(283) Follicular Fluid Insulin, Glucose, and their Correlation with TCA Cycle and Purine Metabolites are Positively Related to Bovine Preovulatory Follicle Maturity

Abstract Authors: Bailey M. Chitwood1, Emma A. Hessock1, J. Lannett Edwards1, Rebecca R. Payton1 , Shawn R. Campagna2 , F. Neal Schrick1, and Sarah E. Moorey1
1 Department of Animal Science, University of Tennessee, Knoxville, TN 37996
2 Department of Chemistry, University of Tennessee, Knoxville, TN 39776

Abstract Text: Physiological maturity of the preovulatory follicle, based on follicle size and estradiol production, is positively associated with bovine oocyte metabolism and competence for embryo development. There is also a positive relationship between follicle maturity and abundance of follicular fluid metabolites involved in glucose metabolism and downstream pathways, but the relationship between follicle maturity and follicular fluid glucose or insulin bioavailability has yet to be examined. Insulin enhances glucose uptake into follicular cells, which then metabolize glucose to generate ATP and other metabolites required to meet the energy demands of the cells themselves and the oocyte. We hypothesized that follicular fluid from greater maturity follicles would have increased bioavailability of glucose and insulin, and that both molecules would be associated with follicular fluid metabolites involved in glucose, purine, and methionine metabolism. The objective was to determine if there was a difference in glucose and insulin concentration in preovulatory follicles of greater and lesser maturity and determine if their correlation to metabolites differed based on follicle maturity. Follicular fluid samples were collected from preovulatory follicles of synchronized Angus cows (n = 136) 19.0 ± 1.71 hours following gonadotropin-releasing hormone administration to induce the preovulatory luteinizing hormone surge (GnRH2). Follicles were categorized as greater or lesser maturity based on serum estradiol at the time of GnRH2 (greater: n = 17, 11.8 ± 0.37 pg/mL; lesser: n = 12, 3.7 ± 0.32 pg/mL). Intrafollicular glucose was measured using a bioluminescent assay, insulin using a bovine specific ELISA, and metabolome profiles generated using liquid chromatography coupled mass spectrometry. Glucose and insulin concentration were compared between follicular fluid from follicles of greater or lesser maturity using analysis of variance. Pearson’s correlation was used to determine the correlation between glucose and insulin with 16 metabolites chosen for their roles in the TCA cycle, pentose phosphate pathway, and methionine, purine, or pyrimidine metabolism. Follicular fluid insulin concentration in follicles of greater maturity was more than 5 times higher compared to that of lesser maturity (P = 0.045; 0.042 ± 0.01 ng/mL vs 0.0073 ± 0.00075 ng/mL, respectively). Similarly, glucose concentration in follicles of greater maturity was over 1.75 times higher than lesser maturity follicles (P = 0.0045; 3.21 ± 0.30 mM vs 1.82 ± 0.28 mM, respectively). While pyruvate, alpha-ketoglutarate, aconitate, citrate/isocitrate, glucose phosphate, and GMP were positively correlated to glucose in greater and lesser maturity follicles (P < 0.05, r > 0.50), insulin’s correlation with metabolites differed based on follicle maturity status. Glutamine, aconitate, glucose phosphate, inosine, and GMP were positively correlated to insulin in follicular fluid of greater maturity follicles (P < 0.05, r > 0.66), but lost correlation in follicles of lesser maturity (P > 0.45). Interestingly, aspartate was not correlated to insulin in follicles of greater maturity (P = 0.22), but gained correlation in lesser maturity follicles ( P = 0.05, r = 0.95).This study demonstrates that greater follicular maturity is associated with higher levels of glucose and insulin bioavailability,which positively correlate with key metabolites involved in energy production, further supporting that greater preovulatory follicle maturity enhances metabolic support of oocyte competence for embryo development. This project was supported by Agriculture and Food Research Initiative Competitive Grant no 2023-67015-39454 from the US Department of Food and Agriculture, USDA Multistate project NE2227, and The University of Tennessee AgResearch and Department of Animal Science.