G. Dijkstra

Immunohistochemical localization and mRNA expression of activin, inhibin, follistatin, and activin receptor in bovine cumulus‐oocyte complexes during in vitro maturation

The aim of this study was to investigate whether bovine cumulus‐oocyte complexes (COCs) synthesize activin A, inhibin, and follistatin and whether they contain activin receptor during in vitro maturation. Therefore, COCs obtained from small and medium‐sized follicles were cultured in M‐199 supplemented with 10% fetal calf serum (FCS) and gonadotropins for 24 hr. At 0, 6, 12, and 24 hr after the onset of culture, COCs were removed for immunohistochemical staining to detect the expression of activin A, inhibin, follistatin, and activin receptor type II proteins. At 0 and 24 hr, COCs were removed and prepared for reverse‐transcriptase polymerase chain reaction (RT‐PCR) to assess the presence of mRNA of these proteins. It appeared that cumulus cells and oocytes express activin, follistatin, and activin receptor proteins as well as their mRNA. While expression of inhibin mRNA was found exclusively in cumulus cells, the inhibin protein was present in cumulus cells and oocytes. Immunohistochemical study both in cumulus cells and in oocytes often showed a moderate and strong staining intensity for activin and follistatin, respectively. Activin staining underwent little or no change during culture except at 24 hr of maturation, where about 60% of the oocytes showed no staining. Follistatin immunoreactivity remained strong in the majority of COCs. At the onset of culture, a spotlike inhibin staining was observed in the oocyte, which increased after 12 hr and was absent at the end of culture. Activin receptor immunoreactivity in cumulus cell membranes and oolemma increased during oocyte maturation to maximum values at the end of culture in most of the COCs. It is concluded that the consistent presence of activin and the increase in activin receptor in cumulus cells and oocytes during in vitro maturation indicate a paracrine and/or autocrine action for activin on bovine oocyte maturation. This action may be modulated by inhibin and/or follistatin. Mol. Reprod. Dev. 49:186–195, 1998.

Penile bulb and its relationship with the pelvic urethra and the penile urethra in the rat: light and scanning electron microscopical observations.

In male rats the urethral tract consists of the following parts: the pelvic urethra, the transitional urethra, the urethral diverticle, and the penile urethra. Perusal of the literature results in only some more general descriptions and a very few which go more into detail. None of the latter deals with all the compartments nor with the relationships between each other. Aim of this study is to give a detailed morphological study of all the urethral compartments and their relationships to provide a platform for further experimental investigations.

31P-NMR DETERMINATION OF PHOSPHOMONOESTERS IN RELATION TO PHOSPHOLIPID BIOSYNTHESIS IN TESTIS OF THE RAT AT DIFFERENT AGES

Changes in the phosphomonoester (PM) peak, as observed in in vivo 31P-NMR spectra, are often attributed to changes in phospholipid synthesis and therefore to changes in cell proliferation. However, this technique provides information about the absolute size of the phosphomonoester pool rather than its turnover rate. To investigate whether there is a good correlation between changes in PM concentration and its turnover rate, we studied the turnover rate of the two major PM compounds, phosphocholine and phosphoethanolamine, in rat testes at different stages of testis development. [3H]Choline and [3H]ethanolamine were injected intraperitoneally into rats at the age of 3, 6 and 13 weeks, respectively. Phosphorylation of these compounds and their incorporation into phospholipids, were followed up to 6 h after injection of the phospholipid precursors. When these data were compared with the changes observed in the in vivo 31P-NMR PM peak, the concentration of the PM compounds appeared to correlate linearly, both with the conversion of choline into phosphocholine, as well with the rate of phospholipid synthesis, and therefore with the rate of cell proliferation. Hence, it is suggested that cell proliferation can be monitored by determining the changes in the PM peak that is observed in in vivo 31P-NMR spectra.

An immunohistochemical study of bovine antral follicles, with special attention to non‐atretic follicles with and without atypical granulosa cells

Inhibin and oxytocin were immunohistochemically demonstrated in all non-atretic and light-atretic follicles > 2 mm from untreated and pregnant mares serum gonadotrophin (PMSG)-treated heifers and cows. Immunostaining for luteinizing hormone (LH) and oestradiol was observed in all non-atretic follicles > 4 mm, but only in follicles from PMSG-treated cows. Inhibin and oestradiol immunoreactivity was restricted to the granulosa. Oxytocin and LH immunoreactivity was visualized in both the theca interna and the granulosa. Within the granulosa, LH immunoreactivity was mainly present in cells that were located near the basement membrane. Normal granulosa cells differed from atypical granulosa cells (AGCs) with respect to their ability to bind LH and oestradiol. It is concluded that immunostaining for alpha-inhibin, oxytocin, oestradiol and LH cannot be used as a marker of follicle quality to discriminate between non-atretic follicles with AGCs and non-atretic follicles without AGCs in mid-luteal bovine ovaries.