Development of in-vitro maturation protocol for rat oocytes; under simple culture vs co-culture with cumulus cell monolayer and its developmental potential via Parthenogenetic/artificial activation.

BACKGROUND: Murine is the most abundantly used as laboratory animal models. There has been a tremendous amount of research including; their evolution, growth, physiology, disease modeling as well as genomic mapping. Rats and mice are the most widely used among them. Although both rats and mice fall under the same category still both are different a lot too. As regarding in vitro maturation and development mouse studies are well established as compared to rats which still lies in the early phase of development. So, we tried to figure out rat oocytes in vitro maturation and their developmental potential by performing 3 experiments i.e. superovulation, in vitro Maturation as simple culture (COC's only), and COC's & cumulus cells co-culture, which later further developed using parthenogenetic activation after IVM. Female Sprague Dawley rat 3-4 week used for these studies, we hyper-stimulated their ovaries using PMSG and hCG 150 IU/kg each. After that, we collected ovaries via dissection and retrieved oocytes. We matured them in TCM 199 supplemented with FSH, Estrogen, EGF, and Pyruvate. After maturation, we activated them using two types of activators i.e. Ethanol 7%, Ionomycin. After that, we saw and compared their developmental potential in vitro. RESULTS: Oocytes matured in COC's and Cumulus cell monolayer co-culture (59% ± 4*) showed significantly more even growth and extrusion of the first polar body as compared to the COC's only culture (53.8 ± 7%*). While oocytes activated using Ionomycin showed more promising development until 8 cells/blastocyst level compared to ethanol 7%. CONCLUSION: we concluded that COC's and cumulus monolayer co-culture is better than COC's only culture. Cumulus monolayer provides extra aid in the absorption of nutrients and supplements thus providing a better environment for oocytes growth. Also, we concluded that matured oocytes showed more developmental capacity after activation via ionomycin compared to ethanol.

Timing of fertile period for successful pregnancy in American Bully dogs.

Determination of the timing of the estrus cycle is essential for fertile mating. There are physiological variations among breeds, between bitches, and between cycles of the same bitch. If serial monitoring and many tools are applied, the exact moment of ovulation could be pin-pointed. However, it leads to time and costly difficulties. Progesterone concentrations during estrus cycles follow a specific pattern, and it is largely used in timing of fertile period. Although it has similar pattern in general, it is likely that breed-specific differences exist. The aim of this study was to investigate the way of timing the fertile period for successful pregnancy in American Bully dogs based on vaginal cytology and progesterone assay with minimized cost. To identify the empirical relations among reproductive characteristics, we performed statistical analyses on data from proestrus-to-estrus 27 American Bully dogs referred for 7 months. We found the significant correlations between the cyclic changes of vaginal cytology and progesterone assay. The relationship of serum progesterone concentrations with the days from vaginal discharge onset was analyzed through linear regression assay. In conclusion, we addressed two standards in the timing of fertile period with a minimal number of progesterone assays in the breeding management of American Bully dogs.