New methods for determining dissociation constants of agonist-receptor complexes

Abstract

Methods are presented which enables the dissociation constant of agonist-receptor complexes (KA) to be estimated without knowledge of the relative intrinsic efficacies. The dissociation constants are estimated from fixed agonist-competitive antagonist concentration combinations which simulate partial agonists. In general, estimating of KA by employing these methods require ф and KB to be known; where ф = antagonist]/[agonist] and KB is the dissociation constant of the competitive antagonist-receptor complex. Some of these methods require only knowledge of ф, while KB may be unknown. A method is also described to estimate KA without knowing the value of KB and the competitive antagonist concentration. In addition to the estimation of KA values, a new method for estimation of the dissociation constants of competitive antagonist-receptor complexes (KB) is reported. One of the new methods was exemplified practically by using sets of experimental agonist concentration-effect curves determined in the absence and presence of increasing concentrations of competitive antagonists. From this pharmacological data the apparent KA value for carbachol (in muscarinic M3 receptors of the guinea-pig ileum) was determined. This action illustrated that the apparent affinity values (KA) and height of the stimulus curve of an agonist may be determined from published pharmacological data. This procedure affords a possibility to establish whether or not spare receptors are present in a particular system. It was also shown that relative affinity values provide more reliable information than does isolated KA values.