alpha 2A- and alpha 2B-adrenoreceptors (AR), identified by Northern blotting in rat myometrium, showed a differential expression during the course of pregnancy. Indeed, the alpha 2A-AR transcript was present at mid-pregnancy, whereas high levels of alpha 2B-AR mRNA could be detected at term. The role of these subtypes in modulating beta 2-AR-stimulated adenylyl cyclase activity was investigated on myometrial membranes from mid-pregnancy and term. At nanomolar concentrations of clonidine (full alpha 2-AR agonist) or oxymetazoline (partial alpha 2A-AR agonist), adenylyl cyclase activity was inhibited by up to 50 +/- 7% at mid-pregnancy or 75 +/- 7% at term, whereas at micromolar concentrations, alpha 2-AR agonists potentiate adenylyl cyclase activity by 140-170% at mid-pregnancy. Both inhibitory and stimulatory components of this biphasic response were blocked by yohimbine, a selective alpha 2-AR antagonist. Preincubation of myometrial membranes with Gi2 and/or Gi3 antisera eliminated alpha 2-AR mediated attenuation or potentiation of isoproterenol-stimulated adenylyl cyclase, thus indicating that both the inhibitory and stimulatory components are mediated via Gi2 and Gi3. In addition, type II and IV adenylyl cyclases were identified by Northern blotting in the pregnant rat myometrium. Altogether these data strongly suggest that the alpha 2A-AR at mid-pregnancy potentiates adenylyl cyclase types II and IV through beta gamma released from Gi2 and Gi3 proteins, whereas the alpha 2B-AR expression at term may be related to persistent inhibition.

The possibility that progesterone or estradiol may regulate expression of G protein in the rat myometrium during the course of pregnancy has been investigated using 1) immunoblot analysis of Gi2 alpha, Gi3 alpha, and Gq alpha subunits and 2) hybridization blot analysis of subunit mRNA. Eighteen hours after administration, estradiol had significantly increased the levels of both Gi2 alpha subunit and Gi2 alpha mRNA (by 40% and 32%, respectively). In control pregnant rats, we observed similar changes at the end of pregnancy, when myometrial concentrations of estradiol had increased, i.e., a 41% increase in immunoreactive Gi2 alpha subunit that correlated with a parallel 45% increase in mRNA levels. In contrast, levels of immunoreactive Gi3 alpha subunit and mRNA, which decreased with advancing gestation, were not influenced by estradiol or progesterone administration. Progesterone administration resulted 30 h later in a significantly decreased level of Gq alpha immunoreactivity (32%) and Gq alpha mRNA (30%). In control rats, Gq alpha protein and mRNA were also significantly lower at midpregnancy under progesterone dominance vs. term. At this stage, a twofold increase in Gq alpha subunit correlated with a 40% increase in mRNA levels. These results demonstrate that myometrial Gi2 alpha and Gq alpha subunits are physiological targets for estradiol and progesterone, respectively, in vivo. Alterations of these G protein levels are discussed in relation to their mediating effects on adenylyl cyclase activity or the phospholipase C pathway during the course of pregnancy.