Background: An increasing number of studies have demonstrated that changes in metabolic levels in follicular fluid are directly related to oocyte maturation, fertilization, the quality of embryo and pregnancy rates. However, whether the intra-follicular neurotransmitters affect the function of granulosa cells and the outcome of in vitro fertilization and embryo transfer (IVF-ET) is not clear.Methods: Human follicular fluid and cumulus granulosa cells (GCs) were harvested from large follicles

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... ained from 27 patients undergoing IVF. Neurotransmitters and steroid hormones in follicular fluid were measured through liquid chromatography–tandem mass spectrometry (LC-MS/MS). Based on the content of Glutamine (Gln) in follicular fluid, the samples were divided into two groups: high Gln level group and low Gln level group. The expression of proliferation-, steroidogenesis- and antioxidant-related genes in GCs was detected by qRT-PCR. In vitro experiments, KGN cells were cultured with or without Gln and NE followed by western blotting to detect PCNA and IDH1 expression. EdU staining and ELISA were conducted to evaluate GCs proliferation and measure the contents of 17 b-estradiol (E2) and progesterone (Prog) in the conditioned medium. Primary and secondary outcomes were the number of mature and retrieved oocytes, and the ratio of high quality embryos, respectively.Results: Gln and norepinephrine (NE) were abundant neurotransmitters in follicular fluid. The contents of Gln and NE in follicular fluid exhibited a significantly positive correlation. The contents of Gln and NE in follicular fluid affected the function of granulosa cells, including their proliferative potential, E2 production and antioxidant capacity. Moreover, the concentrations of Gln and NE in follicular fluid showed significantly positive correlation with isocitrate dehydrogenase 1 (IDH1) expression in GCs. Importantly, the increased Gln and NE in follicular fluid partially affected the quality of embryonic development by regulating IDH1 expression in GCs. In vitro studies further demonstrated that Gln and NE played synergistically function in improving GCs proliferation and increasing the E2 production by upregulating IDH1 expression.Conclusions: These data demonstrate that neurotransmitter metabolic levels in follicular fluid affect the function of cumulus GCs. As important neurotransmitter metabolites, Gln and NE play significant roles in promoting GCs proliferation and increasing E2 production in granulosa cells. In addition, intrafollicular Gln and NE could upregulate the expression of the antioxidative gene IDH1 in cumulus GCs, which could be a good predictor for selecting optimal quality oocytes and evaluating the quality of embryonic development.