Dissociation Constants of Isoprenaline and Orciprenaline and Their Relative Efficacies on Guinea‐pig Isolated Atria Determined by Use of an Irreversible Β‐adrenoceptor Antagonist

Abstract

I Dissociation constants (KA) of isoprenaline and orciprenaline were determined for the positive inotropic and chronotropic responses of guinea-pig isolated atria. Cumulative dose-response curves to the agonists were constructed before and after incubation with and washout of the irreversible ,3-adrenoceptor antagonist, Ro 03-7894 1-(5-chloracetylaminobenzfuran-2-yl)-2-isopropylaminoethanol). 2 After 20 min washout, the curves were displaced to the right with depression of the maxima. After 3 h washout, there was only depression of the maxima. 3 Dissociation constants were determined by plotting reciprocals of molar concentrations before Ro 03-7894 (1/A) against reciprocals of the equiactive concentrations after Ro 03-7894 (1/A'). KA = (slope -1)/intercept. 4 Isoprenaline had a greater affinity (KA) than orciprenaline on both rate and tension. The affinity for rate and tension was identical for both agonists, indicating that the ,B-adrenoceptors were identical. 5 Isoprenaline and orciprenaline produced identical rate response maxima when compared in the same preparation but the orciprenaline tension maximum was only 92.0 + 0.3% that of isoprenaline. 6 These dose-response curves were replotted as response against -log RA/Rt (fraction of receptors occupied) for each agonist concentration, calculated from the equation RA/Rt = (A)/(KA + (A)).

Calculated and actual changes in β-adrenoceptor number associated with increases in rat and guinea-pig cardiac sensitivity

“…This confirms previous reports of an increase in cardiac sensitivity to $-adrenoceptor agonists after chronic catecholamine depletion (see Broadley et a1 1984). There was an increase in basal developed tension after reserpine pretreatment which agrees with previous studies (Broadley & Lumley 1977;Tenner et a1 1982;Torphy et a1 1982) and has been interpreted to indicate a change in the excitationcontraction coupling (Tenner et a1 1982). However, the consensus is that the supersensitivity to padrenoceptor agonists is additional to any change in coupling and in the present study it was associated with an increase in P-adrenoceptor binding sites.…”

Temperature‐induced changes in dissociation constants (KA) of agonists at cardiac β‐adrenoceptors determined by use of the irreversible antagonist Ro 03–7894

“…The depression of maximum responses was generally less at the lower temperature, confirming earlier observations (Broadley & Nicholson, 1980) and reflecting the enhanced receptor sensitivity at the lower temperature. Dissociation constants were calculated only after thorough washout of free Ro 03-7894 since any residual antagonist exerts an additional competitive antagonism which distorts the KA value (Broadley & Nicholson, 1981). Furthermore, care was taken to measure equiactive concentrations on the linear portions of pre-and postRo 03-7894 dose-response curves (see Thron, 1970).…”

“…However, it has recently been shown that the kinetics of agonist and antagonist binding are different (Weiland, Minneman & Molinoff, 1979;1980) and the possibility that the supersensitivity may be the result of an increase in affinity of agonists only, must be considered. With the availability of a 3-adrenoceptor antagonist, Ro 03-7894 (1-(5-chloracetylaminobenzfuran-2-yl)-2-isopropylaminoethanol), which has been shown to behave in an apparently irreversible manner (Nicholson & Broadley, 1978;Nicholson, Broadley, Burden & Natoff, 1979;Rankin & Broadley, 1982), it has been possible to determine pharmacologically the affinities of sympathomimetic amines for cardiac f-adrenoceptors (Broadley & Nicholson, 1981;Siegl & McNeill, 1982). In the present study, the affinities of isoprenaline and orciprenaline were determined as their dissociation constants, in three cardiac preparations of the guineapig, to assess whether changes occur during hypothermia-induced supersensitivity.…”

Is Ro 03–7894 an irreversible antagonist at β‐adrenoceptor sites?

“…The Macmillan Press Ltd 1984 The concentration of Ro 03-7894 used in the present study (0.6 mM) was similar to that used in the initial in vitro studies with this agent (Nicholson & Broadley, 1978;Broadley & Nicholson, 1981).…”

“…In previous studies it has been suggested that the compound Ro 03-7894 (1-(5-chloroacetylaminobenzfuran-2-yl)-2-isopropylaminoethanol) behaves as an irreversible, non-competitive antagonist at Padrenoceptor sites in guinea-pig isolated cardiac tissue (Nicholson & Broadley, 1978;Broadley & Nicholson, 1981;Rankin & Broadley, 1982). The evidence for this classification of the antagonist was based on the observations that it reduced the slope and maxima of positive inotropic and chronotropic concentration-response curves to isoprenaline and orciprenaline without affecting curves established to histamine or calcium (Nicholson & Broadley, 1978).…”