The Study On Regulation Of L-carnitine Supplementation On Fatty Acid Metabolism By AMPKα-ACC-CPT Signaling Pathways In Fish

L-carnitine is an important factor in the regulation of fatty acid metabolism,and has been widely studied in the field of aquaculture.However,the results of studies on L-carnitine supplementation on fatty acid metabolism of fish were not consistent,and even contradictory conclusions appear in the same studies.Many functions of L-carnitine are based on its regulation of fatty acid metabolism.Therefore,it has become the key to solve this problem which in-depth exploration of the mechanism of L-carnitine supplementation in regulating fatty acid metabolism of fish.ATP and acetyl COA are important products of L-carnitine to promote theβ-oxidation of fatty acids,which not only provide sufficient raw materials for many metabolic activities,but also change the energy metabolism of the body.However,changes in the energy state will affect the metabolism of fatty acids through AMPKα-ACC-CPT signaling pathway in body.In this process,the energy state and CPT-I activity of the body are regulated by L-carnitine and AMPKαsignaling system.Therefore,the study of the regulation of L-carnitine supplementation on AMPKα-ACC-CPT signaling pathway is conducive to revealing the mechanism on L-carnitine regulates the fatty acid metabolism.This study was based on the studies on L-carnitine supplementation regulating the growth performance and fatty acid composition of fish with different habits（Cyprinus carpio var.Jian and Phoxinus lagowskii Dybowski）.Firstly,we detected energy state and AMPKαmRNA expression of carp under the condition of intraperitoneal injection carnitine,and discussed whether L-carnitine supplementation can activate AMPKαactivity.Secondly,we studied the regulating effect of L-carnitine supplementation on energy state and AMPKα-ACC-CPT signaling pathway in common carp,and discussed functional mechanism of L-carnitine supplementation regulating fatty acid metabolism.Finally,we compared the regulation differences of L-carnitine supplementation on fatty acid metabolism of two fishes mediated AMPKα-ACC-CPT signaling pathway,and elaborated the interspecific differences of L-carnitine supplementation in regulating fatty acid metabolism of fishes.This study mainly includes four parts.The main results showed as follows:（1）Effects of L-carnitine supplementation on growth performance and fatty acids composition of juvenile carp.The results showed that:At the 4^th week:the growth performance（WGR,SGR,FCR,CF,VI and RIW）of the 200 mg/kg carnitine group were significantly better than those of the control group.The activity of digestive enzymes（trypase,lipase,amylase,Na⁺,k⁺-ATPase,γ-GT and AKP）of carnitine treatment groups was significantly（P<0.05）higher than those of control group.The FH and MLT in the 200 and 800 mg/kg carnitine group were significantly（P<0.05）higher than those in the control group.At the 8^thh week,the growth performance（WGR,SGR,FC,and RIW）of the 200 and 800 mg/kg carnitine groups were significantly（P<0.05）higher than those of the control group.But L-carnitine supplementation had no significant（P<0.05）effect on the CF,HI and RIW.The activity of digestive enzymes（trypase,lipase,amylase,Na⁺,k⁺-ATPase,γ-GT and AKP）of carnitine treatment groups were significantly（P<0.05）higher than those of control group.The FH and MLT in the 200 and 800 mg/kg carnitine groups were significantly（P<0.05）higher than those in the control group.In the challenge experiment,the SR and immune indexes（C3,C4,IgM,ACP and Lys）of carnitine treatment groups were significantly（P<0.05）higher than those of control group.At 4^th and 8^th week:L-carnitine supplementation significantly（P<0.05）affects the fatty acid composition in the liver and muscles in common carp.In carnitine treatment groups,SFA and HUFA contents in the liver were significantly（P<0.05）lower than those in the control group respectively,while the contents of MUFA,PUFA,EFA and∑n3+n6 in the liver were significantly（P<0.05）higher than those in the control group respectively.The contents of SFA,PUFA,HUFA EFA and∑n3+n6 in muscle were significantly（P<0.05）higher than those in control group respectively,and the contents of MUFA in muscle were significantly（P<0.05）lower than those of control group.The above results showed that L-carnitine supplementation had significant effects on the growth and fatty acid composition of common carp.（2）Effect of intraperitoneal injection of L-carnitine on free carnitine content,energy state and AMPKαmRNA expression of juvenile carp.The results showed that:compared with the control group,the free carnitine content of blood,muscle and liver in each carnitine treatment group had a tendency to increase first and then decrease.AMP/ATP in the liver of each carnitine treatment group showed a significantly（P<0.05）decreasing trend,while AMPKα1 and AMPKα2 mRNA expressions were significantly（P<0.05）increased.AMP/ATP in the muscle of each carnitine treatment group showed a significantly（P<0.05）decreasing trend,while AMPKα1 mRNA expressions were significantly（P<0.05）increased and AMPKα2 mRNA expressions were significantly（P<0.05）reduced.The above results demonstrated that L-carnitine can activate AMPKαactivity in common carp,and the activation of AMPKαactivated by L-carnitine may be independent of AMP/ATP.（3）The effects of L-carnitine supplementation on free carnitine content、energy state and AMPK-ACC-CPT signaling pathway of juvenile carp.The results showed that:At 4^th week:The free carnitine content in the liver and muscle of the 200 and 800 mg/kg carnitine groups were significantly（P<0.05）higher than that of the control group.AMP/ATP in the liver of 50 mg/kg carnitine groups was significantly（P<0.05）lower than that of the other three treatment groups respectively.AMP/ATP in muscle of the 200and 800 mg/kg carnitine group were significantly（P<0.05）higher than those of the control group respectively.The relative expressions of AMPKα1,AMPKα2,ACC and CPT-I mRNA in the liver and muscle of carnitine treatment group showed an increased trend,and all of them were significantly（P<0.05）higher than those of the control group respectively.At 8^th week:free carnitine concentration in the liver and muscle of the 200 and 800 mg/kg carnitine groups were significantly（P<0.05）higher than those of the control group respectively.AMP/ATP of50 mg/kg carnitine group in the liver was significantly（P<0.05）higher than those of the other three groups respectively.The AMP/ATP of 50 and 200 mg/kg carnitine group in muscle was significantly（P<0.05）higher than those of the other treatment groups respectively.The mRNA relative expression of AMPKα1,AMPKα2,ACC and CPT-I mRNA of carnitine treatment groups in the liver showed a decreased trend.Their relative expression in 50 and 200mg/kg carnitine groups in the liver were significantly（P<0.05）lower than the control group respectively.The mRNA relative expressions of AMPKα1,AMPKα2,ACC and CPT-I in muscle of carnitine treatment group showed an increased trend,and their relative expression were significantly（P<0.05）higher than those of the control group respectively.The above results showed a difference in the regulatory effect of L-carnitine on AMPKα-ACC-CPT signaling pathway in muscle and liver of juvenile carp.（4）Comparative study of L-carnitine supplementation on fatty acid metabolism by AMPKα-ACC-CPT signaling pathway of common carp and Phoxinus lagowskii DybowskyThe results showed that:the changes on contents of SFA,UFA,EFA and∑n3+n6 in the liver of carnitine treatment groups showed an opposite trend.The changes of SFA and∑DHA+EPA of carnitine treatment groups showed an opposite trend.AMP/ATP in liver of two fish in carnitine treatment group showed a significantly（P<0.05）decrease trend,while AMP/ATP in muscle showed a significantly（P<0.05）increase trend,and AMP/ATP in the appropriate concentration of carnitine group was significantly（P<0.05）different from other groups.The mRNA relative expression of AMPKα1,AMPKα2,ACC and CPT-I in carnitine treatment groups in livers of common carp were decreased trend,and these values in liver of Phoxinus lagowskii were an opposite trend.The mRNA relative expression of AMPKα1,AMPKα2,ACC and CPT-I in carnitine treated groups in muscle of common carp had a significantly（P<0.05）increased trend,and these values in muscle of Phoxinus lagowskii were significantly（P<0.05）decreased.These results indicated that L-carnitine supplementation has different regulatory effects on AMPKα-ACC-CPT signaling pathways in different fish,indicating that there may be a new pathway for the activation of AMPKαsignaling system in fishes.Conclusion:L-carnitine supplementation maybe affect the energy state of fish to activate/inhibit AMPKαsignaling pathway,and then up-regulate/down-regulate the mRNA relative expressions of ACC and CPT-I to regulate fatty acid metabolism of fish.Differences in the fatty acid metabolism of fish regulated by L-carnitine supplementation may be related to the specificity of AMPKαmRNA expression in different fish and tissues.