The Roles Of Neuronal Ensembles In Amygdala In PTSD-like Behavior In Rats

Objective:To explore the effect of different neuronal ensembles in amygdala in PTSD-like behavior,we use single prolonged stress(SPS)as the PTSD model,and integrate behavioral methods,chemogenetics,and immunofluorescence to dissect different neuronal ensembles in amygdala in the development of PTSD-like behaviors and provide theoretical basis for clinical treatments.Methods:Firstly,by using SPS model,the rats were randomly divided into 3groups,including the control,SPS 1d,and SPS 14 d.We used open field,elevated plus maze to test the behavioral change of anxiety,and used immunofluorescence to clarify expression of c-fos which represent the activity of neuron to explore the effect of amygdala activation in PTSD-like behavior.Secondly,we use Designer Receptors Exclusively Activated by Designer Drugs(DREADD)method to inactivate amygdala to verify its role in PTSD.Through microinjection with AAV,then we put the modified form of the human M4 muscarinic(h M4)receptor(h M4Di)into Ce A,which can be activated by CNO.We used open field,elevated plus maze to test the behavioral change of anxiety,to prove if the amygdala participates in PTSD.At last,we used GAD67 as GABAergic neuron marker,and Ca MKII as glutamatergic neuron maker,to label with c-fos,and then observed activation of different neuronal ensembles to reveal related neurobiological mechanism.Results:1.Compared to the control,the rats of SPS 14 d spent less time in central area of open field and in open arms of elevated,but no difference in SPS 1d,which mean SPS induced prolonged anxious behavior.The expression of c-fos in Ce A in SPS 14 d rats significantly rised,but had no difference compared with that in SPS 1d.Additionally,we did not find activation in BLA brain region in either of three groups.2.After inactivated by DREADD in Ce A,the rats of SPS 14 d spent more time in central area of open field and in open arms of elevated,which show an anxiolytic effect.3.After SPS,in immunofluorescence test,the ratio of GAD67+c-fos positive/ c-fos positive is significantly rising in Ce A of SPS 14 d group,comparing to that in the groups of control and SPS 1d.There is no significant differences among three groups in ratio of GAD67+c-fos positive/ c-fos positive in BLA.Furthermore the ratio of Ca MK?+c-fos positive / c-fos positive displayed no difference among three groups in both BLA and Ce A,which means no changes of activities of glutamatergic neurons during the development of PTSD-like behaviors.Conclusion:1.The occurrence of PTSD-like behaviors are accompanied with activation of Ce A but not BLA after SPS.2.Inactivation of Ce A alleviates anxiety-like behaviors of PTSD-like behaviors.3.The activations of GABAergic neuron in Ce A may involve in PTSD-like behavior,which should be verified by further chemogenetic experiments.