Effects Of Music Stimulation On Behavioral Responses Of Positive Emotions And Intestinal Health In Weaned Piglets

In recent years,music has been widely used as an environmental enrichment condition to improve the survival environment of captive animals.Pigs have complex cognition,whether music can bring positive emotional experiences remains to be explored.In addition,the gut is an important digestive and immune organ of the body,and how music can affect the health of the gut needs to be further explored.This study aimed to investigate: 1)the effects of short-term musical stimulation on positive emotional and behavioral responses of weaned piglets;2)effects of music on intestinal barrier function and intestinal health status of piglets.In this study,72 weaned piglets were randomly divided into three groups,which were exposed to 60-70 d B music environment(Mozart sonata K.448),80-85 d B noise environment(mechanical noise)and control group(normal quiet environment),respectively.The piglets were given sound stimulation for 6h every day(10:00-16:00)until the end of the 56-day experiment.The playing behavior,tail wagging behavior,explorative behavior,and aggressive behavior of piglets during the first 3 days of sound stimulation were observed and recorded.On days 3,28,and 56,the jejunal tissue,jejunal mucosa,and cecum contents were collected.HE and transmission electron microscopy were used to observe the microstructure of the intestinal tract.The oxidative stress response of jejunal tissue and the level of brush border enzymes in jejunal mucosa were detected by kits.The relative m RNA expression levels of intestinal permeability related proteins,key node molecules of NF-k B inflammatory signaling pathway,Toll-like receptors,interleukin family,interferon,tumor necrosis factor,colony-stimulating factor,transforming growth factor,chemokines and chemokine receptors were detected by Quantitative Real-time PCR.Targeted metabolomics was used to quantitatively analyze the level of short-chain fatty acids in cecal contents.Finally,the structure of intestinal microflora was analyzed by 16 Sr DNA sequencing.The results were as follows: 1)The playing behaviors of piglets in the music group was significantly higher than that in the control group(P < 0.05),and extremely significantly higher than that of the noise group(P < 0.01).The playing behaviors including flopping,pivoting,pawing,head toss,scampering,social play and climbing,were significantly higher than those of control group(P < 0.05).The playing behaviors in noise group was significantly higher than that in the control group(P < 0.01).The tail wagging behavior and explorative behavior of weaned piglets in the music group were significantly higher than those in the control group(P < 0.01),and those in the control group were significantly higher than those in the noise group(P < 0.01).The aggressive behavior of weaned piglets in noise environment was significantly higher than that in the music group and the control group(P < 0.01),however,there was no significant difference between the music group and the control group(P > 0.05).Therefore,short-term music stimulation induces rich activity behavior of weaned piglets and stimulates the expression of playing behavior,explorative behavior,and tail wagging behavior,while noise stimulates greatly increases aggressive behavior of piglets.2)Pathological sections of the jejunum tissue of weaned piglets showed that the structure of jejunal villi was intact and the epithelial cells were arranged neatly.The ultrastructural observation of jejunum showed that the nucleus in the music group was intact,the chromatin was evenly distributed,and the mitochondrial cristae was intact.In the noise group,more ultrastructural changes occurred,such as nuclear shrinkage,nuclear chromatin condensation,and some mitochondrial cristae disappeared or broken.The results of tight junction protein gene expression showed that on the 3rd day of sound stimulation,music stimulation significantly enhanced the expression of Occludin and Claudin-1(P < 0.05),and noise significantly enhanced the expression of Claudin-1(P < 0.05).On the 28 th day of sound stimulation,music significantly increased the expression of Claudin-4 and E-Cadherin(P < 0.05),while noise significantly decreased the expression of ZO-1,Occludin,and Claudin-1(P < 0.05).On day 56 of sound stimulation,music significantly increased the expression of ZO-1(P < 0.05).In conclusion,music stimulation always increased the relative expression of some tight junction proteins,reduced the intestinal permeability of weaned piglets,and enhanced the structural basis of intestinal health.Noise stimulation enhances intestinal structure in the early stage and improves intestinal permeability in the late stage,indicating that long-term noise environment is a chronic stress stimulus.3)Tests of intestinal immunity in weaned piglets showed: On day 3 of sound stimulation,the expression of IL-1RN,IL-4,IL-17,IL-18,TNF-α,TGF-βR1,CCL-2,CXCL-9,CXCL-10,CCR-5,CXCR-2 and CXCR-3 in the music group was significantly increased(P < 0.05),IL-5 expression was significantly decreased(P < 0.05).The expressions of CSF-1 and CCR-4 in the noise group were significantly increased(P < 0.05),IL-1β,IL-2,IL-6 and CCL-28 were significantly decreased(P < 0.05).On the 28 th day of sound stimulation,music significantly increased the expression levels of Myd88,Ik Bα,NF-k B,TLR-4,TLR-7,IL-1β,IL-2,IL-4,TNFRSF1 B and CSF-2(P <0.05),decreased the expression of TLR-6,TLR-9,IL-1RN,IL-5,IL-6,IL-8,IL-10,IL-12RB1,IL-13,IL-17,IL-18,IL-21,IL-33,IFN-δ1,TGF-βR1,CXCL-9,CXCL-10,CCRL-2,CCR-5,CXCR-3,CXCR-4,CCR-4(P < 0.05).Noise significantly enhanced the expression of CSF-2 and CCL-25(P < 0.05),significantly decreased the expression of Ik Bα,NF-k B,TLR-6,TLR-9,TLR-10,IFN-α1,IFN-β1,IFN-γ,TNF-α,TGF-β1,CXCL-9,CXCL-10,CCR-2,CCR-5,CXCR-3,CXCR-4,CCl-2,CCL-28(P < 0.05).On day 56 of sound processing,the expressions of Ik Bα,TLR-2,TLR-4,TLR-7,TLR-9,IL-1β,IL-1RN,IL-4,IL-5,IL-6,IL-10,IL-12RB1,IL-21,IL-33,TNF-α,TNFRSF1 B,CSF-1,TGF-β1,CCL-2,CXCL-9 and CXCR-3on jejunum of piglets in music group were significantly increased(P < 0.05),the expressions of TLR-3,IL-2,IL-8,IL-13,IFN-α1,IFN-β1,TGF-βR1,CXCL-16,CCR-5 and CXCR-4 were significantly decreased(P < 0.05).Noise significantly enhanced m RNA expression of TLR-10,IL-4,IL-6,IL-10,IL-13,IL-33,TNF-α,CSF-2,CCL-2,CXCL-9,CXCR-3,CCL-25,CXCL-9,CXCL-16(P < 0.05),significantly decreased TLR-2,TLR-4,TLR-6,IL-2,IL-5,IL-8,IFN-α1,IFN-β1,IFN-δ1,TNFRSF1 B,CSF-1,TGF-β1,CXCL-16,CCR-5,CXCR-4,CCL-28,CXCL-10,CCR Expressions of-4,CXCR-2,CXCR-4(P < 0.05).In summary,music can stimulate mucosal immunity,enhance the expression of anti-inflammatory factors,and improve the intestinal immune barrier function.4)The results of oxidative and antioxidant reaction in jejunum tissue showed: Compared with the control group,music stimulation first decreased and then increased the level of catalase in the jejunum of weaned piglets(P < 0.05),while noise stimulation decreased the level of catalase(P <0.01).With the increase of sound stimulation duration,the content of glutathione peroxidase in the music group showed an increasing trend(P < 0.01).However,compared with the control group,the content of glutathione peroxidase in the music and noise groups was lower(P < 0.01).The concentration of superoxide dismutase and reactive oxygen species in the music group were significantly lower than those in the noise group,and the noise group were significantly lower than those in the control group(P < 0.01).In conclusion,sound stimulation reduced oxidative stress in the gut,and music was more effective.5)The results of brush border enzymes in the jejunal mucosa showed: The content of invertase in the music group increased significantly from day 3 to day 28(P < 0.01),but it was significantly lower than that in the control group(P < 0.01).Compared with the control,the invertase content was significantly decreased at 3 and 28 days after noise exposure(P < 0.01).The content of maltase in the music group was always increased(P < 0.01).The levels of lactase and maltase were significantly decreased after exposure to music and noise for 28 and 56days(P <0.01).The results of quantitative detection of short-chain fatty acids in cecal contents showed that:the content of butyric acid in the music group was significantly higher than that in the control group after 28 days of sound stimulation(P < 0.05).On day 56,propionic acid in the music group was significantly higher than that in the control group(P < 0.05).The results of differential expression analysis of metabolites showed that compared with the control group,the relative expression of isovaleric acid in the music group was significantly decreased on the 3rd day,the relative expression of propionic acid in the music group was significantly increased on the 28 th day,and the relative expression of caproic acid,isobutyric acid and isovaleric acid in the noise group was significantly decreased on the 56 th day.Therefore,music can increase the production of short-chain fatty acids in the intestine of piglets and enhance the intestinal chemical barrier.Noise reduces the expression of short-chain fatty acids,which is detrimental to the intestinal chemical barrier of piglets.6)The results of the 16 Sr DNA sequencing showed that the effect of the sound stimulation on the Alpha and Beta diversity of the intestinal microflora was not significant.The results of PCo A analysis showed that there was a certain degree of separation between the microflora between the music and the control group.Pairwise comparison of the sample groups using Metagenome Seq analysis revealed that on day 3 of sound stimulation,music increased the abundance of Prevotella,Lactobacillus,and Faecalibacterium bacteria compared with the control group;noise for 3days increased the abundance of Prevotella,Lactobacillus,Streptococcus,and Roseburia bacteria.Music stimulation for 28 days increased the abundance of Prevotella,Lactobacillus,Fructobacillus,Coprococcus,Roseburia,Butryricicoccus,Ruminococcus and Phascolarctobacterium bacteria.After 56 days of stimulation,increased abundance of Prevotella and Lactobacillus was observed in the music group,and increased abundance of Gemmiger was observed in the noise group.In sum,both music and noise can increase the number of beneficial intestinal flora,and music is more effective in enhancing intestinal microbial barrier.All in all,short-term musical stimulation promotes active behavioral expression and stimulates positive emotions in weaned piglets,whereas short-term noise stimulation leads to increased aggressive behavior.Music can enhance the intestinal mechanical barrier,immune barrier,and chemical barrier of piglets,reduce the level of intestinal oxidative stress,increase the number of beneficial bacteria,and promote intestinal health.Noise damage the intestinal mechanical barrier,immune barrier and chemical barrier to a certain extent,and the effect on microbial barrier and antioxidant capacity is weaker than that of music stimulation.