Abstract

Polycystic ovarian syndrome (PCOS) is the most common cause of infertility in women of reproductive age worldwide. Women with PCOS exhibit the neuroendocrine phenotype of increased luteinizing hormone (LH) pulse frequency, which drives the downstream symptoms of anovulation, cyst formation and hyperandrogenism at the ovary. LH hypersecretion is indicative of impaired central steroid hormone feedback to gonadotropin‐releasing hormone (GnRH) neurons, the final output cell in a large network controlling fertility. Steroid hormone feedback control of GnRH neuron activity and GnRH/LH pulsatile release are hypothesized to be regulated by a population of neurons in the arcuate nucleus (ARC) that co‐express the peptides Kisspeptin, Neurokinin B and Dynorphin, commonly known as KNDy neurons. Therefore, we hypothesized that changes in the regulation of KNDy neuron activity could lead to elevated GnRH neuron activity and GnRH/LH hypersecretion in PCOS. To investigate this hypothesis, we used a prenatal androgen‐induced mouse model of PCOS to determine whether synaptic input to KNDy neurons from the major neurotransmitters GABA and glutamate is altered. Pregnant proDynorphin‐Cre/yellow fluorescent protein (pDyn‐Cre/YFP) transgenic mice were injected subcutaneously with dihydrotestosterone or an oil vehicle on days 16, 17 and 18 of pregnancy. Female PNA (n=4) and prenatal vehicle‐treated (PNV, n=4) pDyn‐Cre/YFP offspring were studied at 60–80 days of age during adult life. Triple‐label immunofluorescence was performed in coronal brain sections for YFP, the vesicular glutamate transporter 2 (vGluT2) and the vesicular GABA transporter (vGaT) to enhance YFP in KNDy neurons and visualize glutamatergic and GABAergic terminals, respectively. The soma of twenty KNDy neurons were analyzed per animal using confocal microscopy. PNA treatment had no effect on the number and density of vGluT2 appositions to KNDy neurons, indicating glutamatergic input is not altered in PNA mice. However, the number (p<0.01) and density (p<0.01) of vGaT appositions to KNDy neurons was significantly reduced in PNA mice when compared to controls. These results indicate that PNA treatment reduces GABAergic input to KNDy neurons. As GABA is classically an inhibitory neurotransmitter, this may lead to increased KNDy neuron firing in PNA mice that drives higher GnRH neuron activity and GnRH/LH pulsatile release, replicating the PCOS neuroendocrine phenotype. The location and phenotype of GABAergic neurons with reduced input to KNDy neurons in PNA mice is so far unknown and deserves further study.