Progress 10/01/05 to 09/30/06
This project examined expression of circadian clock gene transcripts in the ovary of the domestic hen. Because the avian ovary essentially functions as a semi-independent circadian clock, it was pertinent to examine whether the basic molecular mechanisms that govern function of other vertebrate clocks was present and functional in the avian ovary. Because studies of the avian circadian clock genes using quantitative real-time PCR had never been reported, we first designed unique primer sets for three canonical clock genes, Bmal1, Period (Per) and Cryptochrome 1 (Cry1). At the initiation of these studies, the genome of the domestic hen was not yet published, so primers were designed from conserved regions of the genes by comparison of their human, mouse, rat and Drosophila counterparts. We were successful in producing primer sets to identify transcripts for each of the genes. Samples were collected from adult cycling hens at 6 h intervals over the course of a 24 h
period. As controls, we collected samples from the primary clock in the suprachiasmatic nucleus, as well as from the pineal and liver. 24-h oscillations in Per, Bmal1 and Cry1 were demonstrated in all tissues sampled. Importantly, Per and Bmal1 ocillations were out-of-phase with each other, suggesting that a functional clockwork may be present in the ovary. As predicted from the mammalian literature, the phase of the clockwork oscillatory behavior shifted several hours in the peripheral tissues (ovary, liver, pineal) compared to the brain. We next examined whether luteinizing hormone (LH) might act as a factor to set the phase of the ovarian clock. LH did alter expression of both Per and Bmal1, suggesting that endocrine regulatory elements are important in regulating the synchrony of the ovarian clock with the primary clock in the brain. These finding are currently being prepared for publication. These findings demonstrate for the first time that an ovarian clock is present and may be
an important component in the endocrine regulation of the ovary. Alterations of the clockwork mechanism, through light therapy may provide a means to increase reproductive efficiency in the hen.
The impact of the circadian timing system on organismic health is only beginning to be appreciated. Our studies demonstrate that luteinizing hormone regulation of an ovarian clock provides timing cues that are essential for the efficient production of fertilizable ova.
- Howell, R.E., Bahr, J.M. and Tischkau, S.A. 2006. Characterization of the avian circadian system using Bmal1, Cry1, Per, and the effect of luteinizing hormone. Annual Meeting of the Society for the Study of Reproduction, Omaha, NE.