MON-222 Blocking GABAB Receptors (GABABR) from Birth to Weaning in Mice Induces Profound Changes in the Gonadotropic Axis

Abstract

We have previously shown that the administration of a GABAB antagonist (CGP55845) to neonatal BALB/C mice from postnatal day (PND) 2-6 significantly decreased arcuate nucleus (ARC) kisspeptin (Kiss1) expression in both sexes. Furthermore, this neonatal CGP55845 treatment delayed puberty onset in females and altered gonadotropin secretion in both sexes in adulthood. Here our aim was to induce a more sustained inhibition of GABABR signaling and evaluate its effects on the gonadotropic axis. Neonatal Balb/c male (M) and female (F) mice were injected with CGP55845 (1 mg/kg, sc, CGP) or saline (CTRL) from PND2-21, three times/day (8AM, 1PM, 6PM) and sacrificed at 3PM (after two injections on the last day). Serum samples and gonads were collected for hormonal measurements by RIA. Brains were frozen and 500 µm slices were obtained on a cryostat. ARC and antero ventral periventricular nucleus (AVPV) micropunches were obtained. Kiss1, kisspeptin receptor (Kiss1r), prodynorfin (Pdyn), neurokinin B (Tac2), tyrosine hydrolxylase (TH), progesterone receptor (Pgr), GABAB1 and GABAB2 subunits of the GABABR (GABAB1, GABAB2) mRNA expression was assessed in the micropunches by qPCR (control gene: Cyclophilin B). Body weight (BW) and AGI (anogenital distance/BW) were evaluated on PND 7, 14 and 21. In the ARC CGP induced a significant decrease in Kiss1 expression (FCTRL: 1,16±0.24 vs FCGP: 0,67±0.16 vs MCTRL: 1,22±0.24 vs MCGP: 0,70±0.13, CGP vs CTRL p<0,04) and an increase in Pdyn (FCTRL: 0,97±0.26 vs FCGP: 1,77±0,49 vs MCTRL: 1,37±0.32 vs MCGP: 3,01±0.85, CGP vs CTRL: p<0,02) and GABAB1 (FCTRL: 0,42±0.08 vs FCGP: 1,37±0.43 vs MCTRL: 0,74±0.13 vs MCGP: 1,38±0.46, CGP vs CTRL: p<0,04) in both sexes, while a decrease in Tac2 was only observed in males (MCTRL: 1,01±0.14 vs MCGP: 0,45±0.045: p<0,02); no changes were observed in Kiss1r or Pgr expression. In AVPV no CGP-induced changes in Kiss1,Kiss1r, TH, Pgr, GABABR expression were detected. FSH was significantly decreased in CGP-treated males (FSH (ng/ml): MCTRL: 9,3±1,28 vs MCGP: 5,78±0.9: p<0,003). BW was significantly decreased by CGP on PND21 in females while this parameter was decreased at all ages studied in males. CGP significantly decreased AGI in both sexes. These results demonstrate that sustained inhibition of GABABR signaling during the critical postnatal period of development and maturation of the gonadotropic axis profoundly alters key ARC gene expression such as Kiss1, Pdyn, Tac2, altering sexual differentiation (AGI) and BW. In addition, in males FSH levels are decreased to female levels in agreement with feminization of AGI. (Supported by CONICET, ANPCYT, UBA, Fundación René Barón, Fundación Williams).