Effects Of Different Dietary Carbohydrate-to-lipid Ratios And Starvation On GLUT1 Gene Expression And Glucose Tolerance Of Grass Carp,Ctenopharyngodon Idellu And Golden Pompano,Trachinotus Ovatus

Glucose transporter 1(GLUT1),a member of the glucose transporter family,plays an important role in the transport of glucose into cells in the glucose metabolism system of animals.In the present study,we test the hypothesis that the ability of two fish species of different feeding habits: grass carp(Ctenopharyngodon idellu)and golden pompano(Trachinotus ovatus)to clear a glucose load is related to nutrient history of different dietary carbohydrate-to-lipid(CHO:L)ratios and nutritional status(in fed,unfed and refed fish)through the transport and glycolysis of glucose.Firstly,we clone and characterize the complete coding sequence from C.idellus GLUT1 and T.ovatus GLUT1.And then analyze the regulation of GLUT1 expression and glycolytic enzymes in response to a glucose load after different nutritional history.The content of studies and results are as followings:1.Sequence analysis of grass carp and golden pompano GLUT1Sequence alignment and phylogenetic analysis grass carp and golden pompano GLUT1 revealed a high degree of conservation(80–97%)among most fish and higher vertebrates.Multiple sequences alignment showed that the mutation of key function locus may affect the traffic mode and function GLUT1 transport and the glucose utilization ability of fish.The highest ciGLUT1 expression was observed in the heart followed by brain,whereas relatively low values were detected in the muscle,gill,intestine,kidney,spleen and liver.2.Effects of different nutritional status on expression of GLUT1 of grass carp and golden pompanoThe result showed that different dietary carbohydrate had no effect on GLUT1 mRNA of grass carp hepatic and muscle after 8weeks trail,but fasting induced its expression.However,in golden pompano the highest mRNA expression of GLUT1 was found in fish when fed with a low CHO:L ratio diet,and its expression was significantly higher than that of the other two groups,but the expression of GLUT1 gene in the muscle increased with the increase of CHO:L ratio diets.The starvation treatment significantly inhibited the expression of GLUT1 gene in liver and muscle tissue.The above results indicated that3.Effects of different dietary CHO:L ratio on glucose tolerance of grass carp and golden pompanoAfter 8 weeks of feeding trial with different CHO:L ratio diets,the glucose tolerance experiment was conducted.The results showed that after glucose load,the blood glucose concentration of grass carp fed high CHO:L ratio diet was the first to return to its normal level.Then we further explore the reasons why the high CHO:L ratio nutritional history is conducive to the improvement of glucose tolerance.The results showed that the expression level of liver GLUT1 was significantly up-regulated in each treatment group,In terms of dietary carbohydrate levels,the GLUT1 expressions of fish fed HC diet were significantly(P<0.05)higher than the other groups after the glucose load.In addition,the liver and muscle glycogen of grass carp with high CHO:L ratio nutrition background were significantly higher than those of the other two treatment groups at all time points.Compared with the other two treatment groups,the liver GK and PK enzyme activities of grass carps with high glycolipid ratio were significantly increased.These studies suggest that grass carps with a high CHO:L ratio nutrition background may improve their glucose tolerance by promoting the synthesis of glycogen,increasing the activity of key enzymes(GK and PK)of glycolysis and inducing the expression of GLUT1 gene.After the glucose load,the blood glucose levels of golden pompano with different CHO:L ratio nutritional backgrounds increased first and returned to normal condition at 3 h.Furthermore,the blood glucose level of fish fed high CHO:L diet was significantly higher than that of the other two groups during 1-6 h.This result indicated that the high CHO:L ratio nutritional history reduced the glucose tolerance of golden pompano.After glucose load,the activities of hepatic glycolytic enzymes(GK,HK and PK)decreased when the fish fed a long term of high CHO:L ratio diet.This result may be due to the damage of liver function and metabolic dysfunction caused by excessive carbohydrate level supplement.However,the significantly increased glycolytic enzyme activities were found in fish fed the low or moderate CHO:L ratio diet after glucose administration.These results could be an explanation for the limited ability to utilize carbohydrates in golden pompano.4.Effects of different nutritional status on glucose tolerance of grass carp and golden pompanoStarvation can improve the glucose tolerance whose blood glucose level was significantly lower than that of the other two groups at each time point,and it first returned to the normal level at 6 h.Then we further explore the reasons why the starvation could improve glucose tolerance of grass carp.It was found that the expression of GLUT1 in liver and muscle of grass carp,except the hungry refeeding group,increased first and then decreased,and the expression of GLUT1 muscle of fasting fish was higher than that of the other nutritional status at all time points(P<0.05).In addition,the glycogen content in the different treatment groups increased,and it was significantly higher in the starvation group than that in the other two groups.At the same time,the liver GK enzyme activity of grass carp was induced in all the treatments,in which the starvation group increased more significantly,while the PK enzyme activity only increased in the starvation group.The results showed that starvation could improve the glucose tolerance of grass carps,which is related to the increase of liver glycogen synthesis,the induction of glycolysis GK and PK enzyme activity,and the significant enhancement of GLUT1 expression.In conclusion,the different dietary CHO:L ratios and starvation had significant effects on the glucose tolerance of grass carps and golden pompano,in which,golden pompano fed a long-term high CHO:L ratio diet exhibited weaker glucose tolerance,opposite result was ture in grass carp.In grass carp,starvation was beneficial to improve their sugar tolerance,but in golden pompano,continuous feeding significantly improved its glucose tolerance.Under these different nutritional backgroud,the regulation of GLUT1 gene expression and key enzymes of glycolysis(GK,HK and PK)become an important factor affecting the glucose tolerance of these two different feeding habits fish species.