Increased Gene Expression of Follicle Stimulating Hormone Receptor in Cat Cumulus-Oocyte Complexes During Seasonal Anestrus.

Abstract

Reproductive seasonality in the cat is mediated by photoperiod, with longer daylength associated with increased ovarian activity. In the northern hemisphere, estrus occurs from February through August (breeding season) followed by a seasonal anestrus from September through January (non-breeding season). We have previously observed reduced in vitro maturation and in vitro fertilization success in domestic cat oocytes recovered during the non-breeding season. Subsequently, we demonstrated that Grade 1 cumulus-oocyte complexes (COC; a homogenous dark oocyte surrounded by several layers of compacted cumulus cells) collected during this interval of seasonal quiescence require a 10 times higher follicle-stimulating hormone (FSH) concentration to achieve normal meiotic and cytoplasmic maturation in vitro. The objective of the current study was to better understand this seasonal decreased sensitivity to FSH by comparing the expression of FSH receptor (FSHR) mRNA between the breeding and non-breeding season in (1) Grade 1 COCs and (2) whole ovaries (reflecting mRNA expression of the entire intraovarian follicular population). Ovariectomies were conducted on adult cats in June and July (breeding season; Br) and November through January (non-breeding season; NBr). Within each season, ovaries (n = 6 in Br, n = 10 in NBr) were sliced to isolate Grade 1 COCs (n = 12 in Br, n = 11 in NBr) that were pooled before total RNA extraction using a Picopure RNA isolation kit. For whole ovary assessment, six ovaries (n = 3 in Br, n = 3 in NBr) from six females (1 ovary/female) were snap-frozen for total RNA extraction using Trizol reagent. cDNA was generated from COCs and whole ovarian total RNA using random primers and Superscript III reverse transcriptase. Real-time quantitative polymerase chain reaction (qPCR) was performed to compare relative expression levels of FSHR mRNA from pooled cDNA samples from the breeding and non-breeding seasons. All data were normalized to ACTB mRNA expression levels. The significance of expression ratios of FSHR mRNA was tested using the Pair Wise Fixed Reallocation Randomization Test. Results indicated a 2.8-fold increase (P < 0.01) in FSHR mRNA in Grade 1 COCs and a 1.8-fold increase (P < 0.01) in FSHR mRNA expression in whole ovaries during the non-breeding compared to the breeding season. These results suggest that increased FSHR mRNA expression in Grade 1 COCs during the non-breeding season is related to the decreased sensitivity of Grade 1 COCs to FSH previously described during the non-breeding season. A similar trend in expression in whole ovaries indicates FSHR mRNA levels also may be seasonally regulated in the entire intraovarian follicular population. Our results demonstrate for the first time that the seasonal decrease in sensitivity to FSH in intraovarian cat oocytes is not related to decreased FSHR mRNA expression. We propose that this seasonal increase in FSHR mRNA is either due to (1) an increase in expression of a truncated form of FSHR mRNA which may be translated into a non-functional FSHR or (2) an accumulation of FSHR mRNA in the cytoplasm without translation into FSHR protein. Experiments are in progress to analyze mRNA isoform and protein expression as well as FSHR functionality in the breeding versus non-breeding season. (poster)