| In the 21st century, as the human lifespan and demograph are changing, the central nervous system of the disease will be increased significantly with age, including related neurodegenerative diseases ,such as Parkinson's disease ,alzheimer's disease and huntington etc. pathogenesis and pathological aspects have been systematically elaborated, however, this disease treatment strategy is still very limited with them , how the drug effectively transship to nervous system become the most challenging difficulty, enhancing drugs transshipment treatment strategy paid more attention. With degenerative disease of the nervous system and deepen understanding of biologics, We believe that we would have more tharapical opportunities. The research about drug delivery treatment in the central nervous system from laboratory to treatment in clinical practice plays an important role transformation. Parkinson's disease (Parkinson 's, PD) was common in central nervous system, whose pathological invariance is characterized with degeneration and depigmentation of dopaminergic neurons in nigra. The symptoms of rigidity, tremor and serious motionless, etc, are caused by Parkinson's disease . At present there is no method to cure Parkinson's disease. The treatments we grasped included levodopa, dopamine decarboxylase an enzyme inhibitors, catechu - oxygen - methyl transferred enzyme inhibitors, anticholinergic dopamine agonists,and single amine oxidase inhibitors, etc.They can improve symptoms. In addition, various growth factors make neural protection. Levodopa is a method to control clinical symptom.But long-term use will lead to "on-off" phenomenon, dyskinesia and psychiatric disorders.The main reason is the unstable drug concentration.Therefore seeking a stable concentration is significant to treatment. Traditional and new treatments need more suitable transport pathways that will play a role by importing the drug to central nervous system . There are many factors affecting or hindering drug delivery ,especially macromolecular drug delivery in the CNS. It's reported that the way of intranasal dopamine takes positive effect to the stability of dopamine concentration and the Parkinson's disease, but the mechanism is not clear at all. This study aims to observe the influence of intranasal dopamine to MPTP mice and normal dopaminergic neurons and discuss the possible mechanism.1 Through intranasal dopamine, the behavior of MPTP mice has been changed.Climbing pole experiments and hanging experiments can reflect the coordinated ability of mice's body.The more average time mice climbed the pole, the poorer coordinated ability mice have; the lower average hanging score mice attach,the worse body coordinated ability mice have. In climbing pole experiments of mice, we observed data of four groups (n=8) and two time points (the fifth day and the tenth day).In the fifth day, experimental data show that the average total time in the climbing pole test of normal mice is 12.00±0.15 ; the one of model group is 20.34±1.76 ;the one of molel-treatment group is 44.24 + 12.08 ;the one of normal group is 12.34±0.27. According to the statistical software data, there is significant differences with average time of climbing pole between molel group and model-treatment group (p<0.05). It's displayed that body coordination ability of model -treatment group is better than that of model group,by means that the climbing pole time of model- treatment group is shorter than that of model group.In the tenth day,experimental data show that the climbing pole time of normal group is 12.15±1.86;the one of model-treatment group is 26.79±4.24;the one of model group is 36.54 + 6.68;the one of normal-treatment group is 10.68 + 0.65. According to the statistical software data, the climbing pole time of model-treatment group is evidently shorter than that of model group, displaying the body coordination ability of model-treatment group is better than that of model group. Above all, the way of intranasal dopamine make positive effects on the body coordination ability to MPTP mice, indirectly displaying that intranasal dopamine make protecting and promoting role on dopaminergic neurons to MPTP mice.In hanging experiments, observing four groups'data (n = 8) on two time points (the fifth day and the tenth day). The fifth day's data show that the average hanging score of normal group is 3.00±0.00;the average hanging score of model-treatment group is 2.68±0.64; the average hanging score of model group is 1.47±0.68; the average hanging score of model group is 3.00±0.00. According to the statistical software data, the hanging score of model-treatment group is obviously higher than model group,displaying that the body coordination ability of model-treatment group is better than model group. The experimental data of the tenth day show that the average hanging score of normal group is 3.00±0.00;the average hanging score of model-treatment group is 2.35±1.00; the average hanging score of model group is 1.54±0.45; the average hanging score of model group is 3.00±0.00. Above all data also show that the way of intranasal dopamine make positive effects on the body coordination ability to MPTP mice, indirectly displaying intranasal dopamine make a protecting and promoting role on dopaminergic neurons to MPTP mice.2 The immunohistochemical changes of MPTP mice with intranasal dopamine Testing dopaminergic neurons with the way of fluorescence immune tissue chemical, TH immunochemical staining is the classical method reflecting dopaminergic neurons'quantity and shape. The application of immune- fluorescence can also check out the differences of dopaminergic neurons by the fluorescence degrees auxiliarily.According to all above, 4 group mice (n=8) after 10 days'treatment (including normal groups, MPTP model group, MPTP model-treatment group, normal group) were infusioned by polyphosphate,formaldehyded, implated by immunohistochemical staining, and then observed of dopaminergic neurons quantity, shape or fluorescence degrees including olfactory bulb,striatum and nigero under fluorescent microscope.As nigero for example, compared with control group (80.33 + 45.15), the number of positive TH-stained neurons with model group (40.15±10.35, p<0.05)is evidently smaller, the one of model–treatment group (60.25±3.55,p<0.05)is significantly bigger compared with model group. Above all the data, the way of intranasal dopamine is feasible and it has certain protected and promot function to dopaminergic neurons. Its mechanism is probably with exogenous effect, but whether it has endogenous effect remains to be proved.Compared control group (80.33 + 45.15) to treatment group (83.22 + 32.10) , it is no significant differences between them . Similarly, from the immunohistochemical results of olfactory bulb and striatal, there are differences between model-treament group and model group, showing the feasible way of intranasal dopamine and the protected effect of dopaminergic neurons.3 Intranasal dopamine of different doses lead to the changes of immunohistochemical ratsAfter inranasally giving different doses dopamine (1.5 mg/kg, 3mg/kg, 6mg/kg) of chronic long-range (14day), rats are perfusioned, observed TH positive neurons of olfactory bulb, the striatum ,nigero by fluorescence immunohistochemistry methods, and compared with control group . This paper observed the exogenous and endogenous changes and dose - effect relations of intranasal release.SD rats (n = 6) were divided into four groups in accordance with intranasal dopamine doses of 1.5 mg/kg, 3mg/kg, 6mg/kg, blank group (n = 6).Followed chronic long-range medicine-given method before, the animals were executed at last, and TH positive expression were observed by immune fluorescent chemical staining. Through the experimental results, dopaminergic neurons quantity, shape and immune fluorescence degrees have no significant differences between these four groups of 1.5 mg/kg, 3mg/kg, 6mg/kg, blank group . It hints: for normal physiology tissue, approaching effct to dopaminergic neurons is not significant through intranasal delivery; or the viewpoint of way that whether endogenous approaches promot dopaminergic neurons through tyrosine hydroxyl enzyme immunohistochemical staining was not confirmed to the intranasal dopamine mechanism.Conclusion1 Building an ideal model for exploring pathogenesis and seeking medical treatment is extremely important. At present, MPTP model of mice is the most widely used PD model. Previous studies confirm that MPTP model mice were matching clinical PD feature in organization chemistry and biochemistry and it is a kind of ideal PD model. In this experiment , kunming mice seleced were continuous given10d MPTP,in climbing pole and hanging,and their behavior all represent obstacle motion. TH is dopamine synthesis,it can be used as the speed limit enzyme symbol of dopaminergic neurons.In the experiment, model group's TH positive neurons significantly reduced.In two aspects of organization behavior and chemical preparation, this model is successful.2 In the experiment, after intranasal dopamine (0.12 mg/kg) ,it is found this way can significantly ease behavior obstacle on MPTP induced PD model .Meanwhile immunohistochemical method also showed intranasal dopamine eased dopaminergic neurons loss on olfactory bulb, the striatum and nigro, It is confirmed that intranasal dopamine has positive influence to nerve. 3 Different doses of intranasal dopamine dropping, it is showed no obvious difference of dopaminergic neurons in quantity and immune fluorescent histochemical staining among different groups by TH dyeing. Relevant reports said in the literature: intranasal dopamine can cause dopamine receptors and dopamine transhipment increased, and it has positive correlation with intranasal dopamine dose .It hints that the way of intranasal dopamine was probably in ralation with metabolic pathway of dopaminergic neurons, speculating that the way of intranasal dopamine does not significantly affect the metabolic way of tyrosine hydroxyl enzyme in the normal metabolic pathway. So it has no significant differences of the four groups in the experiment. |
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