A Quantitative Ultrastructural Study of Oocytes During the Early Stages of Ovarian Follicular Development in Booroola and Wild-Type Sheep
In the sheep ovary, primordial follicles are formed as an oocyte surrounded by either a single layer of flattened granulosa cells (type 1) or a mixture of flattened and cuboidal granulosa cells (type 1a). Booroola sheep have a mutation in the growth factor receptor, activin-like kinase receptor 6 (ALK6) which is expressed in oocytes of follicles at the type 1 stage of development. In Booroola ewes homozygous for the ALK6 mutation (BB), oocytes undergo precocious maturation that appears to be initiated during the preantral growth phase. The aim of this study was to quantify the ultrastructural features of oocytes of ovarian follicles at the types 1/1a, 2 and 3 stages of development, from BB and wild-type (++) ewes. Ovaries from 6 ++ and 5 BB 4 week old ewe lambs were processed for both light microscopy (LM) and electron microscopy (EM). LM and stereological methods were used to estimate the mean volume of the oocytes of each follicular type and genotype. EM images and point counting or linear intercept counting were used to estimate the volume of smooth endoplasmic reticulum (SER), Golgi, mitochondria, vesicles, lipid, ribosomes, zona pellucida (ZP) and cortical granules (CG), and the surface area of the outer and inner mictochondrial membranes, microvilli and cell junctions. Oocytes of type 1/1a follicles of BB animals had a greater diameter than that in ++ animals (BB: 29.76 ± 0.58 μm vs ++: 27.05 ± 0.30 μm; P < 0.01) but there were no significant genotype differences in the oocyte diameters of type 2 or 3 follicles. As the follicles of ++ animals developed from the type 1 through to the type 3 stage, the volume and 3 surface areas of all sub-cellular structures measured within oocytes increased (P < 0.05). In oocytes of type 1/1a follicles of BB animals, the SER, mitochondria and ZP volumes were greater than in ++ animals (P < 0.05) as were the surface areas of the outer mitochondrial membranes, oocyte membrane, and zonula adherens type junctions (P < 0.05). At the type 2 stage of development the lipid volume was greater in oocytes of ++ animals, and at the type 3 stage of development the ribosomal volume was greater in oocytes of BB animals. These results suggest that the ALK6 mutation in BB animals has influenced the ultrastructural properties of oocytes in the type 1/1a follicle. This early genotype difference in follicular characteristics may influence the rate of follicular development during the early developmental stages.