The Expression of TH、 c-fos and Its Transcriptional Regulation in Locus Coeruleus and Adrenal Medulla on an Experimental Rat Model of Liver Depression Syndrome
|School||Southern Medical University,|
|Course||Traditional Chinese Medicine|
|Keywords||Liver depression syndrome Chronic constraint stress Locus coerule us adrenal medulla Tyrosine hydroxylase C-fos Xiaoyao Powder|
BackgroundSyndrome differentiation and treatment are the essence of the Traditional Chinese Medicine.Being the most prepotentest part of TCM, its procedure the overall concept and individualization have been valued and the philosophy of diagnosis and treatment based on an overall analysis of the illness and the patient’s condition has always taken its effects and made progression in the years of practice. Syndrome,which bases on the theory of Zang and its outer signs in Chinese medicine and insturcts the clinical diagnosis and treatment, has been paid attention in studying of traditional Chinese medicine in recent years. So,the study of nature of syndrome will surely impetus the development of TCM theory and clinical practice, as well as the modernization of TCM to some extend.Liver depression syndrome is a basic syndrome in TCM, which is characteristic of liver-qi stagnation because of the dysfunction and insufficiency of abreaction of liver-qi. As basic syndrome of liver diseases of Chinese medicine is concerned, liver-qi stagnation syndrome is one of common syndromes in clinical practice and highly valuable for profound research. To explore the essence of this syndrome will,as a result,give some new inspiriation in the prevention and treatment of these diseases.Furthermore, by the study o fthis syndrome we may develop an appropriate method in the disclosing the nature of other syndromes in TCM.The nature of liver-qi stagnation syndrome is the focus of TCM theory study today. Some researches indicate that liver-qi stangation snydrome is likely to result from regulating function disorder of high nevrous centre induced by negative psychic stress in which pathological changes of mutfi-system including nevre,endocrine, circulation,digestion,immunity, feeling and motion etc are involved. Now, hypothala-mo-pituitary-adrenal and locus coeruleus-sympathetic-adrenal medulla axis are comprehensively studied from abroad in stress reaction. But in China the study was focus on hypothalamo-pituitary-adrenal axis、hypothalamic-pituitary-thyroid axis or hypothalamic-pituitary-gonadal axis,and the study on LC-NE is very poor. Therefore, we hope to make a contribution to the essence of liver depression syndrome and basic theory of TCM by research into locus coeruleus-sympathetic-adrenal medulla axis.ObjectivesTo set up the effective rat model of liver depression syndrome from chronic constraint stress to find differentia of weight,behavioristics,orrhology and the expression of TH and c-fos in locus coeruleus and adrenal medulla between the rat model and the normal control, and between the rat model and rat treated by Xiaoyao Powder through gene chip.On one hand to research function mechanism of Xiaoyao Powder, on the other hand to explain the essence of liver depression syndrome.Establishment of the model rats:Thirty-six male SD rats(150±20g)were randomly divided into three groups with 12 rats in each after one-week’s adaptation living,which are group A(blank group),group B(model plus Xiaoyao Powder group) and group C(model group).The rats from the group B and C were housed individually in cages while rats from group A were living in two cages averagely.Rats from group B and C were bind in a specific bound frame,which may give chronic immobilization stress every day for 1 hrs to 6 hrs but at unpredictable time, to simulate the cause of depressed emotion.The upset continued for 21 days, and in the restraining moment the rats were deprived from diet or water.Rats from group B were also given Xiaoyao Powder by intragastric administration once a day at eight for consecutive 21 days,while the other two groups were given 2ml physiological saline with the same ways.Dose level for Xiaoyao Powder in the rats was set according to the day dose for a normal adult;it was lOg/kg body weight everyday.We may evaluate the model by the observation of behavior, fluid consumptiontest,laboratory examination and the curative effects. The brain stem and adrenals were properly removed,sliced and stained with hestaining to determine the morphological changes of LC and adrenal medulla.Immunol histochemistry was used to detect the changes of TH, c-fos expression in LC and adrenal medulla, Imagepro plus analysis system was used to determine the changes of staining intensity and the number of TH, c-fos-positive neurons in 3 groups.RT-PCR was performed to determine the changes of TH, c-fos and their mRNA expression in LC and adrenal medulla of rats. The obtained data were statistically analyzed with SPSS 13.0 software.Results1. The model rats displayed the typical appearance such as low activity,slow reaction,taper cry,reduces food and drink,like adherence,and so on.Rats from group B also showed some above abnormalities,but they were not obvious.Rats in group A lived normally.The rats’weight increase was significantly different between groups(P=0.000).It showed rats in group C grew slowest, while rats in group B the second lowest.Fluid consumption test revealed that group C lost appetite for sweet water during the whole procession. Statistics showed there were significant differences between these four groups(P=0.000).The erect time,grooming time and traversing grid also reduced in group C(P<0.01). The serum level of NE was also increased in rat model(P<0.01).2. By HE staining,no pathological changes were observed in LC and adrenal medulla of group B or group C rats.3. Expression results of Immunohistochemical detection of LC and adrenal medulla showed that, the number and mean density of TH and c-fos-positive cells in group C were higher than others(P<0.05).4.The expression of THmRNA and c-fosmRNA by RT-PCR was observed that both were upmost in group C(P< 0.05),while rats in group B was second upmost.The expression of TH was consistant with that of c-fos in chronic constraint stress.ConclusionThe rat model of liver depression syndrome from chronic constraint stress is reliable and credible.The present study demonstrated TH was overexpressed in LC and adrenal medulla in different time periods after stress.This indicates that a lot of catecholamine may be released from LC and adrenal medulla in response to the chronic stress.The released catecholamine can affect the related receptors or targeted organs to adapt to the reactions of stress through regulating the functions of nervous system and metabolism.C-fos expression also increased in LC and adrenal medulla, and c-fos and TH coexisted in the same cells.In addition,c-fos had the binding site on TH promoter,so it might be involved in the regulation of TH expression during the period of stress.It is firstly reported that locus coeruleus-sympathetic-adrenal medulla axis is greatly relational to liver depression syndrome.The present study provides a theoretical base to study the molecular mechanisms and give a beneficial trial for deeper research of the essence of liver depression syndrome.