Differential expression of mRNAs encoding the putative inhibin co-receptor (betaglycan) and activin type-I and type-II receptors in preovulatory and prehierarchical follicles of the laying hen ovary

Abstract

Ovarian follicle development is primarily regulated by an interplay between the pituitary gonadotrophins, LH and FSH, and ovary-derived steroids. Increasing evidence implicates regulatory roles of transforming growth factor-β (TGFβ) superfamily members, including inhibins and activins. The aim of this study was to identify the expression of mRNAs encoding key receptors of the inhibin/activin system in ovarian follicles ranging from 4 mm in diameter to the dominant F1 follicle (~40 mm). Ovaries were collected (n=16) from mid-sequence hens maintained on a long-day photoschedule (16 h of light:8 h of darkness). All follicles removed were dissected into individual granulosa and thecal layers. RNA was extracted and cDNA synthesized. Real-time quantitative PCR was used to quantify the expression of mRNA encoding betaglycan, activin receptor (ActR) subtypes (type-I, -IIA and -IIB) and glyceraldehyde-3-phosphate dehydrogenase (GAPDH); receptor expression data were normalized to GAPDH expression. Detectable levels of ActRI, -IIA and -IIB and the inhibin co-receptor (betaglycan) expression were found in all granulosa and thecal layers analysed. Granulosa ActRI mRNA peaked (P < 0.05) in 8–9.9 mm follicles, whereas ActRIIA rose significantly from 6–7.9 mm to 8–9.9 mm, before falling to F3/2; levels then rose sharply (3-fold) to F1 levels. Granulosa betaglycan mRNA expression rose 3-fold from 4–5.9 mm to 8–9.9 mm, before falling 4-fold to F3/2; levels then rose sharply (4-fold) to F1 levels. ActRIIB levels did not vary significantly during follicular development. Thecal ActRI mRNA expression was similar from 4–7.9 mm then decreased significantly to a nadir at the F4 position, before increasing 2-fold to the F1 (P < 0.05). Although thecal ActRIIB and -IIA expression did not vary significantly from 4 mm to F3, ActRIIB expression increased significantly (2-fold) from F3 to F1 and ActRIIA increased 2-fold from F2 to F1 (P < 0.05). Thecal betaglycan fell to a nadir at F6 after follicle selection; levels then increased significantly to F2, before falling ~50% in the F1. In all follicles studied expression of betaglycan and ActRI (granulosa: r=0.65, P < 0.001, n=144/group; theca: r=0.49, P < 0.001, n=144/group) was well correlated. No significant correlations were identified between betaglycan and ActRIIA or -IIB. Considering all follicles analysed, granulosa mRNA expression of betaglycan, ActRI, ActRIIA and ActRIIB were all significantly lower than in corresponding thecal tissue (betaglycan, 11.4-fold; ActRIIB, 5.1-fold; ActRI, 3.8-fold; ActRIIA, 2.8-fold). The co-localization of type-I and -II activin receptors and betaglycan on granulosa and thecal cells are consistent with a local auto/paracrine role of inhibins and activins in modulating ovarian follicle development, selection and progression in the domestic fowl.