Ultrasonic Vocalization Characteristics Of Male Shank3b Knockout Mouse Model Of Autism In Heterosexual Social Behavior

Objective Autism spectrum disorder(ASD)is a neurodevelopmental disorder.Social dysfunction is one of the key symptoms of ASD.Its pathogenesis is still unclear now.Genetic factors are important factors in the pathogenesis of ASD.For example,SHANK3 is one of the high-risk genes for ASD.Ultrasonic vocalizations(USVs)in mice are calls which frequency is higher than the maximum frequency(20 k Hz)that can be heard by human ear.It is considered to be an important indicator of communication between mice.Previously,the researchers were mainly focused on the social behavior deficit of the same sex in Shank3 KO mice but paid less attention to the change characteristics of USVs under heterosexual behavior.We established a synchronizing USVs and behavior recording system and analyzed the characteristics of USVs in normal mice.Then we did systematically explore the characteristics of USVs changes of Shank3b KO mice during its socialization with heterosexual mice.Our finding may provide an experimental basis for understanding the neural mechanism of heterosexual social interaction in ASD patients.Methods1.The USVs of infant mouse with the maternal separation was recorded by a customized USVs recording system;2.The USVs of the adult mice generated from the heterosexual social interaction test was sampled by a customized USVs and behavior synchronizing recording system,and the characteristics of USVs in young and adult mice were compared;3.The heterosexual social interaction test was done in Shank3b KO male mice and littermate control before and after them get heterosexual social experience;4.The USVs and social behaviors of mice during heterosexual social interaction were recorded by an ultrasonic recording device and infrared video recording equipment respectively;5.Deep Squeak software and Boris software were used to analyze the USVs and social behavior of mice respectively.Results1.On the postnatal day 3(P3)mice,a large number of the USVs were recorded in the infant mice with maternal separation.The distribution pattern of principal frequency of USVs of P3,P5,and P7 was different.The other parameters,including the number,duration,and call length of USVs were similar among these ages;2.No USVs were recorded when the adult female mouse was staying alone.However,large amounts of USVs were found during the adult male mouse and adult female mouse were caged together;3.The principal frequency distribution of USVs from infant and adult mice were significantly different;4.The number of USVs in the first heterosexual social experiment and the second heterosexual social experiment of Shank3b KO male mice was similar to that of the control mice,but showed obvious polarization: “Chatter” and “Silentness”;5.Male Shank3b KO mice were divided into two groups according to the change characteristics of USVs in two times of heterosexual social experiments: the increasing of calls in the “Chatter” USVs group was significantly greater than that in the control mice,while the decreasing of calls in the“Silentness” USVs group was significantly greater than that in the control mice.6.The principal frequency distribution of USVs in the heterosexual social interaction of mice showed three peaks: 50~55 k Hz,60~65 k Hz,and 70~80 k Hz.In the first heterosexual social experiment,the second peak of the principal frequency distribution of USVs in the Shank3b KO USVs increased group was higher than that in the other two groups.In the second time of heterosexual social interaction experiment,the third peak of the principal frequency distribution of USVs in the Shank3b KO USVs increased group was higher than that in the wild type(WT)group,while the first peak was significantly lower than that in the other two groups.The third peak of the principal frequency distribution of USVs in the Shank3b KO group was lower than that in the WT group.7.There was no obvious correlation between the change degree of coincidence rate of USVs and behavior in two times social experiments and the change degree of behavior in Shank3b KO USVs “Chatter” and “Silentness” group.Conclusion1.The number of heterosexual social USVs in Shank3b KO male mice is polarized after having heterosexual social experience: “Chatter” and “Silentness”.2.The principal frequency distribution of USVs in the Shank3b KO group is significantly changed.3.The control mice shows a clear correlation between USVs changing and behavior-changing before and after them getting heterosexual experiences.However,both the “Chatter” and “Silentness” sub-groups of Shank3b KO mice just do a“self-talking” base on the correlation of USVs changing and heterosexual behavior-changing before and after them getting heterosexual experiences.