Establishment Of A Model For The Development Of Acute Stress Disorder To Post-traumatic Stress Disorder And Investigation Of The Mechanism

Stress that exceeds the body’s adaptive capacity can cause psychological and physiological imbalance in the body and even develop into stress disorders.Acute stress disorder(ASD)and post-traumatic stress disorder(PTSD)are the two main categories of stress disorders.The main difference between the two is the duration of symptoms,with ASD diagnosed within 1 month and PTSD diagnosed beyond 1 month.However,the mechanisms of ASD to PTSD are not well understood,and there is a lack of effective drugs for prevention and treatment in the clinic.A good animal model is important for understanding the pathogenesis of the disease and developing drugs for prevention and treatment.However,most of the existing studies for stress disorders have used a single model of conditioned fear,which is inconsistent with the clinical diagnostic criteria for stress disorders.In addition,most current animal models of stress disorders only focus on pathophysiological changes in the PTSD stage,and lack research on the relationship and mechanisms of ASD to PTSD development.A good animal model for the development of ASD to PTSD should have the following three characteristics:it should be able to show clinically consistent symptoms at different stages after stress and exhibit a similar pathogenesis pattern to that of clinical stress disorders,i.e.,good epiphenomenal validity;at the same time,the pathophysiological mechanisms(e.g.,glucocorticoid changes)of the model animals should be consistent with clinical ones,i.e.,good structural validity;finally,the existing therapeutic drugs should be able to perform on the model.Finally,the available drugs can show therapeutic effects on the model,i.e.,good predictive validity.The progression of ASD to PTSD involves multiple biological mechanisms.The mainstream view is that the enhanced consolidation or fading of stress-related memories in the early post-traumatic phase is one of the key reasons for the difficulty in resolving ASD symptoms and thus the development of PTSD.NMD A receptors are also important targets of stress hormones and are one of the important factors mediating the enhanced or diminished consolidation of traumatic memory.Therefore,regulation of NMD A receptor function may have a role in improving stress disorders.However,whether agonism or antagonism of NMDA receptors has an ameliorative effect remains controversial,with both studies in which antagonism of NMD A receptors(ketamine)ameliorates PTSD symptoms and studies in which enhancement of NMDA receptors(D-serine)ameliorates them,and there is a lack of studies related to the effect of lack of NMDA receptor function on the progression of ASD to PTSD.In summary,this study consists of the following three parts:first,an animal model of the development of ASD to PTSD was established,and the model was evaluated based on the dynamic changes of behavioral and pathophysiological indicators in clinical patients after trauma exposure.Then,the established animal model was used to observe the therapeutic effects of the existing PTSD treatment drug Sertraline(Ser),and to preliminarily evaluate the effects and possible mechanisms of the hookwort-derived alkaloid 297(compound 297)on the development of ASD to PTSD,which was screened in the laboratory.Finally,the effects of antagonists and agonists of NMDA receptors and their subtypes on the development of ASD to PTSD were observed on the established model,with the aim of elucidating the role of the functions of NMDA receptors and their subtypes in the development of ASD to PTSD.1.Establishment of a model for the development of acute stress disorder to post-traumatic stress disorderTo establish a mouse model for the development of ASD to PTSD and to provide an animal model for the study of stress disorder mechanism and evaluation of prevention and treatment drugs.Mice were stressed using electric foot-shock for three consecutive days,and behavioral experiments in the acute phase were conducted from day 1 to day 3 after stress;the chronic phase was conducted from day 28 to day 30.The behavioral experiments consisted of an open-field test,a contextual fear text,and a trauma-related cue avoidance test.Using behavioral profile analysis,the most representative index of each behavior was selected,and the "mean ±0.5 times the standard deviation" of the control group data was used as the "critical value" of the selected index.The mice with all three indexes exceeding the threshold value were considered as diseased mice.The results showed that the mice in the model group showed significant active avoidance and re-experiencing behaviors in both the acute and chronic phases;anxiety-like behaviors appeared in the acute phase,but disappeared in the chronic phase;there was a positive correlation between active avoidance and re-experiencing behaviors in the model group in the acute and chronic phases,respectively.The results of behavioral characteristics analysis showed that the incidence of ASD and PTSD in the model mice were 57.7%and 30.8%,respectively,which were similar to the clinical ones.The plasma corticosterone levels in the model mice were significantly higher than those in the normal control group half an hour after the end of stress,and the plasma corticosterone levels in the model mice were significantly lower than those in the normal control group on the 30th day after stress.The results of the above study showed that the acute and chronic phases of the model mice showed similar symptoms as the clinical ones,and the prevalence of ASD and PTSD were similar to the clinical ones,indicating that the model has good apparent validity;the changes of plasma corticosterone levels in the model mice after stress were consistent with the clinical ones,indicating that the model has good structural validity.Therefore,this part of the study successfully established a model for the development of acute stress disorder to post-traumatic stress disorder in mice with good apparent validity and structural validity.2.Study of the effects of sertraline and compound 297 therapeutic administration on the development of stress disordersIn this part of the study,an established animal model was used,and the PTSD treatment drug SER was selected as the positive drug to observe the response of this model to the available treatment drugs,thus evaluating the predictive validity of the constructed model;and the effect of therapeutic administration of compound 297,which was pre-screened in the laboratory,on the development of stress disorder was also observed.In this experiment,the positive drug SER or compound 297 was administered to mice for a single time or for a long time(20 consecutive days)after the end of stress to observe their effects on the behavior and development of stress disorder,respectively.The results showed that single administration of SER had no significant effect on the development of stress disorder in the stressed mice;while the exploration time of mice in the trauma cue avoidance test was significantly higher in the SER long-term administration group than in the stressed group;the maximum startle amplitude in the startle reflex test was significantly lower than that in the stressed group,while the prevalence of PTSD and the conversion rate of ASD to PTSD were significantly lower than those in the stressed group.Single administration of compound 297 significantly increased the freezing time in the acute and chronic phase of situational fear experiments and tended to decrease the exploration time in the trauma cue avoidance test,while the prevalence of PTSD and the conversion rate of ASD to PTSD were higher in the stressed group.Chronic administration of compound 297 to the stressed mice tended to increase freezing time in the chronic phase contextual fear test,and the prevalence of PTSD and the conversion of ASD to PTSD were significantly higher than in the stressed group.Plasma corticosterone levels and the expression of NR2A,NR2B,BDNF and TrkB receptor proteins in hippocampal regions were not significantly different in any of the administered groups compared to the stress group.The above findings suggest that long-term administration of SER helps alleviate the active avoidance symptoms of PTSD and reduces the prevalence and conversion rate of PTSD,indicating that the model has good predictive validity;long-term administration of compound 297 may increase the prevalence of PTSD and the conversion rate of ASD to PTSD.3.Studies on the effect of NMDA receptor function on the development of stress disordersIn this part of the study,we used the established animal model to observe and analyze the role of NMDA receptor function in the development of stress disorder by administering different types of agonists or antagonists of NMDA receptors or their subtypes as instrumental drugs 30 minutes after each stressful episode.D-serine agonism of NMDA receptors,ifenprodil+D-serine combination agonism of NR2A receptors,PEAQX+D-serine combination agonism of NR2B receptors,memantine antagonism of NMDA receptors,PEAQX antagonism of NR2A receptors,and ifenprodil antagonism of NR2B receptors were used in the tests.It was found that neither agonism nor antagonism of NMDA receptors had any significant effect on the indexes related to the avoidance test and the freezing time of the contextual fear test in the acute and chronic phases of stress mice.In the acute phase of the trauma cue avoidance test,there was a tendency for the exploration time of mice in the ifenprodil,PEAQX and ifenprodil+D-serine groups to decrease compared with the model group;in the chronic phase,there was a tendency for the exploration time of mice in the ifenprodil,D-serine,PEAQX,ifenprodil+D-serine and PEAQX+D-serine groups to decrease compared with the model group.After using behavioral characteristics analysis,it was found that the prevalence of agonistic NR2A receptor ASD was significantly higher after each stress,which was statistically significant when compared with the stress control group;the prevalence of PTSD was significantly higher in all administered groups than in the stress group;and antagonizing NMDA receptor,antagonizing NR2B receptor,antagonizing NR2A receptor,agonistic NR2A receptor and agonistic NR2B receptor all elevated the prevalence of stress ASD to PTSD in mice.In addition,the results showed that neither agonism nor antagonism of NMDA receptors had any significant effects on plasma corticosterone levels and expression of NR2A,NR2B,BDNF and TrkB receptor proteins in the hippocampal region of stressed mice during the chronic phase.The above findings suggest that either agonism or antagonism of NMDA receptors or their subtypes may cause aggravation of active avoidance symptoms in acute and chronic phases of stress mice,and enhance the prevalence and conversion rate of PTSD in the chronic phase.4.Conclusion(1)Three consecutive days of electric foot-shock induced anxiety-like behavior,active avoidance behavior,and re-experiencing behavior in mice during the acute and chronic phases after stress.The behavioral characteristics showed that the prevalence of chronic phase in the stressed mice tended to decrease compared with the acute phase,and the mice with ASD symptoms might be more likely to develop PTSD,which was similar to the clinical pathogenesis pattern,indicating that the model had good epistemic validity.The plasma corticosterone levels in the acute phase after stress increased significantly in the model mice,and decreased significantly in the chronic phase,which is consistent with the clinical pattern,indicating that the model has good structural validity.The long-term administration of PTSD treatment drug sertraline can effectively alleviate the active avoidance behavior of stress mice,and reduce the prevalence and conversion rate of PTSD,indicating that the model has good predictive validity.(2)Based on the developmental model of ASD to PTSD established in this study,a single intraperitoneal injection of compound 297 after the end of stress may worsen the re-experiencing and active avoidance symptoms in the acute and chronic phases of stress mice and increase the prevalence of PTSD and the conversion rate of ASD to PTSD in stress mice;long-term administration of compound 297 after the end of stress may worsen the re-experiencing and active avoidance symptoms in the chronic phase of stress and increase the prevalence of PTSD in stress mice.The mechanisms involved need to be further investigated.(3)Based on the ASD to PTSD development model established in this study,modulating the function of NMDA receptors or their subtypes immediately after each stress,either by enhancing or inhibiting their function,may worsen the active avoidance symptoms in stressed mice and increase the prevalence and conversion rate of PTSD in the chronic phase,and the relevant mechanisms involved need to be further investigated.