| Previous study shown that when dietary fishmeal was totally replaced with plant protein,the spotted sea bass(Lateolabrax maculatus),a typical carnivorous fish,showed feed habit shifts of anorexia followed by adaptation even compensation and the early anorexia was the critical reason for its poor growth performance.In the anorexia stage,the fish was in a negative energy balance,however,activation of mTOR in the central nervous system(CNS)decreased the phosphorylation of downstream S6K1,accompanied by further up-regulation of feeding inhibition factors POMC and down-regulation of appetitive peptide AgRP expression in anorectic stage,indicating that the CNS failed to respond effectively to peripheral starvation.However,the key reason for passivation of CNS appetite was remained to be studied.Taste receptor family 1 members(T1Rs)are a class of G protein coupled receptors,which exist of T1R1,T1R2 and T1R3 three subtypes and are directly related to the feeding preferences,swallowing even digestion in fish.In the present study,taste receptor T1R3 was cloned firstly and its protein structure prediction and homology were carried out.Tissue differential expression of three types of TIRs were analyzed.Secondly,detection of the mRNA levels of T1R1,T1R2 and T1R3 in tongues and intestines,as well as levels of small peptides and amino acid transporters in the gut at anorexia,adaptation and compensation stage after replacement of plant protein were carried out.Finally,adding umami attractant in plant protein feed to explore the effects of attractants on the feeding regulation and taste perception.The main results are as follows:1.The open reading frame of T1R3 is 2574 bp,encoding 857 amino acids and belonging to the G protein-coupled receptor,which including seven transmembrane domains;it is well conserved with other carnivorous fish,but are less conservative with herbivore,omnivorous fish,mammals;T1R1 is the highest expressed in the liver,followed by the hypothalamus.There is no significant difference of expressions in the foregut,midgut,hindgut,lip,skin and tongue tissues.T1R2 is expressed in all tissues examined,and the expression differences between the tissues are small.T1R3 is mainly expressed in the foregut,midgut,hindgut,lip,skin,and tongue tissues,while is hardly expressed in the liver,and has a small amount expression in the hypothalamus.2.The taste receptors regulated well to the appetite shift in spotted sea bass fed with plant protein diet.With the increase of adaptability to plant protein,type ? taste buds in tongue tissue were degraded,and the mRNA levels of T1R1 and T1R2 in tongues and intestines were down-regulated.In addition,TIRs were involved in the regulation of amino acid transport in the intestines,therefore,taste receptors played important roles in nutrient sensing in the intestine of spotted sea bass.3.The attractants can improve the short-term anorexia induced by replacement of fishmeal by plant protein,together with a significant reduction of the appetite-suppressing hormone ?-MSH content in plasma.However,the significant reduced expressions of T1R3 and its downstream signaling pathways important proteins IP3R and TRPM5 in the intestines of fish in plant protein group and attractant group indicating that the addition of attractants could not change the nutrient perception,transport and absorption of intestinal taste receptors to plant source contents in a short period of time.This study showed that taste receptor family 1 members can involve in the regulation of feed intake in spotted sea bass,and the tongue,intestine and central nervous system together constitute a powerful taste perception system of fish,and T1Rs play important roles in the nutrients perception and information transfer.This project is helpful to clarify the molecular basis of fish taste perception and feeding regulation mechanism and can serve as the basis for efficient utilization of plant protein feed in the aquatic product. |
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