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Notes: 1. (—)[125I]-Cyanopindolol (CYP) binding to non-β-adrenoceptor sites in dog kidney was characterized in homogenate preparations and their distribution in sections determined using autoradiography.2. In homogenate studies, (—)[125I]-CYP bound to a single population of non-interacting sites (Bmax= 5.45, s.e.m. = 1.00 fmol/mg wet weight; nH = 0.99, s.e.m. = 0.01) with high affinity (KD= 3.84, s.e.m. = 0.76 nmol/l, n= 4).3. In competition studies, compounds selective for α- and β-adrenoceptors, muscarinic cholinoceptors and receptors for 5-HT, histamine and benzodiazepines, calcium channel antagonists, catecholamine uptake inhibitors, MAO inhibitors and adrenergic neurone blockers were ineffective at concentrations of 10 μmol/l.4. Compounds selective for dopamine D1-receptors (fluphenazine, SCH 23390 and SK & F 82526) and D2-receptors (pimozide, domperidone, spiperone, haloperidol, sulpiride, cis- and trans-flupenthixol) competed with similar affinities (5–25 μmol/l) for (—)[125I]-CYP binding.5. In autoradiographic studies, (—) [125I]-CYP binding to non-β-adrenoceptor sites was localized over glomeruli, juxtaglomerular apparatus, distal tubules, blood vessels and medullary rays and tubules.6. It is concluded that in dog kidney, (—)[125I]-CYP binds to a site closely associated with dopamine receptors.

Notes: SUMMARY1. Autoradiographic techniques have been used to examine the location of β-adrenoceptors in the heart and β-adrenoceptors, substance P receptors and muscarinic cholinoceptors in blood vessels.2. Both β1-adrenoceptors and β2-adrenoceptors were present in guinea-pig and human heart, on the myocardium and associated with the cardiac nerves and blood vessels.3. Nerves on the vasculature and vascular smooth muscle contained β-adrenoceptors and muscarinic cholinoceptors.4. Receptors for substance P and β-adrenoceptors, but not muscarinic cholinoceptors were present on endothelial cells.

Notes: Abstract  Regulation of β-adrenoceptor (β-ar) subtypes and transregulation of muscarinic cholinoceptors (mAchr) was examined in regions of rat heart after chronic infusion of (–)-isoprenaline (450 μg/kg per hour) for 14 days. Following (–)-isoprenaline infusion systolic blood pressure was reduced for 10 days but then gradually returned to control levels, whereas heart rate was increased for 7 days before declining to a level significantly above control. Heart weight to body weight ratio was increased in (–)-isoprenaline treated rats. β-ar subtype densities were measured by quantitative autoradiography with [125I]-cyanopindolol (CYP) in sinoatrial node (SA), atrioventricular node (AV), bundle of His (BH), left (LB) and right (RB) bundle branches, interventricular (IVS) and interatrial (IAS) septa, right atria (RA), apex (AX) and mitral valve (MV). β1-ars were reduced by 59.1–74.2% in the AV conducting regions, 53.4% in the SA node and 43.3–53.4% in myocardial areas. β2-ars were markedly reduced in myocardial regions (93.2–98.5%) and in pacemaker and conducting regions (87.7–97.8%). No changes in mAchr densities measured using [3H]-N-methyl scopolamine (NMS) occurred in the AV node, BH, LB, RB, IVS and IAS following (–)-isoprenaline infusion. Densities of β1- and β2-ars and mAchrs were also measured in ventricular homogenates from control and (–)-isoprenaline treated animals. β-ar levels were significantly reduced (P〈0.05) in treated animals and the ratio of β1- to β2-ars increased after treatment. mAchr density in ventricular homogenates measured using either [3H]-NMS or [3H]-quinuclidinyl [phenyl-4-3H]benzilate (QNB) was unchanged. Homogenates of left and right ventricle also showed no change using [3H]-NMS. Organ bath studies were used to investigate the effect of (–)-isoprenaline infusion on negative inotropic and chronotropic effects of the non-selective muscarinic receptor agonist bethanechol in left and right atria, respectively. Lower concentrations of bethanechol (3×10-10 to 10-6 M) produced a negative inotropic response in isolated electrically driven left atria from (–)-isoprenaline treated rats, but not from control rats, with the slope of the curves being significantly different between groups (ANCOVA, P=0.037). At concentrations of bethanechol from 10-6 to 3×10-4 M the negative inotropic response was not changed between (–)-isoprenaline treated and control animals. Bethanechol also produced a negative chronotropic response at lower concentrations (10-10 to 10-6 M) in (–)-isoprenaline treated rats, but not in controls. A second, steeper phase of the negative chronotropic response occurred at concentrations of bethanechol greater than 10-6 M and was also seen in control rats. Expression of M2 (cardiac) mAchrs (m2Achr) in left and right ventricular tissues measured using a quantitative non-competitive polymerase chain reaction (PCR) assay showed a significant (P=0.001) 28.5% increase in expression in left ventricle and a significant (P=0.003) 21.5% decrease in expression in right ventricle after (–)-isoprenaline treatment, compared to controls. There was no significant difference in total ventricular m2Achr expression between the two groups of rats. The results suggest that chronic β-ar stimulation down-regulates both β1- and β2-ars, and appears to differentially transregulate m2Achr expression, but not mAchr protein. Following (–)-isoprenaline infusion, muscarinic receptor mediated responses were sensitised, with no change in receptor densities, suggesting changes occur in the cell signalling system beyond the level of the receptor.

Notes: Abstract  Formoterol is a long acting β2-adrenoceptor agonist designed for the alleviation of the symptoms of asthma. This study examined the effects of 14 day administration of formoterol (200 μg/kg/day i.p.) on β1- and β2-adrenoceptors in guinea-pig cardiac and lung tissue. Quantitative autoradiography was used to measure changes in receptor density and organ bath studies determined alterations in functional response. Formoterol treatment produced marked reductions of between 43% and 77% in β2-adrenoceptor density in all regions of the heart (atrioventricular node, bundle of His, right and left bundle branches, interventricular and interatrial septa, right and left atria, ventricles and apex) and lung (bronchial and vascular smooth muscle and parenchyma) (P〈0.01, n=6). β1-Adrenoceptor density remained unchanged in all cardiac and lung regions. In functional studies (−)-isoprenaline was 4 fold less potent at causing relaxation of carbachol (1 μM) precontracted tracheal smooth muscle (pD2: control 8.49±0.03, formoterol 7.91±0.10, P〈0.001, n=4), but formoterol treatment did not change the ability of (−)-isoprenaline to elicit a maximum response. The pKB values for ICI 118,551, 7.33±0.08 in the control and 7.20±0.01 in formoterol treated animals, were between those expected for β1- and β2-adrenoceptors suggesting involvement of both subtypes in the response. In spontaneously beating right atria and electrically paced left atria, tissues in which responses are largely mediated by β1-adrenoceptors, there was no significant change in responses to (−)-isoprenaline (right atria pD2: control 8.45±0.02; formoterol 8.42±0.11; P=0.77, n=4) (left atria pD2: control 8.25±0.03; formoterol 8.47±0.08; P=0.09, n=4). In the presence of CGP 20712A (100 nM) the pKB values did not change with formoterol treatment (left atria: control 9.59±0.12, formoterol 9.66±0.12; P=0.70, n=4) (right atria: control 8.93±0.11, formoterol 9.11±0.07; P=0.25, n=4). The doses and route of administration of formoterol used in this study differs from those used clinically. However, this study demonstrates that chronic formoterol administration produces selective down-regulation of β2-adrenoceptors in the lung and heart. The changes in the lung are accompanied by a shift to the right in the concentration-response curve to β-agonist stimulation with no change in the maximum response.