The Study Of Corticotropin-releasing Factor On Feeding Regulation In Acipenser Dabryanus

In order to explore the feeding regulation effect of CRF on fish,this experiment firstly cloned CRF system related factors(CRF,UI,UCN2,UCN3,CRF-R1,CRF-R2 and CRF-BP)in Dabry's sturgeon.The cDNA was tested for tissue distribution of CRF-related factors.Furthermore,the expression response of CRF system related factors was investigated by pre-and post-feeding,fasting and re-feeding tests.Finally,the effects of short-term and long-term feeding on CRF were studied by single and continuous intraperitoneal injection of CRF,and the feeding regulation mechanism of CRF system was initially explored.Results were as below.A 953 bp Dabry's sturgeon CRF cDNA fragment was obtained,which has a ORF of 557 bp and encodes 148 amino acids,comprising a 20 amino acid signal peptide and a 41 amino acid mature peptide.CRF is widely distributed in 13 tissues of Dabry's sturgeon,and the expression level in the brain is high.The relative expression of CRF in the esophagus,stomach,duodenum and pyloric cecum is higher in peripheral tissues.A 563 bp UI cDNA sequence was obtained,including a 510 bp ORF encoding 169 amino acids;the obtained Dabry's sturgeon UCN3 cDNA sequence was 575 bp,the 447 bp ORF encoding 148 amino acids;and the Dabry's sturgeon UCN2 was only obtained.The sequence is 222 bp,and part of the CDS is 180 bp,encoding 74 amino acids.The central nervous system showed the highest expression of UCN3 in the brains of 13 detected tissues,the higher UI,the moderate abundance of UCN2,and the highest expression of gastric UI,UCN2 and UCN3 in the peripheral system.UCNs also have higher abundance in the pyloric sac,the liver,the spleen and the esophagus of the Dabry's sturgeon.The cloned CRF-R1 partial CDS 1053 bp can encode 350 amino acid precursor peptides;the CRF-R2 partial CDS 906 bp can encode 301 amino acid precursor peptides.CRF-BP obtained 1072 bp of cDNA,including the complete ORF of 969 bp,encoding 322 amino acids,and the predicted signal peptide contains 22 amino acids.CRF-R1 was abundantly expressed in the brain in 13 detected tissues of Dabry's sturgeon,and the expression of gastric,pyloric,and sputum in peripheral tissues was high;CRF-R2 was expressed in the stomach,and other tissues including brain.The expression level of CRF-BP was the highest in the stomach,followed by the rectum,spleen and sputum.The expression in the eyes,esophagus,valve intestine,middle kidney,liver and pancreas was low,and other tissues including brain expression.The results of pre-and post-feeding tests showed that the expression of CRF mRNA in Dabry's sturgeon was significantly increased after feeding,suggesting that CRF may be used as a short-term satiety factor.The fasting and re-feeding test showed that the expression level of CRF mRNA in the brain was significantly decreased after fasting,suggesting CRF can be used as a long-term satiety factor.Before and after feeding,the expression of UCNs mRNA in Dabry's sturgeon chinensis was significantly increased after feeding,suggesting that UCNs may be used as a short-term satiety factor.The fasting and re-feeding tests showed that the expression of UCNs mRNA in brain was significantly decreased after fasting.After re-feeding,the expression increased,suggesting that UCNs can be used as a long-term satiety factor.Before and after feeding,the expressions of CRF-R1 and CRF-R2 mRNA in D.chinensis after feeding did not change significantly but increased,while CRF-BP did not change significantly.Fasting and re-feeding tests found that on fasting day 3,both CRF-R1 and CRF-R2 were significantly decreased.CRF-R2 was significantly decreased on the 10 th day after fasting,while CRF-BP was not significantly changed during fasting for 1-10 days,suggesting that CRF-Rs participate in Darcy Indole feeding regulation and conduction of satiety,and CRF-R2 and CRF family peptides may have positive regulation,while CRF-BP is not a major factor in feeding regulation.Single intraperitoneal injection of Dabry's sturgeon CRF(50,100 and 200 ng/g BW)significantly reduced food intake by 0-1 h,and 100 ng/g BW CRF significantly reduced D.sinensis intake,while 1-3 h and 3 There was no significant change in food intake at-6 h,and 100 ng/g BW significantly reduced the cumulative food intake for 6 h.The CRF sustained 7-day intraperitoneal injection test showed that both 50 and 100 ng/g BW doses of CRF significantly reduced the daily food intake and cumulative food intake for 1-7 days,while the 200 ng/g BW dose of CRF was injected on day 5.There was a downward trend in food intake but no significant difference.Peripheral injection of CRF can decrease or decrease the expression of UI and UCN2 mRNA in the brain,and the expression of UCN3 increases or does not change,suggesting that CRF may increase/decrease/not affect the signaling pathway mediated by central nervous system UCNs,while the expression of UCNs in peripheral system is less affected.The short-term expression of brain CRF-R1 and CRF-R2 is unchanged but has a downward trend,and the long-term expression is decreased.The decrease of CRF-Rs suggests that CRF may play an anorexia effect by binding to central nervous system CRF-Rs;CRF-BP acts as The binding protein has a significant increase in expression in the long-injected central system,while expression in the peripheral system is decreased or unchanged,suggesting that CRF continuous injection may activate the central system CRF-BP and thereby block CRF and CRF-Rs signaling.In addition to interactions between CRF systems,CRF and other feeding-related factors(NPY,CART,CCK,Ghrelin,and Apelin)also interacted.The results showed that CRF treatment increased the expression of satiety factors in the central system and reduced hunger factors.The amount of expression,further this study found that the expression pattern of feeding-related factors in the central and peripheral tissues after injection of CRF showed opposite results.The above indicates that the interaction of CRF systems and the interaction of CRF and other feeding-related factors suggest that feeding regulation is a complex and elaborate process.In conclusion,CRF,UI,UCN2 and UCN3 can all be used as short-term or long-term satiety factors to regulate the intake of Dabry's sturgeon.CRF-R1 and CRF-R2 are involved in the signaling of satiety in the regulation of food intake.CRF-BP is not the main factor of ingestion regulation.CRF can inhibit the ingestion of sturgeon dactylus sturgeon in a short term or continuously,and cause the expression changes of other relevant factors in the CRF system and other ingestion related factors,indicating that the regulation of ingestion is a complex and delicate process.