| Feeding a high concentrate diet to lactating ruminant easily causes a range of nutritional metabolic diseases,and has a potential risk to threat body health and eventually leading to a decline of milk performance.Feeding high concentrate diet can induce a chronic stress to lactating ruminants.However,the reports on the change of the stress axis hypothalamus-pituitary-adrenal gland(HPA)is really scarce when ruminants were fed with high concentrate diets.Cortisol,the terminal hormone of HPA axis as well as an indicator of chronic stress,can transmit external stimuli to internal tissues and organs,and then to regulate clock genes expressions in different tissues accordingly and ultimately adapting their function to changes in environment.Cortisol can also regulate glucose and lipid metabolism via binding with its specific receptor-glucocorticoid receptor(GR).Therefore,the aim of this study was to explore HPA axis activity when lactating dairy goats were fed with diets enriched with a high proportion of concentrate(65%)for long(19 wks)or short-term(4 wks).Moreover,the alternations of clock genes expression in various tissues,and the expression of genes related to hepatic glucose and lipid metabolism were also detected in this study.1 Effects of long-or short-term feeding a high concentrate diet to lactating dairy goats on HPA axisSixteen lactating dairy goats were randomly divided into three groups,goats in control group were fed with a low concentrate diet(35%concentrate diet,CON,n=5),goats in short-term group were fed with a high concentrate diet for 4 wks(65%concentrate diet,HS,n=4),and goats in long-term group were fed a high concentrate diet for 19 wks(65%concentrate diet,HL,n=7).During the experiment or at the end of the experiment,samples including serum,urine,rumen fluid,and tissures including the hypothalamus,pituitary,adrenal gland,hippocampus,liver,rumen and mammary glands were collected for analysis,respectively.The results showed that the short-term feeding high concentrate had no effect on acute phase proteins(APPs)gene expressions(P>0.05),but Serum amyloid A(SAA)and C-reactive protein(CRP)mRNA expression levels in HL group were significantly higher than that in CON(P<0.05).The level of malondiadehyde(MDA)was significantly elevated,but superoxide dismutase(SOD)enzyme activity was markedly decreased in the liver of HS goats(P<0.05),whereas the antioxidant index was not altered in HL group(P>0.05).Compared to CON group,HS goats showed a significant increase of blood cortisol concentration during lto 3h after morning feeding(P<0.05).In HL goats,plasma cortisol concentration was significantly lower than that in control goats(P<0.05),and the concentration of cortisol in urine had a tendency to decrease compared to CON(P = 0.084).However,the level of cortisol in ruminal fluid and plasma adrenocorticotropic hormone(ATCH)was not altered by long or short-term feeding a high concentrate diet(P>0.05).Real-time PCR results showed that the hypothalamic corticotropin releasing hormone(CRH)mRNA expression was not affected by feeding a high concentrate diet in HS or HL group(p>0.05).Compared with CON,CRHR and POMC mRNA expression in the pituitary was significantly down-regulated in HL but not HS goats(P<0.05).In adrenal gland,both HS and HL goats showed a significant increase of 3β-hydroxysteroid dehydrogenase(3β-HSD)mRNA expression compared to CON(P<0.05),and P450c21 mRNA expression in HS goats was decreased obviously(P<0.05),while P450c21 gene expression was not changed in HL(P>0.05).Glucocorticoid receptor(GR),the specific receptor for cortisol,mRNA expression was significantly up-regulated in the adrenal gland,pituitary,hippocampus,rumen and liver of HL goats compared to CON control goats(P<0.05),and HS group showed a marked increase of GR mRNA expression in adrenal gland as well as in rumeinal epithelial tissues(P<0.05),and a significant decrease of GR mRNA expression in the hypothalamus(P<0.05).Western blot results showed that HL group showed a significant increase of CRHR1 protein expression and a decrease of GR protein expression in the pituitary compared to CON(P<0.05),and the level of cholesterol side-chain cleavage(P450scc)protein had a tendency to decreased(P 0.086),however,the expression of GR proteins in the pituitary and adrenal gland was not changed in HS group compared to CON(P>0.05).These results indicate that short-term feeding a high concentrate diet activated the stress axis activity,however,long-term feeding induced an obvious decrease of HPA axis demonstrating by lower level of cortisol secretion and markedly up-regulation of GR gene expression as a feedback regulation mechanism in the lactating dairy goats.2 Effects of long-or short-term feeding a high concentrate diet to lactating dairy goats on clock genes expression different tissuesCompared with CON,in pituitary of HS goats,the clock genes of per 1 and clock mRNA expression were significant down-regulated,and Bmall mRNA expression had tendency to derease.There was no significant difference in clock genes expression in the pituitary between HL and CON.In hippocampus,perl gene expression in HS and tim gene expression in HL were markedly decreased(P<0.05).In ruminal epithelium,the expression of Bmall,perl,clock and tim mRNA in HL were significantly higher than that in CON,whereas no change were observed between CON and HS.In liver,clock mRNA expression in HL and tim mRNA expression in HS were markedly increased compared to CON,and cryl gene expression in HS had tendency to increased(P = 0.053).In mammary gland,clock mRNA expression in HL and tim mRNA expression in HS were decreased compared to CON.The expression of clock genes in the hypothalamus and adrenal gland did not show a significant difference between HS and HL.The correlation analysis showed that plasma cortisol concentration had a significant positive correlation with tim mRNA expression(R = 0.572,P = 0.021)in the adrenal gland,and perl(R=0.505,P =0.046)mRNA expression abundance in the mammary gland,and showed a tendency of positive correlation with the gene expression of clock(R =0.452,P = 0.079)in mammary gland.Moreover,the level of plasma cortisol showed a negative correlation with Bmal1(R=-0.699,P = 0.003),clock(R =-0.506,P = 0.045)and tim(R =-0.551,P = 0.027)mRNA expression in the ruminal epithelium,and showed a tendency of negative correlation with per1(R =-0.436,P = 0.091)mRNA expression.Moreover,ruminal cortisol concentration showed a strong negative correlation with Bmal1(R =-0.589,P = 0.026)and tim(R =-0.543,P = 0.044)mRNA expression in ruminal epithelium.The results suggest that a general alteration of clock genes transcription was found in goats fed with a high concentrate diet,and there was a significant correlation between the level of cortisol and clock genes expression in the rumen epithelium and mammary gland.3 Effects of long-or short-term feeding a high concentrate diet to lactating dairy goats on glucose and lipid metabolism in the liverThe biochemical analysis demonstrated that HS and HL goats showed significantly increased levels of plasma glucose and hepatic glycogen content compared to CON,yet a significant decrease in plasma triglyceride(TG)level.The concentration of total cholesterol(TCH)in plasma did not show a significant difference between control and treated groups(P>0.05).The concentrations of hepatic TG and TCH were significantly elevated in HS(P<0.05)but not in HL group(P>0.05).In HL group,mRNA expression of Pyruvic carboxylase(PC)and Glucose transporter 2(GLUT2)in liver were significantly up-regulated compared to control(P<0.05).In HS group,mRNA expression of PC,sterol regulation element-binding protein 1(SREBP1),stearoyl-CoA desaturase(SCD),Diacylgycerol acyltransferase(DGAT)and 3-hydroxy-3-methyl glutaryl coenzyme A reductase(HMGCR)in the liver as well as phosphoenol pyruvate carboxykinsae 2(PCK2)protein levels were markedly increased,and SREBP-2 expression had a tendency to increase(P = 0.76),while carnitine palmitoyltransferase-1(CPT-1)and cholesterol 7a-hydroxylase(CYP7α1)mRNA gene expression were markedly decreased compared to control group(P<0.05).These results suggest that feeding a high concentrate diet for a short term enhances gluconeogenesis and promotes lipogenesis in the liver;however,feeding a high concentrate diet for a long term increased gluconeogenesis,but did not affect lipid metabolism in liver. |
