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Feeding N-carbamylglutamate (NCG) and arginine (ARG) improves reproductive measures in pigs and reduces systemic steroid levels in pregnant ewes. We hypothesized that the effects of NCG and ARG on reproduction were due to direct effects on the ovary. Thus, the objectives of this study were to investigate the effects of NCG and ARG on granulosa cell (GC) steroidogenesis, gene expression, and cell proliferation in vitro. GC were collected from small (1-5mm) bovine follicles and treated in vitro with NCG or ARG in serum-free medium for 24h to 48h. Both NCG and ARG inhibited (P0.05) IGF1- and FSH-induced GC estradiol production but only NCG inhibited (P0.05) progesterone production. In contrast, NCG and ARG increased (P0.05) GC numbers induced by IGF1 and FSH. NCG inhibited (P0.05) StAR, CYP11A1 and CYP19A1 mRNA abundance in small-follicle GC, whereas ARG had no effect (P0.10) on StAR, CYP11A1 or CYP19A1 mRNA abundance. We conclude that NCG and ARG may act directly on GC and therefore may regulate ovarian function by slowing follicular differentiation via inhibiting IGF1 action, and steroid synthesis while stimulating GC proliferation in cattle.

Results of in vivo studies indicate dietary N-carbamylglutamate (NCG) and arginine (ARG) can enhance reproductive performance in gilts. It was hypothesized that both NCG and ARG will alter hormone-induced estradiol (E2) production by granulosa cells (GC), explaining why these compounds could improve reproductive performance in pigs. The objective of these studies, therefore, was to evaluate the direct effects of NCG and ARG on porcine GC proliferation and steroidogenesis, using an in vitro cell culture system. The GC from small (SM; 1-5 mm) and large (LG;5 mm) pig follicles were cultured for 2 days in 5% fetal bovine serum and 5% porcine serum-containing medium followed by 2 days in serum-free medium containing 500 ng/mL of testosterone (as an E2 precursor), and NCG or ARG at various doses in the presence of either follicle-stimulating hormone (FSH; 30 ng/mL), insulin-like growth factor-1 (IGF1; 30 ng/mL), or both. Numbers of GC were determined at the end of the experiment and concentrations of progesterone (P4) and E2 in culture medium were determined. Results indicated that LG-follicle GC were more responsive to NCG and ARG than SM-follicle GC. Specifically, in LG-follicle GC, NCG inhibited (P 0.05) basal and FSH-induced P4 and E2 production but stimulated cell numbers; whereas ARG inhibited FSH-induced E2 production and cell numbers. In SM-follicle GC, treatment with NCG and ARG decreased IGF1 plus FSH induced P4 production, but E2 production and cell proliferation were not affected. These studies indicate that NCG and ARG may directly affect follicular function in pigs.