|Cushman, Robert - Bob|
Submitted to: Biology of Reproduction Abstracts
Publication Type: Abstract Only
Publication Acceptance Date: 4/17/2009
Publication Date: 12/1/2009
Citation: Cushman, R.A., Echternkamp, S.E. 2009. Influence of Reproductive Aging of the Cow on Luteal Function and Period 1 mRNA Expression [abstract]. Biology of Reproduction. 2009 Supplement:156 (Abstract #478).
Technical Abstract: In rodents, disruption of the circadian clock genes results in increased incidence of anovulation, irregular estrous cycles, decreased luteal function, and accelerated reproductive ageing. In cattle, reproductive ageing is associated with decreased numbers of follicles in the ovary, decreased luteal function and increased risk of anovulation. The clock gene, Period 1, is expressed in the rodent corpus luteum and may control steroidogenesis. Therefore, we tested the hypotheses that Period 1 mRNA is expressed in the bovine corpus luteum and that luteal Period 1 mRNA levels decrease as part of the reproductive ageing process of cows. Ovaries were collected from young (= 6 yrs; n = 6) and old (> 6 yrs; n = 9) crossbred beef cows by colpotomy on days 3, 6, or 9 after estrus. Immediately prior to surgery, a blood sample was collected by caudal venipuncture and processed for determination of serum progesterone concentration by radioimmunoassay. After removal, ovaries were transported to the laboratory where surface follicles were counted and ovaries were weighed. Corpora lutea were dissected, weighed, and frozen in liquid nitrogen. Total cellular RNA was extracted from luteal tissue and diluted to 50 ng/microliter. Relative levels of messenger RNA for Period 1, 3ß-hydroxysteroid dehydrogenase (3ß-HSD), and GAPDH were determined using previously validated primers in a one-step real-time RT-PCR reaction with 100 ng of tcRNA. Ovarian morphometric data were analyzed using the GLM procedure of SAS with age (young vs. old) as a fixed effect and day of the estrous cycle as a covariate. Relative levels of Period 1 and 3ß-HSD mRNA were analyzed using the GLM procedure of SAS with age (young vs. old) as a fixed effect and day of the estrous cycle and relative levels of GAPDH as covariates. There was no difference in antral follicle number between young and old cows (42.0 ± 10.2 vs. 48.5 ± 8.2 follicles, respectively; P = 0.64); however, luteal weight was decreased in old cows (3.5 ± 0.2 vs. 2.8 ± 0.2 g; P = 0.01) and serum progesterone concentrations tended to be decreased in old cows (4.2 ± 0.3 vs. 3.4 ± 0.4 ng/ml; P = 0.1) when compared to young cows. Serum progesterone concentrations were positively associated with luteal weight and day of the estrous cycle (P < 0.0001). Luteal mRNA levels for 3ß-HSD did not differ between age groups or days of the estrous cycle. However, luteal Period 1 mRNA levels tended to be greater in young cows than in old cows (823.9 ± 179.8 vs. 302.6 ± 223.4 relative units; P = 0.1) and increased with day of the estrous cycle (P = 0.04). From these data, we conclude that reproductive ageing results in decreased luteal function in cows as observed by a trend for decreased serum progesterone concentrations that agrees with previously published data in women and cows. Decreases in serum progesterone concentrations were not due to decreased levels of 3ß-HSD mRNA. Novel findings of this study were that 1) Period 1 mRNA is expressed in the bovine corpus luteum, 2) luteal Period 1 mRNA levels increase during luteal development, and 3) luteal Period 1 mRNA levels tend to decrease with cow age. The exact mechanisms by which circadian clock genes influence steroidogenesis in mammalian corpora lutea remain to be elucidated.