No abstract

Observations on rates of dissipation of responses to various agonists, protection of receptors by residual agonist after washout, and effects of enzyme inhibitors on these processes established that relaxation of rabbit aortic strips is controlled by the concentration of agonist remaining in the environment of tissue receptors. Thus, the rate of relaxation is a valid index of overall drug inactivation. This index of drug disposition was combined with a procedure of oil immersion, which traps a fixed amount of drug in an extracellular compartment only slightly larger than the interstitial space of the tissue. Oil immersion did not detectably alter the functional capacity of the tissue and effectively separated intrinsic disposition from diffusion into the external medium. Examples are given to show that, in combination with inhibitors of specific pathways of disposition, this preparation provides a simple, quantitative, reproducible method for the study of mechanisms of drug inactivation, with the advantages that it (a) allows the study of responses and disposition in the same preparation, (b) allows analysis of the inactivation of any concentration of agonist which produces a recordable response, (c) equates disposition with termination of action, and (d) specifically follows effective drug in the environment of receptors rather than total drug in the tissue.

SUMMARY1. The effect of ambient temperature on the properties of adrenoceptors mediating inotropic responses was assessed in isolated frog hearts on the basis of the effects and tissue uptake of ox-and fl-adrenoceptor antagonists.2. At temperatures of 230 C and above inotropic responses to adrenaline were antagonized by propranolol (0-4-4.0 uM), but were unaffected by phentolamine (26.5 AM) and were potentiated by phenoxybenzamine (POB) (0.7-29.5 AM). Below 170 C the activity of propranolol was reduced at least tenfold, and the a-adrenoceptor antagonists inhibited responses to both adrenaline and isoprenaline, but not those to CaC12.3. The responses of hearts exposed to POB at 140 C and then tested, after thorough washing, at both 14 and 240 C were similarly inhibited at both temperatures, i.e. the usual fl-adrenoceptor response did not appear at the higher temperature. Conversely, exposure to POB at 240 C produced only potentiation at both test temperatures.4

Effects of cocaine on the magnitude of responses to several biologically active amines and on their rates of inactivation were studied in strips of rabbit thoracic aorta in vitro. Although cocaine both potentiated responses to noradrenaline, adrenaline and phenylephrine and slowed their inactivation, the correlation between these two parameters under various experimental conditions was poor, and in all cases the delay in intrinsic inactivation was inadequate to account for the observed potentiation. Potentiation of responses to noradrenaline by cocaine was little decreased in strips stored at 6° C for up to 10 days, although the response to low doses of tyramine was abolished much earlier. Similarly, cocaine clearly potentiated responses to noradrenaline for at least 28 hr at 37° C, at which time responses to noradrenaline alone were markedly decreased. Cocaine potentiated responses to phenylephrine as well after 60 as after 10 min exposure to the amine in strips in which all intra‐neuronal disposition of this amine had been eliminated by treatment with reserpine and iproniazid. Cocaine effectively potentiated responses to histamine, but had only a slight and variable effect on those to 5‐hydroxytryptamine (5‐HT). It did not alter the tissue inactivation of histamine, but did significantly slow the inactivation of 5‐HT. Procaine slowed amine inactivation in the same way and to the same extent as did cocaine, but did not potentiate responses or affect the potentiation produced by cocaine added in its presence. Cocaine potentiated responses to methoxamine to approximately the same degree as it did those to noradrenaline, although studies by the oil immersion technique clearly demonstrated that the aortic strips were entirely incapable of inactivating methoxamine. The observations reported and discussed are incompatible with the hypothesis that cocaine potentiates responses to sympathomimetic amines because it prevents their inactivation by nerve uptake and storage and thus diverts larger amounts of agonist to tissue receptors. It is concluded that potentiation and inhibition of amine inactivation reflect two largely independent actions of cocaine in this vascular smooth muscle preparation, and probably in other organs, and that potentiation is a generally unreliable criterion of the blockade of processes inactivating sympathomimetic amines or of the importance of these processes in terminating the action of the amines.

·ABSTRACTThe present series of experiments provided evidence that the generally accepted distinction between a-and s-adrenergic receptors is not immutable. In frog hearts and rat atria 3H-phenoxybenzamine (3H-POB ) blocked inotropic responses to catecholamines only at temperatures below l7 0 C and potentiated responses ab ove 23 0 C, and considerably more radioactivity was bound to the myocardium at lower temperatures. The converse was true for l4C-propranolol, and the potency of a-and s-adrenergic agonists was shawn to parallel the effectiveness of the blocking agents. Alkylation of a-adrenergic receptors by POB at l4 0 c prevented the appearance of s-adrenergic receptors when the temperature was subsequently raised ta 24 o C.Phentolamine prevented block by 3H-POB and considerably reduced its binding ta the myocardium at low but not at high temperatures. After exposure ta unlabelled POB in the presence of phentolamine at low temperature and thorough washing, exposure to 3H-POB produced a IIpositive label" of the adrenergic receptors, which was localized in a 20,000 9 (ll mitochondrial ll ) and in a IIsoluble protein" fraction. Pretreatment of rats with 6-hydroxydopamine, but not with reserpine, prevented the appearance of oe-adrenergic receptor charac.teristics in left atria at low temperatures; this indicated that adrenergic innervation, but probably not stores of neurotransmitter, is necessary to allow transformation of the receptors. It is suggested that a-and S-adrenergic receptors may be different allosteric conformations of the same molecule.