| Recently,people feel heavy stress generally because of the influence of social work, interpersonal relationship and economic pressure,along with the rapid development of the society and the economy.People pay attention to mitis mental disease such as neurosis from serious mental disease as if schizophrenic in 21 century.The occurrence of anxiety as a subtype of neurosis is increasing year by year.Anxiety which harms people'health becomes increasingly a severe mental disease.The object of this paper layings a Chinese materia medica prescription of treating anxiety----GUANGUIDINGZHIFANG(GGDZ),by consulting a lot of pertinent literature and paleo-modern TCM famous doctors' medicational experience,referring to technological research findings of modern pharmacology,combining clinical and market requirement.This prescription is on the base of composing principles of soothing liver and sending down fire, restoring normal coordination between heart and hidney,relieving convulsion and anchoring the mind.We adopt the international universal anxiety animal models to investigate antianxiety effect of GGDZ.We apply Vogel's conflict test to approach possible GABA nervous pathway of GGDZ antianxiety mechanism.To aim directly at the deficiency of received anxiety animal model of TCM disease-syndrome,this topic explores initially to build a anxiety animal model of TCM disease-syndrome by consulting literature and combining protophase experimental crocidismus.1.Pharmacodynamics research1.1 Effects of GGDZ on the mice EMP testObjective:to observe the antianxiety effect of GGDZ on the mice EMP test.Methods:to adopt the mice EMP model,mice is divided into model,DZP,GGDZâ… , GGDZâ…¡,GGDZâ…¢five groups randomly.GGDZâ… ,â…¡,â…¢groups are administrated orally with 2.5g/kg/d,5.0g/kg/d,10.0g/kg/d respectively,DZP group 2.0mg/kg/d,model group equivalent saline,they are administrated 10 days continuously,the praxiological test, observation,and comparing various praxiological index on the tenth day.Results:DZP and GGDZâ…¡groups can increase the open entries percent(OE%) and open times percent(OT%) significantly,but not raise total entries(TE).Conclusion:GGDZ in the dosage of 5.0 g/kg/d does have antianxiety effect,but its effect doesn't strengthen following the dosage augmentation or decrease.It shows initially that GGDZ has certain intervention on mice anxious behavior.1.2 Effects of GGDZ on the mice light-dark box testObjective:to observe the anti-anxiety effect of GGDZ on the mice light-dark box test.Methods:to adopt the mice light-dark box model,groups and administration are the same as the above test.Mice is administrated 10 days continuously,the praxiological test is carried out on the tenth day.Results:DZP and GGDZâ…¡groups can increase the number of transitions and light times, GGDZâ…¢group can also increase light times significantly.Conclusion:GGDZ does have antianxiety effect in the dosage of ranging from 5.0 g/kg/d to 10.0 g/kg/d,which is the same as DZP effective direction.According to various index in light-dark box test mice,the 5.0 g/kg/d dosage is the best.That is similar to the result of EPM.1.3 Effects of GGDZ on the rat EMP testObjective:to verificate repeatly the antianxiety effect of GGDZ on the rats EMP test by using different animal.Methods:to adopt the rat EMP model,rats are divided into model,DZP, GGDZâ… ,GGDZâ…¡,GGDZâ…¢five groups randomly.GGDZâ… ,â…¡,â…¢groups are administrated orally with 2.0g/kg/d,4.0g/kg/d,8.0g/kg/d respectively,DZP group 1.0mg/kg/d, model group equivalent saline,they are administrated 10 days continuously,the praxiological test,observation,and comparing various praxiological index on the tenth day.Results:DZP and GGDZâ…¡groups can increase the open entries percent(OE%) and open times percent(OT%) significantly,but not raise total entries(TE).Conclusion:this result is the same as the mice EMP test.GGDZ has certain stationary antianxiety effect without increasing the activity of animal.1.4 Effects of GGDZ on the Vogel's conflict testObjective:the Vogel's conflict belongs to a conditional reflex animal model,but EMP is a nonconditional reflex animal model.This test adopts different model to verificate the stability and the reliability of GGDZ on the anti-anxiety effect.Methods:to adopt the Vogel's conflict model,groups and administration are the same as the above test.Rats are administrated 10 days continuously,and they are prohibitted drinking within 48 hours before the last administration,carried out to the un-punished train on the ninth day and the punished train on the tenth day.Results:DZP and GGDZâ…¡,â…¢groups can increase the numbers of licking and shocking very significantly,GGDZâ… can increase above two indexs significantly.Conclusion:GGDZ has better anti-anxiety effect,because All GGDZ groups can increase the numbers of licking and shocking.It proves that GGDZ is very sensitive to the Vogel's conflict model,so we adopt this model to approach effective mechanism of GGDZ.1.5 Effects of GGDZ on the animal model of TCM disease-syndrome testObjective:to adopt acute stress plus EPM to build the animal model of TCM disease-syndrome test.We investigate replicative success of animal model on the aspect of praxiology and GGDZ's effect on this model.Methods:to adopt the mice EMP model,rats are divided into modelâ… ,modelâ…¡,DZP, GGDZâ… ,GGDZâ…¡,GGDZâ…¢six groups randomly.GGDZâ… ,â…¡,â…¢groups are administrated orally with 2.0g/kg/d,4.0g/kg/d,8.0g/kg/d respectively,DZP group 2.0mg/kg/d, modelâ… ,modelâ…¡groups equivalent saline.The process of building the model is:beside modelâ… ,other groups rats are clamped the point of tails with sponge holding forceps,so that rats are infuriated to fight each other.They are stimulated about 30 minutes every time,one time every 4 hour,three times every day,and social-isolated in night.They are built model for 2 days continuously,carried out to EPM test on the tenth day.Results:symptoms after 2 days acute stress:walking frequently in the cage,frighten appearance,screaming,palpi pilosi erection when hearing sound slightly,irritability,tearing and biting each other after removed stimulation,increasing the numbers of defecation, decreasing the quantity of meat and drink;symptoms on EMP test:increasing the numbers of defecation and emiction about the animals of TCM disease-syndrome model.But on the whole, symptoms of DZP,GGDZâ…¡,groups are lighter than other groups.DZP and GGDZâ…¡groups can increase the open entries percent(OE%) and open times percent(OT%) significantly,but not raise total entries.Furthermore,OE%and OT%of modelâ…¡group are lower than that of modelâ… group significantly.Conclusion:symptoms of TCM disease-syndrome model is similar to clinical appearance of depressed syndrome.GGDZ has effect on this model.2.Mechanism study----effects of GGDZ on the GABA pathway2.1 Effects of GGDZ on the amino acid contents in rat brain tissueObjective:to observe the amino acid contents change in rat hippocampus of GGDZ on the Vogel's conflict test,which can appraise the GGDZ's effect on controlling the neuronal excitability.Methods:to adopt the Vogel's conflict model,groups and administration are the same as the above Vogel's conflict test.After finished the test,we put rats to death by decapitation, take out of hippocampus immediately in the ice plate,then detect the amino acid contents with automatic amino acid analyzer.Results:model group shows degression of GABA,Gly contents and augmentation of Glu/GABA ratio.DZP and GGDZâ…¡groups can increase GABA,Gly contents and decrease Glu/GABA ratio significantly.Conclusion:GGDZ can control the neuronal excitability by increasing GABA,Gly contents and decreasing Glu/GABA ratio.2.2 Effects of GGDZ on GAD expression in rat brain tissueObjective:to observe GAD expression in rat hippocampus of GGDZ on the Vogel's conflict test,which can appraise the GGDZ's effect on GABA synthesize.Methods:to adopt the Vogel's conflict model,groups and administration are the same as the above Vogel's conflict test.After finished the test,we put rats to death by decapitation, take out of hippocampus immediately in the ice plate,fix hippocampus with 10% formalin,then detect GAD expression with immunohistochemical method.Results:model group shows degression of GAD expression level.GAD expression level can be increased by DZP and GGDZ.Conclusion:GGDZ can increase GABA contents by upregulating GAD(GABA synthetase)expression.2.3 Effects of GGDZ on GABA-T activity in rat brain tissueObjective:to observe GABA-T activity in rat hippocampus of GGDZ on the Vogel's conflict test,which can appraise the GGDZ's effect on GABA metabolism.Methods:to adopt the Vogel's conflict model,groups and administration are the same as the above Vogel's conflict test.After finished the test,we put rats to death by decapitation, take out of hippocampus immediately in the ice plate,then detect GABA-T activity with ELISA method.Results:model group shows augmentation of GABA-T activity.GABA-T activity can be degraded by DZP and GGDZ.Conclusion:GGDZ can increase GABA contents by inhibiting GABA-T(GABA metabolic enzyme) activity.2.4 Effects of GGDZ on combining activity and combining numbers of GABA with GABAa receptor in rat brain tissueObjective:to observe combining activity and combining numbers of GABA with GABA_a receptor in rat hippocampus of GGDZ on the Vogel's conflict test,which can appraise the GGDZ's effect on the condition of GABA with GABA_a receptor.Methods:to adopt the Vogel's conflict model,groups and administration are the same as the above Vogel's conflict test.After finished the test,we put rats to death by decapitation, take out of hippocampus immediately in the ice plate,then detect combining activity and combining numbers of GABA with GABA_a receptor by ELISA method.Results:model group shows degression of combining activity and combining numbers of GABA with GABA_A receptor.Which can be increased by DZP and GGDZ.Conclusion:GGDZ has antianxiety effect by augmentation of combining activity and combining numbers of GABA with GABA_A receptor.2.5 Effects of GGDZ on the mice spontaneous locomotor activity testObjective:to observe that if GGDZ has sedative side effect by the mice spontaneous locomotor activity test.Methods:to adopt the mice spontaneous locomotor activity model,groups and administration are the same as the above mice test.Mice is administrated 10 days continuously, the praxiological test is carried out on the tenth day.Results:comparing with control group,GGDZ groups don't increase the numbers of spontaneous locomotor activity.There was nosignificant difference about spontaneous locomotor activity between groups.Conclusion:GGDZ does not have sedative side effect in the antianxiety dosage.ConclusionThe praxiology tests verify that GGDZ has stable antianxiety effect.The mechanism research tests prove that GGDZ has antianxiety effect by increasing the contents of GABA and Gly,decreasing Glu/GABA ratio,upregulaing GAD expression,downregulating GABA-T activity,enhancing the combining activity and combining numbers of GABA with GABAa receptor.GGDZ influences the GABA pathway to play a role in anti-anxiey. |
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