Characterization of alpha-adrenoceptors in myometrium of preparturient rats

We describe three methods for the quantitative analysis of the alpha-adrenoceptor subtypes in preparturient rat myometrial membrane fractions. A non-subtype-selective antagonist radioligand. [3H]dihydroergocryptine ([3H]DHE), was used to label all of the alpha-receptors. [3H]DHE bound to both alpha 1- and alpha 2-receptors with indistinguishable affinity. Computer modelling of competition curves of unlabeled selective antagonists or agonists was then required in order to determine reliably alpha 1 and alpha 2 affinities and proportions: the alpha 1-receptors represent 45% and the alpha 2-receptors 55% of the entire alpha-receptor population in rat uterus. The second approach involved the administration of phenoxybenzamine (POB) that irreversibly blocks the alpha 1-adrenoceptors. Myometrial membranes obtained from rats 1 h after the administration of varying amounts of POB showed a dose-dependent reduction in specific [3H]DHE binding. This reduction was accompanied by a progressive increase of the value of the dissociation constant. Our data indicate that a dose of 1 mg of POB left the alpha 2-receptors intact while entirely blocking the alpha 1-receptors in rat myometrium. The third approach utilized the selective radioligand antagonists [3H]prazosin ([3H]PRAZ) and [3H]rauwolscine ([3H]RAUW). The results obtained with these radioligands confirmed our observations on the alpha-adrenoceptor subtypes in experiments with [3H]DHE. The results obtained with the 3 methods are in good agreement. Each approach appears valid and applicable to the characterization of alpha 1- and alpha 2-adrenoceptor subtypes in rat uterus, but the method using [3H]PRAZ and [3H]RAUW demonstrates more directly the presence of the two receptor subtypes.