Effects Of Long-term Social Defeat On Social Behavior And Brain CRF System In Adult Male C57 Mice With Different Susceptibility

Chronic social stress in humans can increase levels of anxiety,depression,and social withdrawal and consequently increase the risk of mental illness.Chronic social defeat stress(CSDS)model is considered as a common model to induce symptoms such as happiness loss,social avoidance and social dysfunction.In addition,the different susceptibility to CSDS in rodent model has good translational value in humans,as humans also show different responses when exposed to similar social stressors.Some people are able to cope with stress,while others develop severe depression when exposed to stress for a long time.CSDS has been found to produce different impacts on anxiety-like behaviors,spatial cognitive function and memory in rodents with different susceptibilities.However,the impacts of chronic social defeat on social behaviors in adult male mice with different susceptibilities to social defeat and its underlying mechanisms in the brain remain unclear.Corticotropin-releasing factor(CRF),which mainly comes from the paraventricular nucleus of the hypothalamus(PVN),is associated with stress-related physiology and behavior and plays a major role in regulating the response of the hypothalamic-pituitary-adrenal(HPA)axis to stress.CRF is the main hormone that triggers adrenocorticotropic hormone(ACTH)release from the anterior pituitary under stress.CRF and its receptors are important mediators of coordinated neuroendocrine and behavioral responses to stress.CRF can act as neurotransmitters/neuromodulators in the brain to exert their effects on behavior.For example,CRF infusion into the nucleus accumbens contributes to partner preference in male prairie voles.CRF exerts its effects by binding with CRF-R1 and CRF-R2 in the brain.CRF-R1 is mainly expressed in the brain and pituitary,while CRF-R2 is only expressed in specific brain regions and some peripheral organs.CRF receptors not only show region-specific distribution but also have different physiological functions.For example,the basolateral amygdala(BLA)plays a key role in anxiety and emotional arousal and exhibits high levels of CRF-R1 expression.CRF-R2 in the nucleus accumbens shell is related to passive coping behavior after partner loss.The medial prefrontal cortex(mPFC),BLA and nucleus accumbens(NAcc)are closely associated with emotional and social behavior.Although many brain regions mediate anxiety symptoms,these three brain regions appear to be preferentially involved.There are direct connections among these three brain regions,which constitute the nervous circuit affected by pressure.Whether chronic social defeat stress has different effects on social behavior and CRF system of adult male mice with different susceptibility remains unclear.Furthermore,the neuronal circuit through which the CRF system regulates social behavior is unclear.We assumed that CSDS may affect emotional and social behaviors differently in animals with different susceptibilities via different alterations in CRF levels.Moreover,we hypothesized that different alterations in the levels of CRF-R1 and CRF-R2 in the mPFC,BLA and accumbens nucleus shell(ACbSh)may be associated with alterations in social behaviors induced by CSDS.Considering that the BLA is related to stress-related behaviors and has certain influence on the HPA axis.According to the results of pharmacological experiments,we speculate that chronic social defeat stress may change the neuronal activity of BLA,and the related neuronal circuit involved in the BLA play an important role in regulating social behaviors.In order to answer the above scientific questions,the chronic social defeat method was used to establish the animal model,and the social approach and avoidance test was used to distinguish susceptible and resilient animals in this study.At the same time,the social behaviors and anxiety-like behaviors of each group were tested.In addition,the number of CRF-,CRF-R1-and CRF-R2-positive neurons in the control,susceptible and resilient groups was determined by immunohistochemistry.Quantitative reverse transcription-polymerase chain reaction(RT-PCR)was used to measure the mRNA expression profiles of both CRF-R1 and CRF-R2 among the groups.Based on the results of the pharmacological experiments,we hypothesized that among the three brain regions concerned in this study,BLA plays an important role in regulation of social behaviors after chronic social defeat.To test our hypothesis,calcium imaging technology was used to detect the real-time activity of BLA when social behaviors occurred.Subsequently,chemical genetic techniques were used to verify the role of BLA-PVN in social behaviors.The aim is to reveal the role of the CRF system and BLA-PVN neural circuits in regulating social behaviors.The main results of the study are as follows:1.Effects of chronic social defeat on behaviorsIn the present study,we found that CSDS reduced the tendency of susceptible adult male C57 mice to approach an unfamiliar individual and increased their avoidance from an unfamiliar CD-1 mouse,but had no effects on resilient individuals.In addition,ten days of chronic social defeat enhanced anxiety-like behavior in susceptible animals,but produced no effects on the resilient group.2.Effects of chronic social defeat on levels of CRF and CRF receptor in specific brain regionsCSDS increased the number of corticotropin-releasing factor(CRF)-positive neurons in the PVN of the hypothalamus and CRF-R2-positive neurons in the ACbSh in both resilient and susceptible animals.Chronic social defeat increased the number of CRF-R1-positive neurons and CRF-R1 mRNA expression in the prelimbic cortex(PrL)and the number of CRF-R2-positive neurons in the basolateral amygdala,but reduced the number of CRF-R2-positive neurons and mRNA expression in the PrL in susceptible animals.3.Effects of glucocorticoid receptor antagonists on behaviorsAfter chronic social defeat stress,intraperitoneally injection of glucocorticoid receptor antagonists resulted in avoidance behavior of mice in the resilient mice,but had no effect on the social approach avoidance behavior of mice in the susceptible group which still showed the avoidance behavior of the unfamiliar CD-1 mice.In addition,there was no significant difference in the total distance of movement before and after drug administration in the open field test,indicating that the effect of the drug on the behavior had nothing to do with the activity of mice.4.Results of calcium imaging and chemical genetics experimentsAfter chronic social defeat stress,calcium imaging technology was used to detect the neuronal activity in BLA brain region of mice before and after drug administration when social behavior occurred.The results showed that neuronal activity in BLA brain region of mice in resilient mice group increased significantly after drug administration.Chemical genetic techniques were used to inhibit the activity of glutaminergic neurons in BLA-PVN neural circuits in mice.Inhibition of BLA-PVN neural circuits reversed the social avoidance behavior induced by social defeat.In addition,there was no significant difference in the total distance of movement before and after CNO administration in the open field test,indicating that the reversal of mouse behavior was caused by the inhibition of virus-controlled neural circuits,and CNO injection had no effect on the activity of mice.Therefore,the different effects of CSDS on sociability and anxiety-like behavior in mice with different susceptibilities may be associated with region-and type-specific alterations in CRF and CRF receptor levels.BLA-PVN neural circuits play a key role in regulating social behavioral abnormalities induced by chronic social defeat.These findings help us understand the mechanism underlying effects of social stress on emotion and social behavior and provide an important basis for the treatment of disorders of social and emotional behavior caused by social stress.