Autonomic and Autacoid Pharmacology最新文献

1 This study examines the cellular localization of α₁-adrenoceptors and demonstrates that binding to intracellular receptive binding sites in native smooth muscle cells may influence the pharmacological profile of agonists or antagonists. The example tissue studied was rat basilar artery.

2 An α₁-adrenoceptor antagonist and fluorescent analogue of prazosin, BODIPY-FL prazosin (QAPB) allowed visualization, with high resolution, of both plasma membrane and cytosolic binding sites on live native cells, as previously shown in cells harbouring recombinant receptors. QAPB-associated fluorescence binding was both time- and concentration- dependent in rat basilar smooth muscle cells and affinity for α₁-adrenoceptors was calculated from specific binding curves as 1.1 nM.

3 Concentration-dependent binding of QAPB detected in smooth muscle cells dissociated from rat basilar arteries was composed of diffuse and clustered fluorescence. Visually the diffuse component of fluorescence was the more evident up to a concentration of 5 nM QAPB. Confocal visualization of an optical section through the cell showed that the clustered component was located mainly intracellularly. In rat basilar artery smooth muscle cells the intracellular binding sites were located in close proximity to the nuclear membrane.

4 3D models of QAPB-associated fluorescence demonstrate that a high proportion of effective binding sites are intracellular, showing not only that a high proportion of receptors are located inside the cell but also that in this location they can bind ligands. This has implications for. pharmacological analysis in relation to the consequences of intracellular binding per se and for differential effects upon the pharmacology of particular ligands according to whether they can enter the cell.

1 Angiotensin and bradykinin facilitate the release of noradrenaline from sympathetic nerve terminals and cause positive inotropy in rat isolated atria and ventricles. The effect of bradykinin was enhanced by the ACE inhibitor, ramiprilat.

2 The facilitated release of noradrenaline in rat ventricle by bradykinin was blocked by the β₂-receptor antagonist HOE-140. This response is also reduced by removing the endocardium, suggesting the release of a mediator from the endocardium.

3 The facilitated noradrenaline release by angiotensin II and bradykinin was blocked by the angiotensin receptor antagonist saralasin to the same extent. In contrast, losartan caused only minor blockade in a range of vascular and cardiac tissues. This suggests that angiotensin and bradykinin exert these responses by interacting with a prejunctional receptor different from the established AT₁ subtype.

4 These results suggest that bradykinin mediates facilitation of noradrenaline release via the local release of angiotensin onto an atypical AT₁ receptor.

1 The aim of this study was to investigate the effect of sertraline, a selective serotonin re-uptake inhibitor, on contractile responses to noradrenaline (NA), KCI, serotonin (5-HT) and electrical field stimulation of rat isolated vas deferens.

2 Pre-treatment with 10^-4 M sertraline showed inhibitory effects on responses to NA, KCI, 5-HT and electrical field stimulation, while pre-treatment with 10 ^-6 and 10^-5 M sertraline caused potentiation of responses to NA (10^-7 and 10^-6 M).

3 A voltage-dependent calcium channel activator, Bay K 8644, restored the inhibited responses when sertraline was washed out of the organ bath, although restoration could not be seen when sertraline was not removed.

4 The inhibition of the contractile responses by sertraline pre-treatment may be via a mechanism through calcium channels which is additional to the selective serotonin re-uptake inhibitory effect of sertraline.

1 In order to more fully understand the role of the α₂-adrenoceptor in brain function, a combination of in vitro and in vivo techniques were utilized including radioligand binding, autoradiography, brain microdialysis and antisense oligonucleotides.

2 Binding studies showed the tritiated form of the selective α₂-adrenoceptor antagonist, RX821002 (methoxy-idazoxan) labelled an apparent single population of sites in rat brain membranes with high affinity (1 nM), for which prazosin had low affinity (1107 nM). Similar studies in rabbit brain membranes found that prazosin and oxymetazoline were able to displace. [³H]-RX821002 in a biphasic manner indicating the presence of subtypes of α₂-adrenoceptors.

3 Receptor autoradiography revealed a distribution of [³H]-RX821002 binding in rat brain consistent with the labelling of all %aL₂-adrenoceptor subtypes, namely α_2A/D-, α_2B and α_2C.

4 In rat, in vivo brain dialysis experiments demonstrated peripherally administered RX821002 elevated basal noradrenaline in frontal cortex and also, although to a lesser extent, in ventral hippocampus. RX821002 was also able to elevate extracellular dopamine in the striatum.

5 A 7-day i.c.v. infusion of an antisense oligonucleotide targeting the α_2A/D-adrenoceptor, resulted in a significant reduction in the autoradiographic density of [³H]-RX821002 binding in specific brain areas, notably the lateral septal nuclei and anterior hypothalamic area.

6 Several years of research by our group has extended our knowledge of the pharmacology and function of the α₂-adrenoceptor and has provided evidence of the roles of this receptor in the control of monoamine turnover. The successful use of antisense technology to knockdown expression of the α_2A/D subtype provides future opportunities to explore the physiology of this receptor subtype.

• A set-up for computerized acquisition and evaluation of haemodynamic data was constructed. Blood flow (BF) in the superior mesenteric and femoral artery of urethane-anaesthetized rats was measured with the ultrasonic transit time shift technique. The signals for arteial blood pressure and BF were fed into a personal computer via an analogue-digital converter. Mean arterial blood pressure, heart rate and vascular conductance (CV) were calculated on-line. For subsequent analysis of the data, algorithms were programmed to filter the data, and to determine average and peak values for each parameter.

• Systemic hypertension induced by phenylephrine (3–300 nmol kg⁻¹), angiotensin II (0.1–3.0 nmol kg⁻¹) and arginine vasopressin (0.03–1.0 nmol kg⁻¹) was accompanied by constriction of the mesenteric artery. In contrast, the femoral artery responded to phenylephrine with constriction, to angiotensin II with dilatation and to arginine vasopressin with dilation followed by constriction. The haemodynamic effects of endothelin-1 (0.03–3.0 nmol kg⁻¹) were generally biphasic, the initial hypotension being associated with dilatation, and the delayed hypertension being accompanied by constriction of both the mesenteric and femoral arterial bed.

• Terbutaline (3–1.0 nmol kg⁻¹) and calcitonin gene-related peptide (0.03–1 nmol kg⁻¹) caused systemic hypotension along with mesenteric and femoral vasodilatation.

• Telmisartan (1 mg kg⁻¹), an angiotensin AT₁ receptor antagonist, dilated the mesenteric artery, but had no effect on femoral VC. In contrast, the α₁-adrenoceptor antagonist prazosin (0.1 mg kg⁻¹), dilated the femoral artery without altering mesenteric VC. Similarly, the β-adrenoceptor antagonist propranolol (1 mg kg⁻¹) had no effect on mesenteric VC, but constricted the femoral arterial bed.

• These data demonstrate that the haemodynamic effects of exogenously administered drugs can widely differ between the mesenteric and femoral arterial beds of urethane-anaesthetized rats. Furthermore, vascular tone of these two arterial beds in maintained by different vasoconstrictor systems. While the femoral artery is mainly under adrenergic control, the renin-angiotensin axis is predominant in the mesenteric arterial bed. In addition, this study also demonstrates that computerized analysis enables quick and accurate estima