GABAergic input through GABA_B receptors is necessary during a perinatal window to shape gene expression of factors critical to reproduction such as Kiss1

Abstract

Lack of GABAB receptors in GABAB1 knockout mice decreases neonatal ARC kisspeptin 1 ( Kiss1) expression in the arcuate nucleus of the hypothalamus (ARC) in females, which show impaired reproduction as adults. Our aim was to selectively impair GABAB signaling during a short postnatal period to evaluate its impact on the reproductive system. Neonatal male and female mice were injected with the GABAB antagonist CGP 55845 (CGP, 1 mg/kg body wt sc) or saline from postnatal day 2 ( PND2) to PND6, three times per day (8 AM, 1 PM, and 6 PM). One group was killed on PND6 for collection of blood samples (hormones by radioimmunoassay), brains for gene expression in the anteroventral periventricular nucleus-periventricular nucleus continuum (AVPV/PeN), and ARC micropunches [quantitative PCR (qPCR)] and gonads for qPCR, hormone contents, and histology. A second group of mice was injected with CGP (1 mg/kg body wt sc) or saline from PND2 to PND6, three times per day (8 AM, 1 PM, and 6 PM), and left to grow to adulthood. We measured body weight during development and parameters of sexual differentiation, puberty onset, and estrous cycles. Adult mice were killed, and trunk blood (hormones), brains for qPCR, and gonads for qPCR and hormone contents were obtained. Our most important findings on PND6 include the CGP-induced decrease in ARC Kiss1 and increase in neurokinin B ( Tac2) in both sexes; the decrease in AVPV/PeN tyrosine hydroxylase ( Th) only in females; the increase in gonad estradiol content in both sexes; and the increase in primordial follicles and decrease in primary and secondary follicles. Neonatally CGP-treated adults showed decreased ARC Kiss1 and ARC gonadotropin-releasing hormone ( Gnrh1) and increased ARC glutamic acid decarboxylase 67 ( Gad1) only in males; increased ARC GABAB receptor subunit 1 ( Gabbr1) in both sexes; and decreased AVPV/PeN Th only in females. We demonstrate that ARC Kiss1 expression is chronically downregulated in males and that the normal sex difference in AVPV/PeN Th expression is abolished. In conclusion, neonatal GABAergic input through GABAB receptors shapes gene expression of factors critical to reproduction.

GABAB Receptor Antagonism from Birth to Weaning Permanently Modifies <b><i>Kiss1</i></b> Expression in the Hypothalamus and Gonads in Mice

“…No such regulation was observed in the adult ARC or AVPV/PeN [31]. In contrast, in neonate GABAB1KO mice, a transient decrease in ARC Kiss1 expression was observed at PND4 [4], and this was validated by the injection of a GABAB antagonist to neonatal BALB/c mice [34], indicating that at this stage of development, GABABR stimulate/maintain Kiss1 expression. Regarding gonadal kisspeptin, it seems to be tightly regulated by circulating LH that stimulates its expression in both ovaries and testes [26, 49].…”

“…Efficiencies of 2 ± 0.15 were considered optimal (corresponding to 115–85%). Quantitative measurements of Kiss1r , Tac2 , Pdyn , Th , Esr1 , Esr2 , Pgr , Gnrh1 , Cyp19A1 , Gad1 , Gabbr1 , Gabbr2 , Cyp17a1 , Hsd3b2 , and Ppib2 cDNA were performed by qPCR in a total volume of 10 µL as previously described [34], whereas Kiss1 qPCR was performed in a total volume of 30 µL. Amplification was carried out in StepOne Real-Time PCR Systems (Applied Biosystems).…”

“…Furthermore, we have recently demonstrated that neonatal treatment of BALB/c mice with a GABAB antagonist from PND2 to PND6 induced downregulation of ARC Kiss1 expression on PND6 in males and females. This downregulation of ARC Kiss1 became permanent exclusively in adult males, long after treatment interruption [34].…”

Endocrine-disrupting chemicals and their effects on puberty

“…MKRN3 itself is down regulated by hypothalamic microRNA, miR-30b, whose expression increases during postnatal maturation [11]. Interestingly, GABA inhibition on GnRH release occurs also at the kisspeptin signalling level [2,12].…”

Metabolomic Analysis of Plasma from GABAB(1) Knock-Out Mice Reveals Decreased Levels of Elaidic Trans-Fatty Acid

“…Therefore, the strain of animal used in our laboratories is different from the original [7] due to the crossing with the FVB strain. Over the years, our colony of mice was maintained by crossing HZ mice since GABA B(R1) knock-out mice are sterile and therefore, to obtain KO mice, we need to cross HZ ± GABA B(R1) mice [22]. The mice used in the present study derived from independent mating of the F12-F16 generation.…”