Tachykinin receptor and neutral endopeptidase gene expression in the rat uterus:: Characterization and regulation in response to ovarian steroid treatment

Tachykinin neuropeptides, such as substance P, are localized to a population of sensory fibers that innervate the mammalian female reproductive tract. In the present study, we have characterized tachykinin NK1 receptor (NK1R), NK2 receptor (NK2R), and NK3 receptor (NK3R) gene expression by semiquantitative RT-PCR in uteri from ovariectomized rats and studied their regulation in response to 17 beta-estradiol (E-2), progesterone (P-4), or a combination of both. In addition, are analyzed the expression and regulation of the neutral endopeptidase 24.11 (NEP), the most important enzyme involved in tachykinin degradation in the rat uterus. In uteri fi om control (olive oil-treated) rats, RT-PCR assays revealed single bands corresponding to the expected product sizes encoding complementary DNA for NK1R (232 bp), NK2R (491 bp), NK3R (325 bp), and NEP (221 bp). The identity of the amplified fragments was confirmed by DNA sequence analysis. Compared with control rats, NK1R messenger RNA (mRNA) was increased by 2-fold in uteri from rats treated with E-2, was decreased by 3.3-fold in rats treated with P-4, and was decreased by 1.8-fold in rats treated with both E-2 and P-4. Uterine NK2R mRNA levels were not altered by any steroid treatment. E-2 treatment decreased by 15-fold NK3R mRNA. P-4 was without effect if administered alone and did not influence the E-2-induced decrease in NK3R mRNA. NEP mRNA levels were about 4-fold lower in E-2-treated than in P-4-treated rats. Functional studies were carried out in uteri from E-2- or P-4-treated ovariectomized rats to characterize the contractile response evoked by the selective tachykinin receptor agonists [Sar(9)Met(O-2)(11)]substance P (NK1R selective), [Nle(10)]NKA-(4-10) (NK2R selective), and [MePhe(7)]NKB (NK3R selective) in the presence of the NEP inhibitor phosphoramidon (1 mu M). A marked correlation was observed between the magnitude of the contractile response to each agonist and the level of expression determined by RT-PCR for each tachykinin receptor. The present findings show that tachykinin NK1R, NK2R, NK3R, and NEP are expressed in the rat uterus and that ovarian steroids differentially regulate their expression.