We investigated the effect of heat stress (HS) on the expression of the GABA receptor in the hypothalamic-pituitary-gonadal (HPG) axis of Wenchang chickens. Real-time quantitative RT-PCR (qRT-PCR) was used to quantify the GABA receptor mRNA levels along the HPG axis of chickens under HS (40±0.5 °C) for 1-6 weeks. Our results showed that the expression of GABA A and GABA B receptor at the mRNAs levels in the tissues of HPG axis exhibited fluctuation and variability. After HS, the mRNA level of GABA A receptor was significantly reduced in the hypothalamus of 1-week-old and in the pituitary of 3-week-old chickens, but significantly increased in the pituitary of 1-, 4-, and 5-weekold chickens. The GABA B receptor mRNA level significantly declined in the hypothalamus of 1-week-old and in the pituitary of 3-week-old chickens, but was significantly upregulated in the pituitary and testis of 1-and 2-week-old chickens. At other time points, the expressions of GABA A receptor and GABA B receptor showed no significant differences compared with control group. These results indicated that the levels of GABA A receptor and GABA B receptor mRNAs varied in different tissues of the HPG axis in chickens of different ages, displaying temporal and spatial variations. GABA receptor behaved as a positively-regulated gene by HS, i.e., its mRNA was increased by HS; similarly, it was a negativelyregulated gene by HS, when its expression was reduced by HS.
ABSTRACT:This study aims to investigate the effect of dietary γ-aminobutyric acid (GABA) on the development of thymus tissue structure and function in chicks under heat stress. One-day-old male Wenchang chicks were randomly divided into control group (CK), heat stress group (HS), and GABA+HS group. The chicks from GABA+HS group were administered 0.2 ml of GABA solution daily by oral gavage (50 mg/kg of body weight). Chicks from HS and GABA+HS groups were subjected to heat stress treatment at 40 ± 0.5°C for 2 h every day. Blood and thymus tissue were collected from the chicks at the end of weeks 1-6. Results showed that the thymus weight and index, thickness of cortex, cortex/medulla ratio, number of lymphocytes, activity of superoxide dismutase, total antioxidant capacity, and glutathione peroxidase, and plasma level of tumor necrosis factor-α in HS group were significantly lower than in CK group (P < 0.05). The Toll-like receptor 2 (TLR2) expression in the late stage of heat stress, malondialdehyde (MDA) content, thymocyte apoptosis rate, number of lymphocytes in the S and G2/M phases, and plasma levels of interleukin-4 and interferon-γ in HS group were significantly higher than in CK group (P < 0.05). In contrast, the integrity of thymus tissue structure of GABA+HS group was improved compared with HS group. The TLR2 expression in the early stage of heat stress and the activity of antioxidant enzymes in GABA+HS group were significantly higher than in HS group (P < 0.05), and the MDA content, thymocyte apoptosis rate, number of lymphocytes in the S and G2/M phases, and plasma level of IL-4 and IFN-γ in GABA+HS group were significantly lower than in HS group (P < 0.05). We concluded that heat stress caused structure damage to thymus tissue of chicks, changed the plasma levels of cytokines, reduced the antioxidant activity, and increased cell apoptosis in chick thymus. GABA alleviated the negative effects on the development of chick thymus, improved the immune function of thymus, and played a protective role by regulating the plasma levels of cytokines and antioxidant activity of thymus tissue.
Under routine feeding conditions, 1-day-old male chicks were randomly divided into control group (CK), heat-stress group (HS), and GABA + heat stress group (GABA+HS). The thymus contents of IL-1, IL-2, TGF-β1, IFN-γ, GH and HSP70 were determined by enzyme-linked immunosorbent assays. The results showed that: (1) IL-1 and TGF-β1 contents of HS group were significantly lower than those of the CK group (p<0.05), and those of the GABA+HS group were significantly higher relative to the HS group (p<0.05); (2) IL-2 and IFN-γ contents of the HS group were significantly higher than those of the CK group (p<0.05), and those of the GABA+HS group were significantly lower relative to the HS group (p<0.05);(3) the thymus GH content of all three groups first increased and then decreased. The expression levels GH of the HS and GABA+HS groups were significantly lower than CK group (p<0.05); and (4) HSP70 expression levels in the thymus were significantly higher in the HS and GABA+HS groups relative to the CK group (p<0.05). These results indicate that heat stress affected thymus development, immune functions, and overall growth of chickens. Furthermore, it was shown that feeding GABA may significantly improve the immune responses of heat-stressed chickens by increasing the expression levels of IL-1 and TGF-β1, and effectively alleviate the negative effects of heat stress on thymus development by changing HSP70 expression and GH secretion.
This study conducted an in-depth investigation on the development of GABAergic neurons and their receptors in HPG axis-related target organs of Wenchang chicks under heat stress. One-day-old healthy Wenchang chicks were randomly divided into control (CK) and heat stress (HS) groups. Chicks in the HS group were placed in a 40±0.5°C climatic chamber for HS treatment from 13:00 to 15:00 daily. By immunohistochemistry and Western blotting, GABA and GABA A receptor (GABA A R) expression in the hypothalamus of the HS group was significantly higher (p<0.05), but GABA B receptor (GABA B R) expression was significantly lower than that of the CK group (p<0.05). Expression of GABA and its two receptors in the pituitary tissues of the HS group was significantly lower than in the CK group (p<0.05). Expression of GABA and GABA B R in ovaries in the HS group was significantly higher, but expression of GABA A R in the testes of the HS group was lower than that of the CK group (p<0.05). In the male chicks, expression of GABA and its two receptors in the hypothalamus, pituitary, and testicular tissues of the HS group was significantly higher than that of the CK group (p<0.05). Western blotting showed that the GABA A R and GABA B R expression of the HS group was significantly higher than that of the CK group at 3 and 5 weeks of age. Thus, HS caused GABAergic nervous system disorder in the HPG axis of Wenchang chicks and seriously hindered the normal development of GABAergic neurons in chicks, leading to the disorder of the expression of GABA and its receptors in tissues.
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