70 EXPRESSION OF NUCLEAR AND MEMBRANE PROGESTERONE RECEPTORS IN THE CANINE OVIDUCT DURING THE PERIOVULATORY PERIOD

Abstract

In the bitch, the oviduct is the site of oocyte maturation (Day 1 to 3 after ovulation), sperm transport/capacitation, fertilization (Day 3 to 4) and embryo development to the morula/blastocyst stage (Day 4 to 8). Unlike other mammals, these events occur in the presence of high (>6 ng mL^–1) and increasing plasma levels of progesterone (P4), but little is known about the regulation of oviductal functions by P4 in the bitch. The objective of this work was to study the mRNA expression of nuclear (PR) and membrane (PGRMC1, PGRMC2, mPRβ and mPRγ) P4 receptors in the canine oviduct during the periovulatory period. Thirty-six Beagle bitches were ovariectomized at 6 stages: anestrus, before the LH peak (pre-LH), after the LH peak (pre-ov) and after ovulation (Day 1, 4 and 7). Three oviductal regions were collected [i.e. ampulla, isthmus and utero-tubal junction (UTJ)]. Total RNA was extracted and then reverse transcribed. The expression of target genes was assessed in duplicate by quantitative PCR (LightCycler^® 480; Roche Diagnostics, Meylan, France) using the relative standard curve method and normalized by the geometric mean of 2 reference genes (RPS19 and GAPDH). Relative amounts of mRNA were compared between groups by ANOVA followed, when necessary, by Duncan's test. The expression of nuclear and membrane P4 receptor mRNA varied according to the stage. Expression of PR mRNA was significantly higher at pre-LH, pre-ov and Day 1 stages [means of 1.8, 1.6 and 1.5 arbitrary units (AU), respectively] than at anoestrus, Day 4 and Day 7 (1, 0.4 and 0.5 AU, respectively) in the ampulla. Same patterns of expression were observed for PR in the isthmus and UTJ. Expression of PGRMC1 and PGRMC2 mRNA were at the lowest level during anoestrus (1 AU) and increased significantly from pre-LH to Day 7 in the ampulla (from 2.2 to 8.3 AU and from 1.3 to 5.4 AU for PGRMC1 and PGRMC2, respectively) and in the isthmus (from 0.4 to 2.6 AU and from 0.5 to 1.8 AU for PGRMC1 and PGRMC2, respectively). In the UTJ, mRNA levels for PGRMC1 and PGRMC2 were the highest at Day 4 (3.9 AU) and pre-LH (2.1 AU), respectively, compared to other stages. Expression of mPRβ mRNA did not vary according to the stage in the ampulla and the isthmus, whereas it was significantly lower at Days 4 and 7 (0.6–0.7 AU) compared to other stages (1–1.2 AU) in the UTJ. Expression of mPRγ was significantly higher at Day 7 (5.0 AU) compared to other stages (0.2–1 AU) in the ampulla and was significantly higher at both anoestrus (1 AU) and Day 7 (0.9 AU) compared to other stages (0.02–0.09 AU) in the isthmus, whereas it did not vary significantly in the UTJ. In conclusion, our data suggests that P4 may be an important regulating factor of oviductal functions and could mediate its actions through genomic as well as non-genomic pathways.