Open Access
Anthony, Kate Lordan
Graduate Program:
Animal Science
Master of Science
Document Type:
Master Thesis
Date of Defense:
November 16, 2016
Committee Members:
  • Francisco Javier Diaz, Thesis Advisor
  • Alan Leslie Johnson, Committee Member
  • Ramesh Ramachandran, Committee Member
  • ovary
  • follicle
  • chicken
  • broiler breeder hen
  • microarray
  • steroidogenesis
  • neural
  • prepubescent
  • feed
  • gene expression
  • VIP
  • vasoactive intestinal polypeptide
Modern broiler chicken lines have been selected for high growth rate and increased feed consumption. The fast growth of the broiler chick is offset by poor breeding performance of the broiler breeders (parent lines). In the adult broiler breeder hen, excessive feed consumption causes severe ovarian dysfunction and results in superfluous follicle growth and double preovulatory hierarchies, leading to reduced fertility and egg production. Modern management practices involve severe restriction of feed to help minimize reproductive dysfunction and maintain productivity. However, little is known about the effects of overfeeding or feed restriction on ovarian function in the prepubescent hen. The objective of this thesis was to identify differences in morphology, gene and protein expression within the ovarian cortex that distinguish the follicular growth environments of full-fed (FF) and restricted-fed (RF) prepubescent hens. Current work was performed on ovaries of 10-and 16-week-old-hens. Results showed, at both ages, heavier ovaries in FF hens, with a shift towards greater proportion of larger (>0.3 mm) cortical follicles and fewer smaller (<0.1 mm) cortical follicles as compared to RF. Expression of proteins and mRNA of factors known to be involved in follicular development were measured. Increased follicular development seen in FF was not associated with changes in mRNA of TGF-β ligands (AMH, BMP6, BMP15, or GDF9) or protein of their signaling transcription factors (SMAD1/5/8 or SMAD2/3). Nor were there differences in immunolocalization of the modified histone proteins associated with transcription activity; trimethylated histone H3K4, acetylated histone H3K9, or trimethylated histone H3K27. However, serine phosphorylation, which localized to the oocyte cytoplasm, revealed vesicle-like structures in the RF hens, while tyrosine phosphorylation localized more prominently to the surface of granulosa cells from RF hens. Exploration of possible pathways and genetic markers associated with follicle growth using microarray analysis of the ovarian cortex transcriptome of the 16-week-old hens uncovered differentially expressed genes (DEG) involved in steroidogenesis and neuroactive ligand-receptor interaction. Transcripts with greater abundance in the full-fed hens included those involved in cholesterol biosynthesis (HMGCR and DHCR24) and transport (STAR and STARD4), steroid biosynthesis (CYP11A1, HSD3β2, HSD17β1 and CYP19A1), and G protein-coupled receptor VIPR2. Two transcripts with less abundance in FF at 16 weeks were adenylyl cyclase inhibitors CHRM5 and DRD4. This pattern was also seen at 10 weeks but with less DEG represented in each group. DEG with greater abundance at 10 weeks included DHCR24, STAR and STARD4, CYP11A1 and CYP19A1, as well as VIPR2. One adenylyl cyclase inhibitor, DRD4, had less abundance in FF at 10 weeks. Vasoactive intestinal polypeptide (VIP) and its receptors (VIPR1 and VIPR2) had become factors-of-interest in this thesis because of their role in steroidogenesis in mammalian ovaries and in larger follicles of adult hens. But little is known about the role of VIP in the chicken cortex. The final group of experiments in this thesis focused on cortical follicle culture to test effects of VIP on STAR mRNA expression. The results showed that STAR abundance was maintained in follicles treated with VIP equal to that of fresh samples, but follicles cultured with no steroidogenic activator could not maintain STAR. Unexpectedly, abundance of VIPR2 also was not maintained, while that of VIPR1 was 5 fold higher than that of fresh in the VIP-treated follicles. The roles of these receptors within the cortex have not been determined, and most likely the expression of each changes as follicles develop, suggesting greater expression of VIPR1 during proliferation and VIPR2 during differentiation. But taken together, our observations suggest that excessive energy intake leads to increased follicle growth and abundance of transcripts involved in steroid biosynthesis and neuronal activity in ovaries of immature broiler breeder hens and that VIP promotes activation of steroidogenic transcripts in cortical follicles.