EFFECT OF CHRONIC STRESS ON REPRODUCTIVE CYCLE AND EXPRESSION PATTERN OF RFRP-3 GENE IN ADULT FEMALE RATS
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Date
2023
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West Bengal University of Animal & Fishery Sciences, Kolkata
Abstract
The present study was conducted to analyze the effect of chronic stress on the reproductive cycle and expression pattern of the RFRP-3 gene in adult female rats. The experiments were performed on adult female Wistar rats which were divided into two groups viz. Control and the Stressed groups, and were housed individually in polypropylene cages. The rats of the Stressed group were subjected to 3 hrs. of restraint or immobilization stress daily in a modified rat restrainer for 28 days. The reproductive cycle of the female rats was monitored and the different stages of the oestrous cycle (proestrus, estrus, metestrus and diestrus) were determined through the visual assessment of the vagina of the rats and the vaginal exfoliated cell cytology. All rats from both the Control and the Stressed groups were tested for behavioural traits after the completion of the trial period. Different tests viz. open field locomotion test, Porsolt’s forced swim test, Light/Dark Transition test and the Elevated plus maze test were performed to observe the anxiety/depressive behaviour of the rats.The experimental animals of both groups were sacrificed at the end of the trial period. Plasma was separated from the collected blood samples and was estimated for haematological parameters (Hb, PCV, TEC, TLC and DLC), biochemical parameters (Blood glucose, Total antioxidant, T3, T4 and Cortisol) and reproductive hormones (LH, FSH, Progesterone and Estradiol). The tissue samples collected for histological studies were ovary and for the study of the expression of the RFRP-3 gene were brain (hypothalamus). During the visual assessment of the vagina, the vaginal opening of the rat appeared to be wide and moist and the tissues appeared pink during proestrus. During estrus, the vagina appeared similar to proestrus but was less pink and moist. In metestrus, a pale and dry vaginal opening was seen and during the proestrus, the vaginal opening was very moist, small and closed. The result of the vaginal cell cytology showed that a mixture of epithelial cells was seen in the proestrus. The estrus stage was characterized mainly by the presence of anucleated cornified (keratinized) cells. Metestrus shows the presence of leucocytes and some keratinized cells. Polymorphonuclear leucocytes were seen in diestrus. The behavioural analysis of the rats showed that the rats of the stressed group showed more depressive/anxiety-like behaviour when compared to the control group. The PCV and Hb of the stressed rats were highly significantly(P<0.01)lower than the control rats. The TLC and the TEC between the two groups were found to be not statistically significant (P>0.05). A significant (P<0.05) variation was observed in the blood glucose, T3, T4 and Cortisol between the two groups. The variation in the total antioxidant was found to be non-significant (P>0.05). A significant (P<0.05) variation was observed in the LH of the two groups. The variation in the mean with standard error of FSH, Estradiol (E2) and Progesterone of the two groups viz. the Control and the stressed group was found to be not statistically significant (p>0.05). A significant (P<0.05) increase in the expression of the RFRP-3 gene was seen in the hypothalamus of the stressed rat. The results lead us to conclude that chronic stress causes significant changes in the normal blood physiology of the rat. Chronic stress also alters the concentration of the reproductive hormones which confirms the relationship between stress-induced female infertility and the expression of RFRP-3.