(154) Heat-Induced Hyperthermia After a GnRH-Induced LH Surge Impacts Circulating and Periovulatory Intrafollicular Metabolites in Lactating Dairy Cows

Rebecca R. Payton¹, Sarah E. Moorey¹, Shawn R. Campagna², F. Neal Schrick¹, Ky G. Pohler³, and J. Lannett Edwards¹

1. Department of Animal Science, The University of Tennessee Institute of Agriculture and AgResearch, Knoxville, TN, USA, 37996

2. Department of Chemistry, The University of Tennessee, Knoxville, TN, USA, 37996

3. Department of Animal Science, Texas A & M University, College Station, TX, USA 77843

Abstract Text:

Impacts of varying levels of hyperthermia on the periovulatory follicle after the LH surge include components (i.e., follicular fluid proteome and cumulus/granulosa cell transcriptomes) known to potentiate ovulation, corpus luteum formation, and influence developmental competence of the maturing oocyte within. The aim of this study was to examine periovulatory follicular fluid and circulating metabolite levels in hyperthermic cows, mindful that temperature-related changes in serum may or may not influence follicular fluid composition. Cows were transported to a climate-controlled facility 35 h after PGF_2α administration.  After GnRH administration (40 h after PGF_2α) to induce an LH surge, THI (temperature-humidity index) was steadily increased for ~12 h to induce hyperthermia (final THI 83.2 ± 0.2). The average rectal temperature of hyperthermic cows ranged from 38.9 to 40.2°C (overall average 39.5 ± 0.1°C).  Thermoneutral cows were maintained at a THI of 65.8 ± 0.1; rectal temperatures ranged from 38.2 to 38.7°C and averaged 38.4 ± 0.1°C. After ~12 h, hyperthermic cows were abruptly returned to thermoneutral conditions.  Periovulatory follicle fluid was aspirated 16 h (average 16.4 h; range 15.3 - 18.2 h) after GnRH (E2:P4 ratio: 1.91 ± 0.27; follicle diameter: 16.7 ± 0.5 mm) and blood was obtained via coccygeal venipuncture. Serum and follicular fluid samples from 27 cows (12 hyperthermic and 15 thermoneutral) were submitted for ultra-high performance liquid chromatography-high resolution mass spectrometry metabolomic analysis.  Mixed model hierarchical regression was performed with metabolite abundance as the dependent variable. Using the mean rectal temperature of individual cows as the independent variable, best fit models were obtained by backwards manual selection after ensuring lack of collinearity among regressors (e.g., follicular fluid and serum hormone levels, follicle sizes, days in milk, milk yield, and parity) and data normality. A total of 67 unique metabolites were identified. In serum, 37 of 39 metabolites were negatively related to varying levels of hyperthermia.  In the follicular fluid, 17 metabolites were related to hyperthermia (P < 0.05); all but one decreased with increasing rectal temperature. Twelve follicular fluid metabolites (70%) were also related to hyperthermia in the serum. However, heat-related effects on five others were noticed in the follicular fluid but not serum.  Serum metabolites were enriched for glycine and serine metabolism (P = 0.0025, FDR = 0.0025). Methionine metabolism (P = 0.0151, FDR = 0.0085), homocysteine degradation (P = 0.0169, FDR = 0.0085) and ammonia recycling (P = 0.0582, FDF = 0.0195) were overrepresented pathways in follicular fluid. Serine/glycine metabolism provides building blocks for rapidly dividing cells and maintains redox status. Intrafollicular methionine relates to fertility outcomes and is necessary for redox maintenance while homocysteine, a degradation product of methionine, plays important roles in cellular growth and oxidative stress. Interestingly, the glycine/serine pathway in serum feeds into the homocysteine and methionine pathways altered in follicular fluid, suggesting heat-induced changes in circulating metabolites directly impact the follicular fluid milieu. Similarly, follicular fluid estradiol was a significant regressor for >90% of serum metabolites indicating influence of intrafollicular constituents on circulating molecules.  Cumulatively, findings provide important insight regarding how varying degrees of hyperthermia induce functional changes in systemic and periovulatory follicular fluid metabolomes. This project was supported by the state of Tennessee through UT Institute of Agriculture, AgResearch, the Department of Animal Science, and the USDA National Institute of Food and Agriculture, Multistate Project No. NE2227.