Effects Of Enriched Environment On BDNF Expression In Midbrain Of The Rat Model Of Parkinson’s Disease

Background:Parkinson’s disease(PD)is a common neurodegenerative disease,which is characterized by pathological the death of dopaminergic neurons in the substantia nigra pars compacta and the accumulation of Lewy bodies.Its typical symptoms include tremor,muscle rigidity,bradykinesia,and postural abnormalities.At present,therapies of PD include drug therapy and surgical therapy,but none of them are successful to avoid the progression of degeneration.Studies have shown that enriched environment has a positive impact on neurodegenerative diseases such as Huntington’s disease,Alzheimer’s disease and Parkinson’s disease,but the mechanism remains unclear.Brain-derived neurotrophic factor(BDNF),a neurotrophic factor,plays an important role in maintaining neuronal function and structure,involved in the pathology of many neurodegenerative diseases.This study is to investigate the effects of enriched environment on BDNF expression in the midbrain of PD rats.Part I:Effects of 6-OHDA on behavior and dopaminergic neurons in midbrain of ratsObjective:To study the effects of 6-OHDA on on behavior and dopaminergic neurons in midbrain of rats and to establish an effective and stable 6-OHDA PD rat modelMethods:6-hydroxydopamine was injected to damage the striatum.At 2 weeks,3 weeks,9 weeks,and 12 weeks after operation,apomorphine-induced rotation test and openfield test were performed to evaluate changes of animals’ behavior.Nissl staining and tyrosine hydroxylase(TH)immunohistochemical staining were used to observe dopaminergic neurons in midbrain of rats.Results:(1)Apomorphine-induced rotation test:24 of 32 rats had more than 210 rotations within 30 minutes and this could last for at least 12 weeks.(2)Openfield testCompared with the sham-operated rats,the 6-OHDA group has less total travel distance(length),less total travel distance in the peripheral area(distl),less moving fast time in the peripheral area(MF1),or less average speed(Vmeanl)and more resting time in the peripheral area(RT1).After 6-OHDA injection,the activity of the rats in the open field during the 2nd week and the 3rd week was not statistically different(P>0.05).Rats showed decreased length,MF1,Vmeanl,dist2,MF2,Vmean2 at the 9th after6-OHDA injection than those at the 2nd week.Rats showed decreased length,MF1,Vmeanl,dist2,MF2,RT2%,RT2 at the 12th after6-OHDA injection than those at the 2nd week.Rats showed decreased length,MF1 and increased RT1 and RT1%at the 9th after6-OHDA injection than those at the 3rd week.Rats showed decreased length,distl,MF1,Vmeanl,RT2%,RT2,MF2 at the 12th after6-OHDA injection than those at the 3rd week.No difference was observed of activities in openfield test between rats at 9th week or 12th week.(3)Nissl staining:In the sham-operated group,cells in leisioned striatum and substantia nigra were densely arranged,and the cytoplasmic Nissl body was abundant,while cells in the 6-OHDA group were sparsely arranged.As the gap increases,the intracytoplasmic Nissl body was scarce.(4)TH immunohistochemical staining:Rats at the 2nd,3rd,9th,or the 12th week after injection of 6-OHDA showed decreased TH-positive fibers or cells in leisioned striatum and substantia nigra than those in the sham group(P<0.05).At the 3rd,9th,and 12th weeks after 6-OHDA injection,rats had less TH-positive fibers or cells in leisioned striatum and substantia nigra than those at the 2nd week(P<0.05).Conclusion:Injecting 6-OHDA into striatum induce an effective and stable PD rat model.The damage of 6-OHDA to dopaminergic neurons in midbrain of rats gradually stabilized 3 weeks after operation,and motor dysfunction of rats increased with time.Part II:Effects of enriched environment on BDNF expression in midbrain of the rat model of Parkinson’s diseaseObjective:To study effects of enriched environment on BDNF expression in midbrain of the rat model of Parkinson’s disease and try to find out the possible mechanism of enriched environmentMethods:Successful rat models of Parkinson’s disease induced by 6-hydroxydopamine(6-OHDA)were divided into 2 groups:6-OHDA+EE group and 6-OHDA+standard environment(SE)group.Rats accepted sham-operation were divided into 2 groups:Sham+EE group,sham+SE group.Six weeks after exposure to different environments,behavior tests were performed to evaluate animals’ behavior.Nissl staining and tyrosine hydroxylase(TH)immunohistochemical staining were performed to observe neurons in midbrain of rats.The level of brain-derived neurotrophic factor(BDNF)in midbrain was detected by western blotting.Results:(1)Successful PD model:14 of 20 rats accepted 6-OHDA injection were successful PD models,and the success rate was 70%.(2)Behavioral testsa.Apomorphine-induced rotation test:The number of rotation in 30 minutes of the 6-OHDA+EE and 6-OHDA+SE rats was 237.00± 17.70 cycles and 236.83±14.15 cycles,respectively.There was no significant difference between the two groups(t=0.019,P>0.05).No rotation was observed in sham+EE or sham+SE groups.b.Open-field test:Differences were found in length,distl,RT1,RT1%,MF1,and Vmeanl in the four groups of rats(P<0.05).Compared with rats in sham+EE group,Rats in 6-OHDA+EE group showed decreased length,distl,and Vmeanl(P<0.05).Compared with rats in sham+SE group,rats in 6-OHDA+SE group showed decreased length,distl,MF1 and Vmeanl,but increased RT1 and RT1%(P<0.05).Compared with rats in 6-OHDA+SE group,Rats in 6-OHDA+EE group showed increased length,distl and Vmeanl(P<0.05).No difference was observed between sham+EE group or sham+SE group.(3)Nissl stainingIn the leisioned striatum and substantia nigra of rats in the sham+EE and sham+SE groups,cells were densely arranged,with abundant cytoplasmic Nissl bodies.However,in the leisioned striatum and substantia nigra of rats in the 6-OHDA+EE and 6-OHDA+SE groups,the arrangement of cells was sparse,the intercellular space was increased,and Nissl body in the cytoplasm was scarce.(4)TH immunohistochemical stainingRats in the 6-OHDA+SE group,showed decreased TH-positive fibers or cells in the injured striatum and substantia nigra than those in the sham+SE group(P<0.05).Rats in the 6-OHDA+EE group,showed decreased TH-positive fibers or cells in the injured striatum and substantia nigra than those in the sham+EE group(P<0.05).Rats in the 6-OHDA+EE group,showed increased TH-positive fibers or cells in the injured striatum and substantia nigra than those in the 6-OHDA+SE group(P<0.05).Rats in the sham+SE group,showed decreased TH-positive fibers or cells in the injured striatum and substantia nigra than those in the sham+EE group(P<0.05).(5)BDNF expression in the midbrain of rats exposed to different environmentsThe levels of BDNF in the injured striatum and substantia nigra of rats in 6-OHDA+SE group were lower than those in sham+SE group(P<0.05).The levels of BDNF in the injured striatum and substantia nigra of rats in 6-OHDA+EE group were lower than those in sham+EE group(P<0.05).The levels of BDNF in the injured striatum and substantia nigra of rats in 6-OHDA+EE group were higher than those in 6-OHDA+SE group(P<0.05).The levels of BDNF in the injured striatum and substantia nigra of rats in sham+SE group were lower than those in sham+EE group(P<0.05).Conclusion:Enriched environmental may improve motor function and promoting recovery of dopaminergic neurons in rat models of Parkinson’s disease by up-regulating BDNF in the midbrain of rats.