, which regulates morphological plasticity, energy metabolism, membrane transport, gap junction permeability and immunological responses in these cells. Noradrenergic effects on astrocytes are essential during consolidation of episodic, long-term memory, which is reinforced by beta-adrenergic activation. Glycogenolysis and synthesis of glutamate and glutamine from glucose, both of which are metabolic processes restricted to astrocytes, occur at several time-specific stages during the consolidation. Astrocytic abnormalities are almost certainly important in the pathogenesis of multiple sclerosis and in all probability contribute essentially to inflammation and malfunction in Alzheimer's disease and to mood disturbances in affective disorders. Noradrenergic function in astrocytes is severely disturbed by chronic exposure to cocaine, which also changes astrocyte morphology. Development of drugs modifying noradrenergic receptor activity and/or down-stream signaling is advocated for treatment of several neurological/psychiatric disorders and for neuroprotection. Astrocytic preparations are suggested for study of mechanism(s) of action of antidepressant drugs and pathophysiology of mood disorders.

Regulatory effects of reboxetine treatment alone, or following paroxetine treatment, on brain noradrenergic and serotonergic systems.

When patients do not respond to an initial antidepressant, one clinical approach is to switch to an agent in a different pharmacological class. However, few studies have examined the neurochemical consequences of this practice. To study this, we examined changes in binding sites in rat brain for norepinephrine (NET) and serotonin transporters (SERT), alpha1, alpha2, and beta1 adrenergic receptors after chronic administration of paroxetine (PRX), reboxetine (RBX), or PRX followed by RBX. We also examined the effects of these treatments on mRNA expression for tyrosine hydroxylase (TH). RBX treatment for 3 weeks reduced NET binding significantly, by approximately 40% in terminal field areas, and 6 weeks of RBX reduced it even more, by approximately 60%. RBX treatment for 3 and 6 weeks reduced beta1 adrenergic receptor-binding sites equally, by 50-60%. At no time did RBX treatment reduce SERT-binding sites. PRX treatment had no effect on beta1 adrenergic or NET-binding sites, but reduced SERT-binding sites by 75-80%. Neither treatment altered mRNA for TH, alpha1, or alpha2 adrenergic receptor-binding sites. When 3 weeks of RBX treatment followed 3 weeks of PRX treatment, NET-binding sites were reduced to the same extent as measured after 6 weeks of RBX treatment alone, indicating that PRX pretreatment may have 'primed' the subsequent regulatory effect of RBX on the NET. Thus, pretreatment of rats with PRX actually enhanced at least one regulatory effect of RBX treatment on the noradrenergic system, and did not interfere with any other pharmacological effect caused by RBX treatment.

The subdivision of beta-adrenoceptors in the cardiovascular system of the rat.

1 The antagonism by the beta-adrenoceptor blocking drugs, propranolol (non-selective) and practolol (beta-selective), of the cardiovascular actions of isoprenaline has been investigated in the rat. 2 All doses of practolol (0.1, 1 and 3 mg/kg) blocked the cardio-accelerator action of isoprenaline but only the largest dose blocked the vasodilator effect. 3 All doses of propranolol (0.01, 0.03 and 0.1 mg/kg) blocked the vasodilator effect of isoprenaline but only the largest dose diminished the tachycardia. 4 It is concluded that in the rat, as in other species, beta-adrenoceptors may be subdivided into beta 1 (cardiac) and beta 2 (peripheral vascular) types.

The existence of a new subtype of alpha-adrenoceptor on the rat anococcygeus is revealed by SGD 101/75 and phenoxybenzamine.

1 Noradrenaline and Sgd 101/75 (4(2-imidazoline-amino)-2-methylindazol-chlorhydrate) acted as full agonists in contracting the rat anococcygeus. 2 Very low concentrations of phenoxybenzamine (0.3 nM for 2-30 min) reduced preferentially the effects of Sgd 101/75. Preparations made insensitive to Sgd 101/75 by phenoxybenzamine still contracted to noradrenaline, this contraction occurring in the presence of high concentrations (400 microM) of Sgd 101/75. 3 It is concluded therefore that the alpha 1-adrenoceptor in this preparation is not a homogeneous entity and must be subdivided into at least two subtypes.

Distribution and types of adrenoceptors in the guinea-pig ileum: the action of alpha- and beta-adrenoceptor agonists.

1 Segments of guinea-pig ileum and the myenteric plexus-longitudinal smooth muscle preparation were used for a study of the actions of adrenaline, noradrenaline, isoprenaline, ephedrine and phenylephrine on the responses of coaxially stimulated ileum at different distances from the ileocaecal valve.2 The responses of the ileum to electrical stimulation were suppressed by adrenaline, nonadrenaline and ephedrine, while phenylephrine and isoprenaline inhibited them only partially.3 The twitch inhibition elicited by these adrenoceptor agonists was the same at all distances from the ileocaecal valve. There was no significant difference between their cumulative and non-cumulative concentration-response curves.4 Smooth muscle relaxation was induced only by isoprenaline and contraction only by phenylephrine at all distances from the ileocaecal junction. Adrenaline and noradrenaline evoked smooth muscle contraction in the terminal (0 to 20 cm), a concentration-dependent, biphasic response in the intermediate part (21 to 50 cm) and a relaxation in the proximal ileum (> 50 cm from the ilecocaecal valve). Ephedrine did not change significantly the smooth muscle tension in the terminal and the intermediate segments and induced smooth muscle relaxation in the proximal ones.5 Ouabain and a potassium-free solution did not appear to influence the prejunctional action of noradrenaline nor the amplitude of smooth muscle relaxation in the proximal ileum, whereas the concentration-contractor response curves were significantly depressed and shifted to the right by ouabain and in a potassium-free solution.6 The brief initial (phasic) contraction induced by acetylcholine was not influenced during the sustained increase or decrease in tension induced by catecholamines. On the contrary, the stimulatory catecholamine actions disappeared or were changed to smooth muscle relaxation by acetylcholine pretreatment. Potassium chloride pretreatment did not change the character of the adrenoceptor agonist action of the agonists studied.7 Since there is a similar prejunctional action at all distances from the ileocaecal valve and a different postjunctional effect of the adrenoceptor agonists at different distances from the ileocaecal junction, it could be suggested that in the guinea-pig ileum there are at least two alpha-adrenoceptors (inhibitory prejunctional-alpha(2), stimulatory postjunctional-alpha(1)), an inhibitory postjunctional beta-adrenoceptor and an as yet uncharacterized inhibitory postjunctional receptor.8 Based on the specific postjunctional action of phenylephrine and the prejunctional action of ephedrine in the guinea-pig ileum, these drugs could be used with success as ;specific' alpha(1)- and alpha(2)-adrenoceptor stimulants.

Classification of beta-adrenoceptors in isolated bronchus of the pig.

1 (+/-)-Isoprenaline (Iso), (-)-adrenaline (Ad), (-)-noradrenaline (NA), the beta 2-selective adrenoceptor agonist (+/-)-fenoterol (Fen) and the beta 1-selective adrenoceptor agonist (+/-)-RO363 caused concentration-dependent relaxation of preparations of pig bronchus pre-contracted with carbachol 40-ng/ml (0.22 microM). Iso, Ad, NA and Fen caused complete relaxation of carbachol-induced tone, but RO363 caused relaxation equivalent to only 59% of the maximal response to Iso. 2 When relaxation responses to these amines were plotted as a % of their maximal effects, comparison of EC50 values showed that the order of potency was RO363 greater than Iso greater than NA greater than Fen greater than Ad (14.4:4.6:1:0.4:0.3). 3 pA2 values determined for the beta-adrenoceptor antagonists propranolol (non-selective) and atenolol (beta 1-selective), or the partial agonist salbutamol (beta 2-selective) using Iso as agonist were 8.3, 7.3 and 4.4 respectively. The pA2 value for atenolol using RO363 as the agonist was 7.6. 4 These results indicate that porcine bronchus contains a homogeneous population of beta 1-adrenoceptors.

Receptors involved in nerve-mediated vasoconstriction in small arteries of the rat hepatic mesentery.

1. We have investigated the neurotransmitters and receptor subtypes involved in nerve-mediated vasoconstriction in small arteries of the rat hepatic mesentery. 2. A dense sympathetic innervation was demonstrated using catecholamine histochemistry and antibodies against the synaptic vesicle protein synaptophysin. 3. Reverse transcription-polymerase chain reaction (RT-PCR) demonstrated very strong expression of the alpha1A-adrenergic, neuropeptide Y (NPY) Y1, P2X1- and P2X4-purinergic receptors, moderate expression of the alpha2B-adrenergic receptor and the purinergic P2X5- and P2X7-receptors and weak expression of the alpha1B-, alpha1D-, alpha2A- and alpha2C-adrenergic receptors and the P2X2- and P2X3-purinergic receptors. NPY2 and P2X6 receptor expression was absent. 4. Electrical field stimulation (10 Hz, 10 s) produced contractions which were abolished by tetrodotoxin (10(-6) M) and/or guanethidine (GE, 5 x 10(-6) M) and a combination of benextramine (10(-5) M) and alpha,beta-methylene ATP, (alpha,beta-mATP, 3 x 10(-6) M) or PPADS (10(-5) M). Selective alpah1-adrenergic receptor antagonists showed the potency order of prazosin > WB-4101 > 5-methyl-urapidil > BMY 7378. Yohimbine (10(-8) M, 10(-7) M), alpha,beta-mATP (3 x 10(-6) M) and PPADS (10(-5) M) each enhanced the response to nerve stimulation. 5. Some experiments demonstrated a slow neurogenic contraction which was abolished by GE or the selective NPY1 receptor antagonist 1229U91 (6 x 10(-7) M). 6. We conclude that nerve-mediated vasoconstriction results from the activation of postsynaptic alpha,beta-adrenergic and P2X-purinergic receptors and under some conditions, NPY1 receptors. Neurotransmitter release is modulated by presynaptic alpha2-adrenergic receptors and possibly also P2X-purinoceptors. The major postsynaptic subtypes involved were well predicted by mRNA expression as measured by RT-PCR, suggesting that this technique may be a useful adjunct to studies aimed at identifying functional receptor subtypes.