The affinity of betaxolol, a beta 1-adrenoceptor-selective blocking agent, for beta-adrenoceptors in the bovine trachea and heart.

1. The specificity of betaxolol, a beta-adrenoceptor antagonist, for beta 1- and beta 2-adrenoceptors was compared with that of other beta-antagonists, atenolol, ICI-118551, butoxamine and (+/-)-propranolol, in the bovine trachea and heart by competitive interaction with [3H]-CGP12177 as a radioligand. 2. The radioligand Kd values were 0.75 +/- 0.12 and 1.60 +/- 0.11 nM in the trachea and heart, respectively, and the Bmax values were 34.00 +/- 4.41 and 21.54 +/- 2.94 fmol mg-1 protein, respectively. 3. Using ICI-118551, we determined the ratio of beta 1:beta 2-adrenoceptors in the trachea and heart to be approximately 29:71 and 56:44, respectively. 4. In the trachea, a beta 2-predominant tissue, betaxolol and atenolol were more selective for beta 1-adrenoceptor binding sites than beta 2-adrenoceptor binding sites, whereas ICI-118551 and butoxamine were more selective for beta 2-adrenoceptor binding sites. 5. The beta 1-selectivity of betaxolol was 2.2 and 2.7 fold higher than that of atenolol in the bovine trachea and heart. These findings suggest that betaxolol may be useful in the treatment of hypertension, cardiac arrhythmia and angina pectoris.     

Beta-adrenoceptor antagonist activities and binding affinities of timolol enantiomers in rat atria

S-Timolol is an effective anti-glaucoma drug, but has potentially hazardous side effects. Recently, R-timolol, also, has been reported to be effective in lowering elevated intraocular pressure. In the present study, the beta-adrenoceptor antagonist activities and binding of R- and S-enantiomers of timolol have been examined on rat atrial preparations. The beta-antagonistic activities were investigated using spontaneously beating rat heart atria. Both timolol enantiomers inhibited (-)-isoprenaline-induced chronotropic action competitively. S-Timolol was about 54 times more potent than R-timolol. The apparent binding affinities of timolol enantiomers to beta 1- and beta 2-adrenoceptors were determined by a radioligand binding assay using (-)-[125I]iodocyanopindolol (ICYP) as a marker and CGP 20712 A as a beta 1- and ICI 118,551 as a beta 2-adrenoceptor antagonist. Both enantiomers of timolol inhibited ICYP binding in nanomolar concentrations with Hill coefficients near unity. Neither enantiomer showed selectivity between beta 1- and beta 2-adrenoceptors, but R-timolol was approximately 30 times less active than S-timolol. It is concluded that R-timolol is a relatively potent non-selective beta-adrenoceptor blocking agent, but may possibly exert a more localized beta-adrenoceptor action in the eye than S-timolol, thus improving the safety of ocular timolol therapy.     

The inhibitory effects of iberiotoxin and 4-aminopyridine on the relaxation induced by beta 1- and beta 2-adrenoceptor activation in rat aortic rings.

1. In rat aortic rings contracted by phenylephrine, the relaxation induced by isoprenaline was partly inhibited by iberiotoxin, (ibTX), tetraethylammonium, 4-aminopyridine (4-AP) and 1,9-dideoxyforskolin, but not by glibenclamide. 2. In the presence of 4-AP, 1,9-dideoxyforskolin failed to inhibit further the relaxant response to isoprenaline. Cromakalim-induced relaxation was inhibited by glibenclamide. 3. In the absence of endothelium, ibTX and 4-AP still inhibited the relaxant response to isoprenaline. 4. The inhibitory effect of ibTX on the relaxant response to isoprenaline was eliminated by pretreatment with ICI-118,551, a beta 2-adrenoceptor antagonist, but not by atenolol, a beta 1-adrenoceptor antagonist. 5. The inhibitory effect of 4-AP on the relaxation induced by isoprenaline was abolished by atenolol, but not by ICI-118,551. 6. The inhibitory effect of ibTX on the isoprenaline-induced relaxation in the presence of atenolol was completely abolished by MDL 12,330A, an adenylate cyclase inhibitor. Further, the inhibitory effect of 4-AP on the isoprenaline-induced relaxation in the presence of ICI-118,551 was markedly reduced by MDL 12,330A. 7. The relaxation induced by dibutyryl cyclic AMP was partly inhibited by 4-AP but not by ibTX. However, in the presence of KT5720, an inhibitor of cyclic AMP-dependent protein kinase, ibTX failed to inhibit further the relaxation induced by isoprenaline. 8. These results suggest that, in rat aortic rings, KCa channels are involved in the relaxation induced by isoprenaline. In addition, KCa channels are mainly activated by beta 2-adrenoceptors through cyclic AMP-dependent pathways. Further, the inhibition of isoprenaline-relaxation by 4-AP may be related to the activation of beta 1-adrenoceptors and cyclic AMP formation.     

The beta 1- and beta 2-adrenoceptor affinity of atenolol and metoprolol. A receptor-binding study performed with different radioligands in tissues from the rat, the guinea pig and man

The radioligand binding technique was used to perform a systematic investigation of the beta 1- and beta 2-adrenoceptor affinity of atenolol and metoprolol in tissues from the rat, the guinea pig and man. Radioligands, [125I](+/-)hydroxybenzylpindolol, [125I](-)pindolol, [3H](-)dihydroalprenolol and [3H](-)CGP12177, with different degrees of lipophilicity were used in the binding experiments. In membrane preparations of rat ventricular myocardium and uterus, the number of specific binding sites was similar when comparing experiments performed with the different radioligands. The percentage of the beta 1- and beta 2-adrenoceptor subpopulations in the tissues studied was not dependent on the radioligand or displacing compound used. Furthermore, the affinity of metoprolol and atenolol for beta 1- and beta 2-adrenoceptors was independent of the radioligand used or the tissue studied. The beta 1-adrenoceptor affinity of metoprolol was about 6-7 times higher than that of atenolol, while the beta 1-adrenoceptor selectivity was similar (about 30-fold) for the two beta-blockers. It is concluded that the physical-chemical properties of the radioactive ligands and beta-blockers studied do not affect the results obtained from beta-adrenoceptor-binding experiments in cellular membrane fractions. The beta 1- and beta 2-adrenoceptor affinities did not change in any experiments performed in tissues from the rat, the guinea pig and man for either atenolol or metoprolol.     

Genistein potentiates the relaxation induced by beta1- and beta2-adrenoceptor activation in rat aortic rings

In rat aortic rings, genistein, an inhibitor of tyrosine kinase, but not daidzein, an inactive analogue of genistein, potentiated the relaxation induced by isoproterenol. Atenolol, a beta1-adrenoceptor antagonist, or ICI-118,551, a beta2-adrenoceptor antagonist, inhibited the relaxation induced by isoproterenol. The potentiating effect of genistein on the relaxation induced by isoproterenol in the presence of ICI-118,551 was apparently greater than that in the presence of atenolol. In the presence of ICI-118,551, theophylline, an inhibitor of cyclic adenosine monophosphate (cAMP)-dependent phosphodiesterase (cAMP-PDE), markedly inhibited the potentiating effect of genistein on the isoproterenol-induced relaxation, whereas in the presence of atenolol, theophylline only partly inhibited the potentiating effect of genistein. The relaxation induced by forskolin, an activator of adenylyl cyclase, was potentiated by genistein or theophylline. In the presence of theophylline, the relaxation induced by forskolin was not further affected by genistein. Genistein also inhibited the activities of cAMP-PDE. In the presence of atenolol, but not ICI-118,551, iberiotoxin, an inhibitor of Ca-activated K channels, inhibited the relaxation induced by isoproterenol and the potentiating effect of genistein. In the presence of atenolol, quinacrine, an inhibitor of phospholipase A2, and metyrapone, an inhibitor of P-450 enzymes, but not alpha-naphthoflavone, an inhibitor of P-450 enzymes, indomethacin, a cyclooxygenase inhibitor, or AA861, a 5-lipoxygenase inhibitor, inhibited the potentiating effect of genistein. These results suggest that the potentiation of the beta1-adrenoceptor-induced relaxation by activation of genistein may mostly be due to inhibition of cAMP-PDE activities. In addition, the potentiation of the relaxation induced by activation of beta2-adrenoceptors by genistein may be related to the inhibition of arachidonic acid metabolism and cAMP-PDE activities.     

Beta 1- and beta 2-adrenoceptor antagonist activities of ICI-215001, a putative beta 3-adrenoceptor agonist.

1. The present study was undertaken to characterize the beta 3-adrenoceptor agonist activity of ICI-215001 and to determine whether it exhibits additional activities on beta 1- and beta 2-adrenoceptors in isolated spontaneously beating atrium, trachea and ileum of guinea-pig. 2. In guinea-pig atrium, isoprenaline, a non-selective beta-adrenoceptor agonist, caused concentration-dependent, positive chronotropic effects that were inhibited by atenolol, a selective beta 1-antagonist. ICI-215001 also competitively antagonized the increase in heart rate caused by isoprenaline. 3. ICI-215001 exhibited low intrinsic activity at increasing the beating rate of atrium and no activity on resting or induced tone of tracheal strips. 4. In strips of guinea-pig trachea, contracted submaximally with carbachol, isoprenaline, caused concentration-dependent relaxations. Both ICI-118551, a selective beta 2-adrenoceptor antagonist, and ICI-215001 competitively inhibited the relaxations caused by isoprenaline. 5. In isolated strips of guinea-pig ileum longitudinal smooth muscle contracted with histamine, isoprenaline and ICI-215001 caused relaxations which were inhibited by alprenolol, a beta-adrenoceptor antagonist with modest affinity for beta 3-adrenoceptors, but were resistant to ICI-118551 and atenolol. 6. These results indicate that ICI-215001 exhibits beta 3-adrenoceptor agonist activity as demonstrated by relaxations mediated via atypical beta-adrenoceptors in the longitudinal smooth muscle of guinea-pig ileum. Further, the studies demonstrate that ICI-215001 can act as an antagonist at beta 1- and beta 2-adrenoceptors in situations where its intrinsic agonist activity is low.     

Characteristics of 125I-iodocyanopindolol binding to beta-adrenergic and serotonin-1B receptors of rat brain: selectivity of beta-adrenergic agents

The present study was designed to examine the specificity of beta-adrenergic antagonists for beta 1-, beta 2-adrenergic and 5HT1B-serotonergic receptors by the competitive interaction with 125I-iodocyanopindolol (125I-ICYP) as a radioligand. The beta 1-adrenoceptors were preferred by acebutolol, atenolol, betaxolol, practolol, and l-, dl- and d-metoprolol, while butoxamine and lCl-118,551 preferred beta 2-adrenoceptors. The selectivities of these beta 1- and beta 2-antagonists are well-known, but alprenolol which is known as a non-selective antagonist was 7.2-fold more selective for the beta 2-adrenoceptors in the present study. All beta-antagonists used were more selective towards beta-adrenoceptors as compared with 5HT1B-receptors. Good correlations were observed between the potencies of beta-adrenoceptor antagonists for inhibition of 125I-ICYP binding to beta 1- and beta 2-adrenoceptor sites and their potencies for inhibiting the binding of the same radioligand to 5HT1B-serotonergic receptor sites. These results suggest that beta-adrenoceptor antagonists can bind to beta-adrenoceptors and 5HT1B-receptors.     

Demonstration of the beta 2-adrenoceptor intrinsic efficacy of AY-28,925 using the PGF2 alpha-contracted guinea-pig trachea

The abilities of AY-28,925, labetalol and medroxalol to relax the PGF2 alpha-contracted isolated guinea-pig trachea have been investigated to compare their activities at beta 2-adrenoceptors. Maximum relaxation induced by AY-28,925 was significantly greater than that induced by either labetalol or medroxalol. This relaxation occurred in a concentration-dependent manner over a range of concentrations consistent with the previously determined affinity of AY-28,925 for beta-adrenoceptors. ICI-118,551 inhibited AY-28,925-induced relaxation in a concentration-dependent manner with a pA2 value similar to that determined for ICI-118,551 inhibition of the selective beta 2-adrenoceptor agonist salbutamol, but not the selective beta 1-adrenoceptor agonist norepinephrine. The Schild plot slope for ICI-118,551 inhibition of AY-28,925 or salbutamol did not differ significantly from unity, while that for inhibition of isoproterenol (a non-selective beta-adrenoceptor agonist) did. It is concluded that AY-28,925 is a more efficacious relaxant of tracheal smooth muscle than either labetalol or medroxalol, and that this relaxant activity is the result of its greater intrinsic efficacy at the beta 2-adrenoceptor.     

α-Methyl-5-HT, a 5-HT2 receptor agonist, stimulates β2-adrenoceptors in guinea pig airway smooth muscle

Alpha-methyl-5-HT is widely used as a high-affinity 5-HT(2) receptors agonist, though some studies have postulated that this drug also activates other serotonergic receptors. In the present work, we found that a wide range of concentrations of alpha-methyl-5-HT induced biphasic responses (contraction followed by relaxation) in guinea pig tracheal rings. The relaxing phase caused by 32microM alpha-methyl-5-HT was blocked by 0.1microM propranolol. Furthermore, during an ongoing histamine-induced contraction, alpha-methyl-5-HT (0.1-100microM) produced a concentration-dependent relaxation starting at 10microM. This relaxation was fully abolished by 0.1microM propranolol or 1microM ICI 118,551 (a selective beta(2)-adrenoceptor antagonist). Additionally, in electrophysiological recordings, 32microM alpha-methyl-5-HT also enhanced the membrane K(+) currents of single tracheal myocytes, an effect reverted by propranolol and ICI 118,551, and mimicked by 0.1microM salbutamol. Thus, we concluded that alpha-methyl-5-HT activates beta(2)-adrenoceptors in guinea pig tracheal smooth muscle at concentrations >or=10microM. This effect must be taken into account when this drug is used in airway smooth muscle and in other tissues expressing beta(2)-adrenoceptors.     

Pharmacological analysis of atypical beta-adrenoceptors in the guinea pig gastrointestinal tissue systems

The purpose of the present study was to characterize the atypical beta-adrenoceptors involved in relaxant responses in guinea pig gastric fundus, duodenum and ileum in functional experiments with catecholamines (isoprenaline, noradrenaline and adrenaline), beta 3-adrenoceptor agonists (BRL37344 and CGP12177A) and a non-selective beta 1-, beta 2- and beta 3-adrenoceptor antagonist bupranolol, and to obtain further evidence to clarify whether there is a tissue difference in atypical beta-adrenoceptors in the guinea pig gastrointestinal tissue systems. The atypical beta-adrenoceptors are present in gastric fundus, duodenum and ileum of guinea pig. In the presence of propranolol (1 microM) or atenolol (100 microM) plus butoxamine (100 microM), bupranolol caused a concentration-dependent rightward shift of the concentration-response curves for catecholamines and beta 3-adrenoceptor agonists. There was not a significant difference of pA2 values for bupranolol against these agonists between gastric fundus, duodenum and ileum of guinea pig. These results suggest that guinea pig gastric fundus, duodenum and ileum relaxation are mediated predominantly by an atypical beta-adrenoceptor population whereas the classical beta 1- or/and beta 2-adrenoceptors play a subordinate function role and that the receptors of three tissues are pharmacological identified by functional approaches. There is not a tissue difference in atypical beta-adrenoceptors in the guinea pig gastrointestinal tissue systems between stomach and ileum.     

The distribution of beta-adrenoceptors in dog kidney: an autoradiographic analysis

The distribution of beta-adrenoceptors in slide-mounted dog kidney sections was determined using the radioligand (-)-[125I]cyanopindolol ((-)-[125I]CYP) and autoradiography. Using conditions designed to prevent (-)-[125I]CYP binding to non-beta-adrenoceptor sites, biochemical studies revealed that (-)-[125I]CYP binding equilibrated within 150 min (K1 = 3.2 X 10(8) M-1 min-1), was saturable (KD = 30.72 +/- 2.96 pM; Bmax = 0.57 +/- 0.03 fmol/section, n = 4) and stereoselective with respect to the stereoisomers of propranolol and pindolol. Delineation of beta-adrenoceptor subtypes with the selective beta 1-adrenoceptor antagonist betaxolol and beta 2-adrenoceptor antagonist ICI 118,551 demonstrated that the proportions of beta 1-: beta 2-adrenoceptors was between 1:6 and 1:11. Autoradiographic studies showed that beta 1-adrenoceptors were localized on the juxtaglomerular apparatus and glomeruli, while beta 2-adrenoceptors were localized on medullary rays. The distribution of beta-adrenoceptors with respect to renal function in the dog kidney is discussed.     

Effect of SR59230A on atypical beta-adrenoceptor mediating relaxation in the Guinea Pig gastric fundus

The purpose of the present study was to clarify whether atypical beta-adrenoceptors which presented in the guinea pig gastric fundus are beta(3)-adrenoceptors or putative beta(4)-adrenoceptors. In the presence of both the selective beta(1)-adrenoceptor antagonist atenolol (10(-4) mol/l) and the selective beta(2)-adrenoceptor antagonist butoxamine (10(-4) mol/l), the selective beta(3)-adrenoceptor antagonist SR59230A caused a concentration-dependent rightward shift of the concentration-response curve to catecholamines (isoprenaline, noradrenaline and adrenaline) and beta(3)-adrenoceptor agonists (BRL37344 and CGP12177A) in the guinea pig gastric fundus. Schild plot analyses of SR59230A against these agonists gave pA(2) values of 7.35 +/- 0.03 (isoprenaline), 7.26 +/- 0.04 (noradrenaline), 7.26 +/- 0.05 (adrenaline), 7.79 +/- 0.03 (BRL37344) and 6.74 +/- 0.03 (CGP12177A), respectively, and all Schild slopes were not significantly different from unity. These results suggest that atypical beta-adrenoceptors mediating relaxant responses of these agonists in the guinea pig gastric fundus are beta(3)-adrenoceptors rather than putative beta(4)-adrenoceptors.     

Species variation and stereoselectivity in the metabolism of nicotine enantiomers

1. High performance liquid radiochromatographic systems have been developed for the identification and quantification of 7 urinary metabolites of both S-(-)-[3H-N'-CH3]nicotine and R-(+)-[3H-N'-CH3] nicotine in guinea pig, hamster, rat and rabbit. 2. 3'-Hydroxycotinine was a major urinary metabolite of both S-(-)-nicotine and R-(+)-nicotine in guinea pig, hamster and rabbit. Cotinine was not generally a significant urinary metabolite of either nicotine enantiomer, except in rat, where it constituted 14.6 and 10.4%, respectively, of the total radiolabel in the urine after administration of [3H]-S-(-)-nicotine or [3H]-R-(+)-nicotine. Nicotine N'-oxide was an important urinary metabolite of both nicotine isomers in guinea pig and rat, but in both cases, was not observable in hamster and rabbit. No N-methylated urinary metabolite of S-(-)-nicotine could be detected in any of the species examined. In R-(+)-nicotine experiments, only guinea pig afforded N-methylated metabolites. Significant amounts of 2 unidentified polar, non-basic urinary metabolites of both S-(-)- and R-(+)-nicotine-treated animals were observed. 3. Analysis of the comparative metabolism of the nicotine enantiomers in the four animals species studied, showed that stereoselective differences in the formation of oxidative metabolites existed, particularly in the formation of 3'-hydroxycotinine and nicotine-N'-oxide. A clear stereospecificity was observed in the guinea pig, in that only the R-(+)-nicotine enantiomer was N-methylated in this species. 4. Sex differences appear to exist in the metabolism of nicotine enantiomers in the rat. Female rats excreted more of the unidentified polar metabolite B than male rats, whereas the converse was true for nicotine-N'-oxide. In experiments with R-(+)-nicotine, urinary levels of 3'-hydroxycotinine and R-(+)-nicotine in female rats were higher than in male rats. Conversely, higher amounts of nicotine-N'-oxide were observed in the urine of male rats compared to those in female rats.     

Beta 2-adrenoceptor-mediated increase in the slow inward calcium current in atrial cells

The effects of procaterol (beta 2-adrenoceptor agonist) on the slow inward calcium current (Isi) were examined in single cells of guinea-pig heart. Procaterol increased Isi in atrial cells, in the presence of atenolol (beta 1-adrenoceptor antagonist). The effect was abolished by ICI-118551 (beta 2-adrenoceptor antagonist). However, procaterol did not modify the membrane currents in ventricular cells. These results suggest that beta 2-adrenoceptor agonists bind to beta 2-adrenoceptors in atrial cells and augment Isi, but beta 2-adrenoceptors are not present in ventricular cells.     

Functional beta1- and beta2-adrenoceptors in the left and right atrium of pre-hypertensive rats

There is a small increase in the functional beta2-adrenoceptor response on the spontaneously hypertensive rat (SHR) left atrium in the early stages of hypertension. In the present study, the functional beta1- and beta2-adrenoceptors of the left and right atrium in SHR pre-hypertension and age-matched (5-week-old) Wistar Kyoto (WKY) rats were characterized. Contractility methods with isoprenaline, T-0509 (a selective beta1-adrenoceptor agonist) and procaterol (a selective beta2-adrenoceptor agonist) were used. At 5 weeks, the SHRs were pre-hypertensive. Isoprenaline was more potent on the left atrium of 5-week-old SHRs than WKY rats. Bisoprolol, a selective beta1-adrenoceptor antagonist, was more potent against isoprenaline and T-0509 on the SHR than WKY rat left atrium. ICI 118,551, a selective beta(2)-adrenoceptor antagonist, was more potent against procaterol and T-0509 on the SHR than WKY rat left atrium. The results with bisoprolol and ICI 118,551 suggest that there are more functional beta(1)- and beta(2)-adrenoceptors on the left atrium of 5-week-old SHRs than WKY rats. Isoprenaline, T-0509 and procaterol were equipotent on the right atrium of 5-week-old WKY rats and SHRs. Bisoprolol was more potent against isoprenaline, T-0509 and procaterol on the SHR than WKY rat right atrium. ICI 118,551 was more potent against T-0509, but not isoprenaline and procaterol, on the SHR than WKY rat left atrium. This suggests there are more functional beta1-adrenoceptors, and probably more functional beta2-adrenoceptors, on the right atrium of 5-week-old SHRs than WKY rats. These functional differences in beta1- and beta2-adrenoceptor-mediated responses of the left and right atria of pre-hypertensive SHRs cannot be caused by hypertension, and may be associated with the onset of hypertension.     

Effects of betaxolol on cardiohemodynamics and coronary circulation in anesthetized dogs: comparison with atenolol and propranolol

Effects of betaxolol, a cardioselective beta-adrenoceptor antagonist, on cardiohemodynamics and coronary circulation were investigated in two kinds of anesthetized open-chest dog preparations in comparison with those of atenolol and propranolol. When administered intravenously, betaxolol, atenolol and propranolol produced dose-dependent decreases in the heart rate (HR), maximum left ventricular dP/dt [+)dP/dt), cardiac output (CO) and mean arterial pressure (MAP). Although all three drugs were almost equipotent in decreasing HR, betaxolol was much less potent than atenolol and propranolol in decreasing (+)dP/dt. Betaxolol decreased the total peripheral resistance (TPR), whereas atenolol and propranolol increased it. In another series of experiments, when administered intravenously, betaxolol, atenolol and propranolol all produced a decrease in the myocardial oxygen consumption (MVO2) and an increase in the atrioventricular conduction time (AVCT). All three drugs were nearly equipotent in decreasing MVO2, although betaxolol was less potent than the other two drugs at higher doses (greater than 300 micrograms/kg). Prolongation of AVCT with propranolol was stronger than those with betaxolol and atenolol. These results suggest that, unlike atenolol and propranolol, the decrease in TPR as well as beta 1-adrenoceptor blockade may be responsible for both the hypotensive effect of betaxolol and the decrease in MVO2 with betaxolol. The result that the cardiodepressant effect of betaxolol was much less potent than those of atenolol and propranolol suggests that betaxolol would be more beneficial than the others in the treatment of ischemic heart disease.     

Sympathetic nerve stimulation activates both beta 1- and beta 2-adrenoceptors of SA and AV nodes in anesthetized dog hearts.

We investigated blocking effects of the selective beta 1-adrenoceptor blocker atenolol (0.1-100 micrograms/kg, i.v.), the selective beta 2-adrenoceptor blocker ICI 118,551 (1-1000 micrograms/kg, i.v.) and the combination of the two drugs on positive chronotropic and dromotropic responses to norepinephrine (NE) released by stimulation of the sympathetic nerves in anesthetized, neurally decentralized, open-chest dogs after atropine was given. Stimulation of the intracardiac sympathetic nerves to the SA nodal region or to the AV nodal region selectively increased heart rate or decreased AV conduction time, respectively. ICI 118,551 inhibited the chronotropic or dromotropic response to each stimulation in a dose-dependent manner, but its inhibition of the dromotropic response was less than that of the chronotropic response. Atenolol similarly inhibited either the positive chronotropic or dromotropic response to each stimulation in a dose-related manner. The combination of atenolol and ICI 118,551 attenuated the responses to each stimulation more than atenolol alone. These data indicate that sympathetic nerve stimulation activates both beta 1- and beta 2-adrenoceptors of the SA and AV nodes and that the proportion of beta 2-adrenoceptor-mediated effects on the AV node is less than that on the SA node. These results suggest that neurally released NE in part controls physiological functional cardiac responses mediated through beta 2-adrenoceptors, in addition to the responses predominantly mediated through beta 1-adrenoceptors.     

Selective regulation of beta 1- and beta 2-adrenoceptors in the human heart by chronic beta-adrenoceptor antagonist treatment.

1. In 44 patients undergoing coronary artery bypass grafting, the effect of chronic administration of the beta-adrenoceptor antagonists sotalol, propranolol, pindolol, metoprolol and atenolol on beta-adrenoceptor density in right atria (containing 70% beta 1- and 30% beta 2-adrenoceptors) and in lymphocytes (having only beta 2-adrenoceptors) was studied. 2. beta-Adrenoceptor density in right atrial membranes and in intact lymphocytes was assessed by (-)-[125I]-iodocyanopindolol (ICYP) binding; the relative amount of right atrial beta 1- and beta 2-adrenoceptors was determined by inhibition of ICYP binding by the selective beta 2-adrenoceptor antagonist ICI 118,551 and analysis of the resulting competition curves by the iterative curve fitting programme LIGAND. 3. With the exception of pindolol, all beta-adrenoceptor antagonists increased right atrial beta-adrenoceptor density compared to that observed in atria from patients not treated with beta-adrenoceptor antagonists. 4. All beta-adrenoceptor antagonists increased right atrial beta 1-adrenoceptor density; on the other hand, only sotalol and propranolol also increased right atrial beta 2-adrenoceptor density, whereas metoprolol and atenolol did not affect it and pindolol decreased it. 5. Similarly, in corresponding lymphocytes, only sotalol or propranolol increased beta 2-adrenoceptor density, while metoprolol and atenolol did not affect it and pindolol decreased it. 6. It is concluded that beta-adrenoceptor antagonists subtype-selectively regulate cardiac and lymphocyte beta-adrenoceptor subtypes. The selective increase in cardiac beta 1-adrenoceptor density evoked by metoprolol and atenolol may be one of the reasons for the beneficial effects observed in patients with end-stage congestive cardiomyopathy following intermittent treatment with low doses of selective beta 1-adrenoceptor antagonists.     

Photoaffinity labeling of beta-adrenergic receptors in mammalian tissues

Photoaffinity labeling of beta 1- and beta 2-adrenergic receptors in plasma membranes from various mammalian tissues has been been performed utilizing the recently developed beta-adrenergic antagonist probe [125I]para-azidobenzylcarazolol. Tissues studied and their proportions of beta 1 and beta 2 receptors were: rat lung (18% beta 1, 82% beta 2), rabbit lung (72% beta 1, 28% beta 2), guinea pig lung (15% beta 1, 85% beta 2), dog lung (20% beta 1, 80% beta 2) and rabbit skeletal muscle (10% beta 1, 90% beta 2). As assessed by autoradiograms of sodium dodecyl sulfate-polyacrylamide gel electrophoresis, two to three specifically protected bands of Mr 62,000-65,000, 50,000-55,000 and 38,000-42,000 were observed in each tissue system. In each case, beta-adrenergic agonists and antagonists protected against photolabeling with appropriate beta 1 and beta 2 selectivity. Thus, in rat lung the beta 2 selective antagonist ICI-118,551 was more potent in blocking incorporation than the beta 1 selective antagonist betaxolol, whereas in rat, dog and guinea pig lung and rabbit skeletal muscle epinephrine was more potent than norepinephrine in blocking labeling, indicating a beta 2 specificity in these tissues. Conversely, in rabbit lung membranes, norepinephrine was approximately equipotent with epinephrine in blocking photoincorporation, indicating a beta 1 selectivity. In some systems protease inhibitors, especially those specific for metalloproteases (EDTA, EGTA), markedly diminished the amount of the smaller Mr peptides. For example, in rat lung the ratio of Mr 62,000:47,000:36,000 peptides changed from 30:40:30 to 60:35:5 in the presence of inhibitors. These results demonstrate the applicability of using [125I]para-azidobenzylcarazolol to covalently label mammalian beta-adrenergic receptors and suggest that mammalian beta 1 and beta 2 receptor binding sites primarily reside on peptides of Mr 62,000-65,000 and that smaller ligand binding fragments may arise by proteolysis.     

Uptake of radioligands by rat heart and lung in vivo: CGP 12177 does and CGP 26505 does not reflect binding to beta-adrenoceptors.

The biodistribution of (-)-4-(3-t-butylamino-2-hydroxypropoxy)-[5,7-3H-benzimidazol-2-one (CGP12177, a non-selective beta-adrenoceptor antagonist) and 1-[2-(3-carbamoyl-4-hydroxy)-(5-3H-phenoxy)]-2-propanol methanesulfonate, (CGP26505, a beta 1-adrenoceptor antagonist) was studied in rats pretreated with various alpha- and beta-adrenoceptor blocking drugs (5 min before 3H injection, in dosages at which the drugs demonstrated the expected selectivity). Cardiac and pulmonary radioactivity were measured after 10 min, when specific binding was maximal. Uptake of [3H]CGP12177 was linked to binding to beta-adrenoceptors since it was not affected by prazosin or yohimbine, and was equally well inhibited by propranolol, unlabelled CGP12177 and isoprenaline. Moreover, atenolol and CGP20712A inhibited [3H]CGP12177 uptake in heart (predominantly beta 1-adrenoceptors) more potently than ICI 118,551, while in lungs (predominantly beta 2-adrenoceptors) ICI 118,551 was more potent than atenolol or CGP20712A. In contrast, [3H]CGP26505 uptake in the target organs was equally effectively inhibited by propranolol and ICI 118,551, and significantly lowered by alpha-adrenoceptor antagonists. We conclude that [11C]CGP12177, but not [11C]CGP2605 will be suitable for positron emission tomography imaging of beta-adrenoceptors in animals.