REPRODUCTION RESEARCH Increased potency of a1-adrenergic receptors to induce inositol phosphates production correlates with the up-regulation of a1d/Gha/phospholipase Cd1 signaling pathway in term rat myometrium M Dupuis, E Houdeau 1 and S Mhaouty-Kodja CNRS UMR 7079, Laboratoire de Physiologie et Physiopathologie, 75231 Paris Cedex 05, France and 1 INRA UMR 1054, Unite ´ de Neuro-Gastroente ´rologie and Nutrition, 31931 Toulouse Cedex 09, France Correspondence should be addressed to S Mhaouty-Kodja who is now at CNRS UMR 7148, Colle `ge de France, 11 place Marcelin Berthelot, 75231 Paris Cedex 05, France; Email: sakina.mhaouty-kodja@college-de-france.fr M Dupuis is now at Rudolf Virchow Center, DFG Research Center for Experimental Biomedicine, University of Wuerzburg, Wuerzburg, Germany Abstract In the present study, we studied the potential regulation by rat myometrial a1-adrenergic receptors (a1-AR) of the newly identified Gha protein/phospholipase Cd1 (PLCd1) signaling pathway and compared myometrial inositol phosphates (InsP) production and activity of the uterine circular muscle in response to a1-AR activation between mid-pregnancy and term. For this, we quantified the level of rat myometrial a1-AR coupling to Gha protein by photoaffinity-labeling, the cytosolic amount of PLCd1 enzyme by immunoblotting, and the expression level of a1-AR subtypes by RT-PCR. The results showed an increased level of a1-AR/Gha protein coupling and the amount of PLCd1 at term (C147 and C65% respectively, versus mid-pregnancy). This was correlated with an up-regulation of a1d-AR subtype (C70% versus mid- pregnancy). Incubation of myometrial strips with phenylephrine (Phe), a global a1-agonist, increased InsP production in a dose-dependent manner at both mid-pregnancy and term, but with an enhanced potency (tenfold decrease in EC 50 value) at term. Phe also dose-dependently induced contraction of the circular muscle at both mid-pregnancy and term. However, unlike InsP response, no amelioration of potency was observed at term. Similar results were obtained with the endogenous agonist norepinephrine. Our results show, for the first time, that rat myometrial a1d-AR/Gha/PLCd1 signaling pathway is up-regulated at term. This is associated with an increased potency of a1-AR to elicit InsP production but not uterine contraction at this period. It is thus hypothesized that a1-AR, through activation of Gha/PLCd1 system, are not primarily involved in the initiation of labor but may rather regulate responses such as myometrial cell proliferation or hypertrophy. Reproduction (2008) 135 55–62 Introduction It is well established that catecholamines (norepinephrine, epinephrine) regulate uterine contractility. The first evidence came from Ru ¨sse and Marshall’s works, which showed that norepinephrine, through activation of a-adrenergic receptors (a-AR), enhances the effects of uterotonic agents by improving the excitability of the uterine muscle in the guinea pig (Russe & Marshall 1970). Moreover, Legrand and Maltier have reported evidences for a noradrenergic transmission in the control of both pregnancy and parturition in rat. For instance, adminis- tration of propranolol, a b-AR antagonist, advanced pregnancy (Maltier & Cavaille ´ 1975). In contrast, adminis- tration to near term rats of prazosin, a powerful a-AR antagonist, reduced myometrial excitability and delayed parturition (Legrand & Maltier 1986). Norepinephrine signals through nine different G-protein-coupled ARs divided into three subfamilies, namely a1(a1a, a1b, a1d), a2(a2a, a2b, a2c), and b (b1, b2, b3). Using specific radioligands, we have shown that b- and a2-AR are co-localized in the longitudinal muscle, whereas a1-AR are expressed in the circular layer of rat myometrium (Legrand et al. 1991, 1993). Co-activation of b- and a2-AR increases myometrial cAMP production at mid-pregnancy (Mhaouty et al. 1995), thereby resulting in a relaxation of the longitudinal uterine smooth muscle. Near term, progesterone withdrawal associated with increase in estradiol concentrations trigger desensitization of b-adrenergic signaling pathway (Cohen-Tannoudji et al. 1991) and a2-AR shift to inhibitors of adenylyl cyclase (Mhaouty et al. 1995). These events probably participate in the initiation of labor. q 2008 Society for Reproduction and Fertility DOI: 10.1530/REP-07-0332 ISSN 1470–1626 (paper) 1741–7899 (online) Online version via www.reproduction-online.org