Study On The Characteristics And Pathological Mechanism Of PTSD Induced By Combined Stress In Adult Rat

Posttraumatic stress disorder(PTSD) is a permanent chronic psychological dysfunctioncaused by stressor such as trauma. The main affairs leading to PTSD is life-threatening,death, severe injury, terrors and intensive fear or helpless feeling. Although body isWell-tended appearance, the patients of PTSD suffered torture psychiatrically, even lossnormal function and pose a huge burden on society. Currently, the occurrence of PTSDexhibited an increasing tendency due to severe natural disaster, war, terrors and otherstressing affairs. But there are not effective measures to prevent and cure the disease. Somany governments and militaries all over the world pay more attention to the disease and itbecome the hot-point of researching fields.The beginning research of of PTSD is late, but the progress is rapid. There areremained some basic questions to be solved in the current:1. As a necessary experimentaltool for PTSD research, the establishment of aminal models is interested by researchers.But there is still no one widely accepted animal models of PTSD. Several animals of PTSDhave been established in previous researches, such as single-prolonged stress(SPS) model.Because of the limit of animals models in some degree, it is urgent to create a valid modelsto mimic the occurrence and development of PTSD in human.2. The mechanism of PTSDoccurrence have been made some progresses, but still exists many questions to be clarified.For example, the effects of different region of brain in the happen of PTSD is worth to bediscuss deeply. At the same time, there are some conflicted aspects in current researchingresults. Hippocampus is closely relevent to learning and memory, besides it is an importantstress-regulated region in the central nervous system. Many researches indicated thathippocampus may play crucial effects in develop of PTSD. Some researchers found thechange of hippocampal volume in patients with PTSD, but the detailed characteristics andmechanism of which in PTSD developing process are not clear and need to be deeplystudied.3. The regulation of neuroendocrine system plays key roles in occurrence of PTSD. There are more reports about hypothalamic-pituitary-adrenal (HPA) axis, but the researchof glutamate (Glu)/gamma-aminobutyric acid (GABA) is emerging and attractingincreasingly. Glutamine is an excitory neurotransmitter generally existing in the brain,which can form consciousness and memory through sensory afferent. GABA is aninhibitory neurotransmitter. The balance of Glu/GABA is remarkably important formaintaining the function of brain. During excessive stress, GABA decreased and Gluexhibited hyper-excitable activity, which led to the injury of hippocampal neurons andstrengthen trauma-related memories. It is concluded that: the dysfunction of Glu/GABA andtheir receptors may be one of the considerable mechanisms of PTSD pathological process.There are still to be confirmed through experiments.In order to solve above questions, a new PTSD model–combined stress model, inadult rats was established through thinking of the advantages in current several PTSDanimal models. The method is as follows: First, the adult rats were restrained for2hoursaccompanying inevitable electrical stimulation (4mA,60V) lasting2seconds withrandom intervals of30-120seconds. After restraining, rats proceeded fatigue swimming(water temperature is20℃) up to sink into the bottom of water for30seconds. Aftermodeling, the rats were regularly feed in cages and freely drink water. SPS modelestablished according to regular method is for positive control and normal rats for negativecontrol. All rats for experiment were divided into normal group, SPS model group andcombined stress model group after one-week accommodation. Specialized laboratoryassistant observed the general behaviors with four weeks after modeling. To avoid theinfluence of acute stress reaction, the examination of system behavior, learning andmemory functions, pathological morphology were carried out at14day after stimiulation.Furtherly, the change of pathological characteristic in important region of brain related toPTSD development and the relationship of Glu/GABA transmitters and their receptors inhippocampus was discussed. Through above measurements, it can provide theoreticalfoundation and experimental proofs for thorough researches on mechanism and drugs forprevention and treatment PTSD in the future.Results and conlusions:1. Study on the behaviors, learning and memory: The results of qualitation andquantitation were as fellows:(1) Behaviors in general: Responses to the capture of combined stress group were higher than the results in control group(P＜0.01)，and alsoshowed higher than those in the SPS group(P＜0.05), which indicated that the rats incombined stress group was prone to attacking and escaping behaviors than the other twogroups.(2) Open field test: The rats in combined stress group showed significant differencein the crossing and standing behaviors，comparing to the control(P＜0.01)and SPS groups(P＜0.05)，while no difference in the grooming behaviors, it indicated that the rats incombined stress group showed less locomotors and more anxiety.(3) Elevated plus mazetest: There were clear differences in the OE%，OT%and OE+CE between the cominedstress group and control group(P＜0.01), and the results of OE%，OT%and OE+CEdisplayed different between the complex stress group and SPS group(P＜0.05)， nodifference were found in dipping head or standing, which showed the rats of comined stressgroup reduced the locomotors and explained more anxious in mood.(4) Water maze test:No difference were found on the first escape latency of the1st and2nd day on the test (16thand17th after stress), the escape latency was prolonged on the3rd and4th day(18th and19th after stress) between the comined stress group and normal group(P＜0.01); nosignificant difference was found between the comined stress group and SPS group, but thelatencies on3rd and4th day were increased remarkably indicating that the space memory ofrat PTSD model was damaged(P＜0.05), furthermore, the space memory in rat ofcomined stress group was injured more obviously.(5)Water maze exploration test: Therewere differences on the times of target quadrant and crossing platform in combined stressgroup than in normal group(P＜0.01), more significant changes were found in rats ofcomined stress group than those in SPS groups(P＜0.05), it indicated that the spacememory of rats in compined stress group was affected more severely. As a result, thecombined stress rats might mimic the clinical symptom of PTSD patients and the combinedstress model is an easy and ideal animal model for PTSD.2. Study on the pathomorphology：The pathomorphologies on cortex and hippocampusof rat in control, SPS and combined stress groups on the14th day after stress were observedwith H.E, Nissl and Holmes staining. Almost no clear changes were found in the cortex ofthree groups, no neuronal degeneration was found. In the control group, the neurons ofsubregion in hippocampus were well distributed, the granular cells of DG were in the closeset, no cell prominency were found; the cell bodies in the CA3were comparatively large and more cross sections of dendrite were evident in the stratum radiatum; the cell bodies inthe CA1region distributed regularly, and the cross sections of dendrite were seen. The celldensity of CA3and DG regions in the combined stress group decreased than control group,and the deeply dying and irregular particles of subregions might be the fragment of deadcell nucleus. Intercellular space of CA3region was widened, the DG region showedunderstain in cluster. As to the Nissl’s staning, the cells in PTSD group were in line and thecell bodies were round or in arris, the cell bodies were expanded, the cell membranes wereuncompleted, the Nissl’s bodies were understained in different degrees. In some cells, theNissl’s bodies completely disappeared, but no differences were found in the combinedstress and SPS groups. In the studies of Holmes staining, the hippocampus neurons inPTSD group damaged more clearly, the axonotmesis was found, and also the nerve fiberwere found short and lined untidily. No significant differences were found in SPS and CSgroups. These results indicated that the neuron degeneration took place in the hippocampusof PTSD model, and the changes in the CA3and DG regions were more obviously,furthermore, the pathological lesion in the rat of combined stress group was more severity.3. The study of the pathogenesis：The changes of receptors of Glu/GABA in the cortexand hippocampus among three groups. The results were as follows:(1)The concentration ofGlu in hippocampus of PTSD model increased more evidently than that in control group onthe14th day after stress, and the concentration of combined groups was higher than that ofSPS group(P＜0.05)；the content of GABA in hippocampus was lower than that in thecontrol group, but no significant difference was found between combined stress group andSPS group.(2)The concentration of GluR2expression of hippocampus in PTSD modelsdecreased compared with that in control group, the result in combined stress groupdecreased more obviously than that in SPS group(P＜0.05).The expression of GABAARγ2was down regulated，but there was no difference between SPS and combined stress groups.These results showed that dysfunction of Glu/GABA system may be one of the PTSDpathogenesis.