In vitro and in vivo studies on the selectivity of beta-adrenoceptor antagonist drugs in the guinea-pig

Abstract

The main aim of this study was to determine to what extent estimates of the receptor (b2:b1) selectivity of b-adrenoceptor antagonists obtained in vitro in the guinea-pig might reflect the tissue (trachea:heart) selectivity of the same antagonists in vivo in the same species. To achieve this aim in vitro experiments with guinea-pig isolated atria (rate) and intrinsic-tone trachea (relaxation) were carried out under optimal experimental conditions for adrenoceptor selectivity studies. The b-adrenoceptor agonists used in vitro were fenoterol, noradrenaline and isoprenaline and various b-adrenoceptor antagonists were investigated viz a-methylpropranolol, 4-BIP [(p)-l-(4-benzimidazoloxy)-3-isopropylamino-2-propanol], 5-BIP [(p)-l-(5-benzimidazoloxy-3-isopropyl amino-2-propanol] and IPS 339 [(t-butylamino-3-ol-2-propyl)oximino-9-fluorene]. All these antagonists had been claimed in prior studies as having some degree of b2-selectivity but the published data on which this conclusion was based were not obtained under conditions considered optimal for the determination of selectivity. Receptor selectivity estimates, calculated from the difference in pA2 values obtained on trachea, using fenoterol as agonist and on atria, using noradrenaline as agonist, were obtained for each antagonist and were 11.0 for a-methylpropranolol, 2.5 for 4-BIP and 4.9 for IPS 339; no estimate was calculated for 5-BIP. Experiments conducted with carbachol - contracted tracheal preparations from the same animals showed these to be unsuitable preparations for the estimation of receptor selectivity since conditions satisfying Schild plot theory could not be met.a-Methylpropranolol, being the most selective of these antagonist compounds, was selected for study in vivo. Before an in vivo estimate of its selectivity could be obtained an experimental model in the intact guinea-pig, which mirrored the in vitro responses, had to be developed. The experimental model chosen involved measuring heart rate, tracheal segment pressure and diastolic blood pressure responses to isoprenaline and other selective agonists in anaesthetized guinea-pigs and led to the following conclusions on methodology. Isoprenaline was selected as agonist since it gave short-acting, reproducible responses and could be given in a cummulative dose to enable several doses of the b-adrenoceptor antagonist to be investigated in any one guinea-pig. Endogenous catechol amines were depleted by the use of reserpine. Cardiovascular reflexes were reduced or inhibited by bilateral vagotomy. Responses were also obtained in the presence of the a-adrenoceptor antagonist phenoxybenzamine, to mimic as far as practicable conditions used in the in vitro experiments.Since only one b2-selective adrenoceptor antagonist, a-methylpropranolol, had been selected for the in vivo study, two other selective antagonists, for which satisfactory in vitro data were already available, were also studied in vivo. They were ICI 118,551 (erythro-DL-l-(7-methylindan-4-yloxy)-3-isopropylamino-butan-2-ol), having a b2-selectivity of 53.7 and atenolol, having a b1-selectivity of 27.8 (which is equivalent to a b2-selectivity of 0.036). Estimates of tissue (trachea:heart) selectivity in vivo were calculated from the difference in qin vivo pA2q values obtained on tracheal segment pressure and heart rate, using isoprenaline as agonist. These estimates were less than the receptor (b2:b1) selectivity estimates determined in vitro, i.e. they were 1.7 for a-methylpropranolol, 4.0 for ICI 118,551 and 0.23 for atenolol. a-Methylpropranolol selectivity increased marginally to 2.0 if the b2-selective agonist rimiterol was used in place of isoprenaline. Vascular:heart selectivity estimates were also obtained, but only for comparative purposes, and were 1.1 for a-methyl propranolol, 2.8 for ICI 118,551 and 0.12 for atenolol.It is concluded that the estimates of tissue (trachea:heart) selectivity of b-adrenoceptor antagonists obtained in vivo in guinea-pigs, usually with isoprenaline (non-selective) as agonist drug, follow the same relative trend as those obtained in vitro for the same tissues (isolated trachea, atrial rate) in the same species, but are quantitatively less. If selectivity in vitro is obtained using selective agonists instead of isoprenaline, then because of the possibility of heterogeneous b-adrenoceptor populations this difference is even more exaggerated. The use of selective agonists in vivo , to try and allow for the possibility that responses could be mediated by both subtypes of b-adrenoceptors, was hindered by technical problems due to, for example, vasoconstrictor actions (in the case of noradrenaline) and a prolonged duration of action of the agonists.