IN SITU AND IN VITRO IMMUNOLOCALIZATION OF OVIDUCTIN BINDING SITES ON HAMSTER UTERINE EPITHELIAL CELLS AND DETECTION OF A HAMSTER OVIDUCTIN HOMOLOGUE IN THE FEMALE RAT REPRODUCTIVE TRACT

Oviductin is an oviduct-specific and high-molecular-weight glycoprotein that has been suggested to play important roles in the early events of reproduction. The present study was undertaken to localize the oviductin binding sites in the uterine epithelial cells of the golden hamster (Mesocricetus auratus) both in situ and in vitro, and to detect a hamster oviductin homologue in the female rat reproductive tract. Immunohistochemical localization of oviductin in the hamster uterus revealed certain uterine epithelial cells reactive to the monoclonal anti-hamster oviductin antibody. In order to study the interaction between hamster oviductin and the endometrium in vitro, a method for culturing primary hamster uterine epithelial cells has been established and optimized. Study with confocal microscopy of the cell culture system showed a labeling pattern similar to what was observed using immunohistochemistry. Pre-embedding immunolabeling of cultured uterine epithelial cells also showed gold particles associated with the plasma membrane and microvilli. These results demonstrated that hamster oviductin can bind to the plasma membrane of certain hamster uterine epithelial cells, suggesting the presence of a putative oviductin receptor on the uterine epithelial cell surface. In the second part of the present study, using the monoclonal anti-hamster oviductin antibody that cross-reacts with the rat tissue, we have been able to detect an oviduct-specific glycoprotein, with a molecular weight of 180~300kDa, in the female rat reproductive tract. Immunohistochemical labeling of the female rat reproductive tract revealed a strong immunolabeling in the non-ciliated oviductal epithelial cells and a faint immunoreaction on the cell surface of some uterine epithelial cells. Ultrastructurally, immunogold labeling was restricted to the secretory granules, Golgi apparatus, and microvilli of the non-ciliated secretory cells of the oviduct. In the uterus, immunogold labeling was observed on the cell surface of some uterine epithelial cells. Furthermore, electron micrographs of ovulated oocytes showed an intense immunolabeling for rat oviductin within the perivitelline space surrounding the ovulated oocytes. The findings of the present study demonstrated that oviductin is present in the rat oviduct and uterus, and it appears that, in the rat, oviductin is secreted by the non-ciliated secretory cells of the oviduct.

Variation in offspring size towards the high elevational range edge of a montane annual

Understanding the ecological determinants of species’ distribution is a fundamental goal of ecology, and is increasingly important with changing limits to species’ range. Species often reach distributional limits on gradients of resource availability, but the extent to which offspring provisioning varies towards range limits is poorly understood. Selection is generally expected to favour higher provisioning of individual offspring in environments with short growing seasons and limited moisture, nutrients, or hosts for parasitism. However, individual provisioning may decline if parent size is limited by resources. This thesis focuses on three major questions: 1) does seed size vary over an elevational gradient? 2) does this variation respond adaptively towards the range limit? and 3) is potential elevational variation environmentally or genetically controlled? I tested variation in seed investment towards the upper elevational limit of the hemiparasitic annual herb Rhinanthus minor, sampled across an elevational range of 1,000m in the Rocky Mountains of Alberta, Canada. I also used a reciprocal transplant experiment to address the heritability of seed mass. Seed mass increased marginally towards higher elevations, while seed number and plant size declined. There was a strong elevational increase in seed mass scaled by overall plant size. Therefore, investment in individual seeds was higher towards the upper range edge, indicating potential adaptation of the reproductive strategy to allow for establishment in marginal environments. Genetic, environmental, and genotype-by-environment interactions were observed in transplanted populations, but the relative proportions of these effects on seed size were unclear.