| BackgroundPerinatal and early childhood are critical periods for the development of the brain.During this critical period,brain neurons quickly proliferate and differentiate,forming many synaptic connections with each other,and eventually develop into a complex central nervous system.However,environmental stress can greatly affect brain development.For example,children who need surgical interventions come in contact with many stressors: mental,pain,inflammation and anesthesia,which may affect their brain and behavioral development.In 2016,the US Food and Drug Administration issued a warning that repeated or prolonged use of general anesthetics and sedatives at the end of three months of pregnancy or under three years of age(>3 hours)may affect the brain development of the fetus and children.Perioperative anesthesia in infants and young children causes neurological damage and dysfunction that has become a clinical and social concern worldwide.Sevoflurane is the most commonly used inhalation anesthetic in infants and young children.In recent years,more and more studies have reported that sevoflurane can induce cognitive disorders in the developing brain,including social interaction disorders,and surgical trauma can also cause cognitive disorders.However,the specific mechanism is not yet clear,and urgently needed to be solved in order to guide the rational use of drugs in the clinic.Part one1.Specific Aims:To evaluate the effect of sevoflurane anesthesia and the sevoflurane anesthesia combined with laparotomy on C57/BL6 mice in neonatal period about their social interaction.2.Methods:(1)Selection of animal models:The study found that mice(C57BL/6J)had better social communication tendencies and social novelty preferences.Therefore,C57BL/6J mice born on the 7th day were selected for this experiment.(2)Establishment of animal model:(1)Anesthesia group: neonatal(7 days)C57BL/6J mice "inhaled 3 % sevoflurane + 60 % oxygen" for 2 hours(22-25 °C at room temperature);(2)Surgical group: neonatal(7 days)C57BL/6J mice "inhaled 3 % sevoflurane + 60 % oxygen" 2 hours + exploratory laparotomy(room temperature maintenance 22-25 °C);(3)Control group: neonatal(7 days)C57BL/6J mice "inhaled 60 % oxygen" for 2 hours(22-25 °C at room temperature).After completion of the model,the neonatal C57BL/6J mice were fed by the dam.On day 30,the social ability of each group of C57BL/6J mice was tested.(3)Sociability test(three-room social communication model):The first stage is the habituation: firstly,the mice to be tested are placed in the central room,and the two doors are open.There is an identical cage on each side of the room,and mice can freely adapt to the environment in each room.The anymaze software automatically records the time and frequency of its entering and exitsing each room,as well as the time and frequency of the active sniffing of the iron cage,which is 5 minutes.The second phase tests is social interaction(Sociability): The first stranger is placed in a cage and observes the mice being tested for free activities and active contact with strangers.The anymaze software automatically records the time and frequency of its entering and exitsing each room,as well as the time and frequency of the active sniffing of strange similar or the empty cage,which is 10 minutes.The third phase tests is social novelty preferences(Social Novelty): The second newcomer is unfamiliarly placed in a cage in the opposite room,and then the behavior of the tested mouse were observed for its free movement and active exposure to a stranger of the same kind.The anymaze software automatically records the time and frequency of its entering and exitsing each room,and as well as t The time and frequency of free activity and active contact with new or original strangers,which is 10 minutes.After obtaining the experimental data,the significance and correlation of each behavior parameter were analyzed.3.ResultsSurgical group was partially inhibited during the infancy period(day 30),but their social behavior tendencies were normal.The time of offspring sniffing strange 1 is much longer than when they sniff the empty cages(Control: P <0.0001;Anesthesia: P<0.05;Surgery: P<0.001);the social novelty were significantly decreased(Control: P=0.01;Anesthesia: P=0.001;Surgery: P=0.17).4.ConclusionNeonatal C57BL/6J mice were inhaled with 3 % sevoflurane 2-hour combined laparotomy,which inhibited their social interaction and induced cognitive impairment during the early childhood(day 30).Part two1.Specific Aims:To evaluate the effects of sevoflurane anesthesia and the sevoflurane anesthesia combined with laparotomy on the social communication behaviors of early adult C57/BL6 mice.2.Methods(1)Selection of animal models:The study found that mice(C57BL/6J)had better social communication tendencies and social novelty preferences.Therefore,this experiment selected C57BL/6J mice as early as the 60 th day of the year.(2)Establishment of animal model:(1)Anesthesia group: During the early adulthood(60 days),C57BL/6J mice were "inhaled 3 % sevoflurane + 60 % oxygen" for 2 hours(22-25 °C at room temperature);(2)Surgical group: During the early adulthood(60 days),C57BL/6J mice were inhaled with 3 % sevoflurane + 60 % oxygen for 2 hours + intraperitoneal exploration(room temperature maintained at 22-25 °C);(3)Control group: During the early adulthood(60 days),C57BL/6J mice were inhaled with 60 % oxygen for 2 hours(22-25 °C at room temperature).On the 1st,3rd,and 7th days after completion of the model,the social ability of each group of C57BL/6J mice was tested.(3)Sociability test(three-room social communication model):The first stage is the habituation: firstly,the mice to be tested are placed in the central room,and the two doors are open.There is an identical cage on each side of the room,and mice can freely adapt to the environment in each room.The anymaze software automatically records the time and frequency of its entering and exitsing each room,as well as the time and frequency of the active sniffing of the iron cage,which is 5 minutes.The second phase tests is social interaction(Sociability): The first stranger is placed in a cage and observes the mice being tested for free activities and active contact with strangers.The anymaze software automatically records the time and frequency of its entering and exitsing each room,as well as the time and frequency of the active sniffing of strange similar or the empty cage,which is 10 minutes.The third phase tests is social novelty preferences(Social Novelty): The second newcomer is unfamiliarly placed in a cage in the opposite room,and then the behavior of the tested mouse were observed for its free movement and active exposure to a stranger of the same kind.The anymaze software automatically records the time and frequency of its entering and exitsing each room,and as well as t The time and frequency of free activity and active contact with new or original strangers,which is 10 minutes.After obtaining the experimental data,the significance and correlation of each behavior parameter were analyzed.(4)Western blotting was used to measure the expression of PSD95 in hippocampus of C57BL/6J mice.3.ResultsIn the anesthesia group,the social communication behavior was inhibited on the first day after exposure to sevoflurane.The social interaction was normal,and the time of offspring sniffing strange 1 is much longer than when they sniff the empty cage(Baseline: P<0.0001;Day1: P<0.0001;Day3: P<0.05;Day7: P<0.001);The social novelty was significantly decreased(Baseline: P<0.01;Day1: P=0.4212;Day3: P<0.01;Day7: P<0.01).In the surgery group,social communication behavior was suppressed in the week after the operation.The social interaction was normal,and and the time of offspring sniffing strange 1 is much longer than when they sniff the empty cage(Baseline: P<0.0001;Day1: P<0.05;Day3: P<0.05;Day7: P<0.001);The social novelty was significantly decreased(Baseline: P<0.01;Day1: P=0.4403;Day3: P<0.01;Day7: P=0.068).4.ConclusionInhalation of 3 % sevoflurane in C57BL/6J mice in early adulthood may inhibit their social interactions,but the phenomenon is short-lived.C57BL/6J mice inhaled 3 % sevoflurane combined with laparotomy will inhibit their social interactions and the phenomenon is sustainable. |
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