Relaxation Ofcat Trachea by 13-Adrenoceptor Agonists

Br. J. Pharmac. (1983), 78,417-424 Relaxation of cat trachea by 13-adrenoceptor agonists can be mediated by both f3I- and 132-adrenoceptors and potentiated by inhibitors of extraneuronal uptake Stella R. O'Donnell & Janet C. Wanstall Pharmacology Section, Department of Physiology and Pharmacology, University of Queensland, St Lucia, Brisbane, Qld. 4067, Australia 1 Responses (relaxation) to the P-adrenoceptor agonists, isoprenaline, fenoterol or noradrenaline, were obtained on cat tracheal preparations contracted with a submaximal concentration of carbachol (0.5 JAM). 2 The relative potencies of isoprenaline: fenoterol: noradrenaline were 100:8.1:10.7. From this, it was concluded that responses were mediated predominantly by 131-adrenoceptors but that a minor population of P2-adrenoceptors might also be involved. 3 Schild plots for the selective antagonists atenolol (Pi1-selective) or ICI 118,551 (32-selective) were in different locations, i.e. were separated, depending on whether the antagonist was antagoniz- ing noradrenaline or fenoterol. This supported the conclusion that 12- as well as Pl-adrenoceptors were involved in mediating the response. In this respect, cat trachea resembles cat atria (rate responses). 4 In the presence of atenolol the concentration-response curves to fenoterol became biphasic. This was interpreted as indicating that the P2-adrenoceptors were too few in number to elicit a maximum tissue response. 5 Responses to isoprenaline of cat trachea were potentiated by the extraneuronal uptake inhibitor drugs, corticosterone and metanephrine. This indicated that extraneuronal uptake could modulate P-adrenoceptor-mediated responses (relaxation) of cat trachea. 6 Cat trachea resembles guinea-pig trachea in that (i) the P-adrenoceptor population mediating relaxation is mixed (PI + P2) and (ii) responses to isoprenaline are modulated by its extraneuronal uptake. However, cat trachea differs from guinea-pig trachea in that the predominant 1- adrenoceptor sub-type is Pi not P2- Introduction In cat trachea, the P-adrenoceptors mediating relaxa- interest to determine whether the influence of ex- tion were classified by Lulich, Mitchell & Sparrow traneuronal uptake on ,B-adrenoceptor-mediated (1976) as P1-adrenoceptors. Thus they appeared to tracheal relaxation was also different in the two be different from the P-adrenoceptors mediating re- species. This question was relevant to the hypothesis sponses in guinea-pig trachea, which were originally that extraneuronal uptake may be more closely as- said to be P2-adrenoceptors (Farmer & Levy, 1970). sociated with P2-adrenoceptors than pl- More recently, it has been possible to demonstrate adrenoceptors (Bryan, Cole, O'Donnell & Wanstall, that PI as well as P2-adrenoceptors can mediate relax- 1981). An extraneuronal uptake of catecholamines ation of guinea-pig trachea (Furchgott, 1976; has been demonstrated in the trachealis smooth mus- O'Donnell & Wanstall, 1979a). Therefore it was of cle cells from both species, using a fluorescence his- interest to determine whether tracheal relaxation in tochemical technique (Anning, Bryan & O'Donnell, cat could also be mediated by both sub-types of 1978) and responses to isoprenaline on guinea-pig P-adrenoceptor. trachea are known to be potentiated when this up- Because the P-adrenoceptor sub-type predomin- take is inhibited by extraneuronal uptake inhibitor antly responsible for tracheal relaxation was appar- drugs (O'Donnell & Wanstall, 1976). Experiments ently different in guinea-pig and cat, it was also of have now been carried out to determine whether 0007-1188/83/020417-08 $01.00 'P) The Macmillan Press Ltd 1983 418 STELLA R. O'DONNELL & JANET C. WANSTALL potentiation of isoprenaline by extraneuronal uptake used (see Results), values of concentration-ratio inhibitor drugs can be demonstrated on cat trachea. (CR) were calculated from the EC50 in the presence A preliminary report of some of these data formed of the antagonist divided by the EC50 in the absence part of a communication to the British Pharmacologi- of antagonist (control). It was not necessary to cor- cal Society (Cole, O'Donnell & Wanstall, 1982). rect these CR values since, in a separate series of experiments, it was shown that there was no signific- ant change in sensitivity of the preparations with time Methods and/or pre-exposure to the agonists. Plots of log Cat trachealpreparations (CR-1) vs. log molar concentration of antagonist (log [B]) were obtained according to the method of Cats of either sex, weighing 1.5 to 3.5 kg, were pre- Arunlakshana & Schild (1959). A linear least treated with reserpine (0.1 mg/kg i.p. 18 h previous- squares regression analysis (Snedecor & Cochran, ly). They were anaesthetized with ether and then 1967) was used to obtain the line of best fit through exsanguinated. The trachea was removed and single the combined data points from a number of cats, and ring preparations (up to 4 preparations from each this line is referred to as a Schild plot. None of the cat) were set up in Krebs solution at 37°C and gassed Schild plots had slopes which differed significantly with 95% 02 and 5% CO2. In some experiments the from 1.0 (Table 2). Thus pA2 values could be calcu- whole tracheal ring was mounted, at a resting tension lated for each concentration of antagonist used from of 1.5 g, around two horizontal stainless steel pins the equation pA2 = log (CR-1) - log [B] and a mean (one fixed and one linked to a Statham Universal pA2 value obtained for each cat. For each agonist- transducer, UC3 + UL5) and changes in isometric antagonist combination a mean pA2 value ± s.e. was tension recorded. In other experiments, most of the then calculated from a number of animals. cartilage was removed from the ring and the remain- ing smooth muscle preparation was set up, at a ten- Effects ofextraneuronal uptake inhibitors on sion of 300 mg, so that changes in length could be isoprenaline-induced relaxation ofcat trachea recorded isotonically using a modified Statham lOB strain gauge. No quantitative differences were ob- a-Adrenoceptors and neuronal uptake were blocked served between these two types of preparation. Prep- with phentolamine (10 Mm) and cocaine (10jM) re- arations taken from the region of the trachea nearest spectively. Concentration-response curves to isop- to the larynx are referred to as from the cervical renaline were obtained in the absence and presence trachea and those taken just before the trachea bifur- of metanephrine (50 Mm) or of increasing concentra- cated into the bronchi are referred to as from thoracic tions (1, 10 and 50 Mm) of corticosterone. The in- trachea. hibitors were in contact with the tissue for 30 min Preparations were contracted with a submaximal before, and also during, the isoprenaline concentration (0.5 pM) of carbachol. Cumulative concentration-response curve. The corticosterone concentration-response (relaxation) curves were ob- stock solution was made up in absolute ethanol. tained to P-adrenoceptor agonist drugs and EC50 Therefore in the cortiscosterone experiments, a con- values (molar concentration of agonist producing stant concentration of ethanol (17.2 mM) was main- 50% maximum response) interpolated. The potency tained in the organ bath throughout the experiment of the agonist drugs was expressed as the negative log by adding, when necessary, an appropriate amount of EC50. absolute ethanol to the Krebs solution in the organ bath before obtaining an isoprenaline concentration- Schildplotsforfi-adrenoceptorantagonists on cat response curve. Potentiation of isoprenaline, i.e. shift trachea of the log concentration-response curve to a lower concentration range, was calculated from [neg log a-Adrenoceptors, neuronal uptake and extraneuron- EC50 (inhibitor present) - neg log EC50 (control)]. In al uptake were blocked with phentolamine (10 Mm), 4 of the 10 experiments with metanephrine the cocaine (10 AM) and metanephrine (50 AM) respec- potentiation of isoprenaline was corrected to allow tively (in contact with the tissue for 30 min before and for antagonism of isoprenaline due to the weak P- then throughout the experiments). Concentration- adrenoceptor antagonist action of metanephrine response curves to either fenoterol (32-selective (Kenakin, 1980). In order to do this, concentration- agonist) or noradrenaline (pl-selective agonist) were response curves were obtained for both fenoterol and obtained in the absence and presence of increasing isoprenaline (in random order), in the absence and concentrations of a ,-adrenoceptor antagonist. The presence of 50 Mm metanephrine. Fenoterol is not a contact time for the antagonist drugs was 60 min. substrate for extraneuronal uptake and metane- Except in the experiments with atenolol using phrine antagonized responses to fenoterol, i.e. fenoterol as agonist, when a different approach was shifted the fenoterol concentration-response curves P-ADRENOCEPTORS AND EXTRANEURONAL UPTAKE IN TRACHEA 419 to a higher concentration range. Thus corrected val- solutions were made in Krebs solution and kept on ues for the potentiation of isoprenaline by metanep- ice during the course of the experiment. hrine, due to inhibition of extraneuronal uptake, The composition of the Krebs solution was (mM): were obtained by adding the value for the antagonism NaCl 114, KCl 4.7, CaCl2 2.5, KH2PO4 1.2, MgSO4 of fenoterol (in log units) to the value for the potenti- 1.2, NaHCO3 25, glucose 11.7, ascorbic acid 1.1. ation of isoprenaline (in log units). Statistical analyses Cat right atrial

Load more