In order to explore the role of acetylcholine in the pathogenesis of Parkinson's disease (PD), the changes in the concentration of acetylcholine (Ach) in the striatum, the apoptosis of substantia nigra cells, the ultrastructure and the changes of Nissl cells in rats during the morbidity of PD, and the corresponding behaviors in rats with PD were observed. Rat PD model was established by using the modified Thomas method. Eighty-one rats were randomly divided into normal control, sham operation and PD groups and their behavior features were observed at post-operative day (POD) 7, 14 and 21 as three subgroups (n=9 each). The concentration of Ach in the striatum was determined by using high-performance liquid chromatography. The apoptosis of substantia nigra cells was assayed by using TUNEL method. The ultrastructural changes in the substantia nigra were observed under the electron microscopy, and the survival of neurons in the substantia nigra area was examined by using Nissl staining. In PD group at POD 7 to 21, the damage in the substantia nigra area was gradually aggravated, the concentration of Ach, apoptosis rate and turns of rotation were gradually increased, and the number of Nissl cells was gradually reduced over the time as compared with the normal control and sham operation groups (all P<0.05). It was concluded that there exist dynamic changes in Ach concentration, ethology and apoptosis of the substantia nigra cells during the morbidity of PD, suggesting the contribution of apoptosis to the morbidity of PD, and critical role of Ach in the pathogenesis of PD.
Acetylcholine ; pharmacology ; Animals ; Corpus Striatum ; drug effects ; metabolism ; pathology ; Disease Models, Animal ; Male ; Parkinson Disease ; metabolism ; pathology ; Rats ; Rats, Wistar

In order to explore the role of acetylcholine in the pathogenesis of Parkinson's disease (PD), the changes in the concentration of acetylcholine (Ach) in the striatum, the apoptosis of substantia nigra cells, the ultrastructure and the changes of Nissl cells in rats during the morbidity of PD, and the corresponding behaviors in rats with PD were observed. Rat PD model was established by using the modified Thomas method. Eighty-one rats were randomly divided into normal control, sham operation and PD groups and their behavior features were observed at post-operative day (POD) 7, 14 and 21 as three subgroups (n=9 each). The concentration of Ach in the striatum was determined by using high-performance liquid chromatography. The apoptosis of substantia nigra cells was assayed by using TUNEL method. The ultrastructural changes in the substantia nigra were observed under the electron microscopy, and the survival of neurons in the substantia nigra area was examined by using Nissl staining. In PD group at POD 7 to 21, the damage in the substantia nigra area was gradually aggravated, the concentration of Ach, apoptosis rate and turns of rotation were gradually increased, and the number of Nissl cells was gradually reduced over the time as compared with the normal control and sham operation groups (all P<0.05). It was concluded that there exist dynamic changes in Ach concentration, ethology and apoptosis of the substantia nigra cells during the morbidity of PD, suggesting the contribution of apoptosis to the morbidity of PD, and critical role of Ach in the pathogenesis of PD.

Objective To study the effect of mild brain hyothermia on cerebral ischemic injury. Methods Global cerebral ischemia was established by modified Pulsinelli 4-vessel occlusion model. Forty-eight Sprague-Dawley rats were divided into 4 group: a sham-operated group, a normothermia (37～38℃) ischemic group and a mild is-chemic hypothermia (31～32℃) group; the mild ischemic hypothermia was subdivided into 4 groups with the hypo-thermia lasting for 30 min, 60 min, 120 min and 240 min, respectively. After 240 rain of reperfusion following 20 min cerebral ischemia, the levels of nitric oxide products nitrite (NO2) ,endothelin-1 (ET1) , tumor necrosis fac-tor alpha (TNFα) and interleukin-1 beta (IL-1β) in brain tissue and the lactate dehydrogenase (LDH), aspartate aminotransferase(AST) , creatine kinase(CK) and its brain band isoenzyme (CK-BB) in plasma were measured. Results The levels of IL-1β,TNFα, ET1 and NO2. in brain tissue, and the amounts of LDH, AST, CK and CK-BB in serum were higher in normothermia ischemic group than those in sham-operated group (P <0.05). Mild hypother-mia lasting for 60 min to 240 min markedly decreased the levels of IL-1β, TNF-α, ET1 and NO2 in brain tissue, and the amounts of LDH, AST, CK and CK-BB in serum in normothermia ischemic group, when compared with normo-thermia ischemic group (P < 0.05 or P < 0.01). Mild hypothermia lasting for 30 min did not influence the content of IL-1β, TNF-α, ET1 and NO2 in brain tissue when compared with normothermia isehemia group (P > 0.05). Con-clusion Mild brain hypothermia post-ischemia can significantly suppress the inflammation response in ischemic brain tissue and stabilize the function of cell membrane. The best neuroprotection of mild brain hypothermia must be carried out immediately and last for more than 60 minutes.

BACKGROUND: It is indicated in the study of recent years that gingko biloba extract (EGB) is a kind of natural cleaner of free radical and it protects the body from the damage induced by free radical and improves cerebral circulatory disturbance and neuronal function. But the experimental or clinical study on the effects of EGB on high neural functional activity, like cognition, is relatively lagged.OBJECTIVE: To probe into the function of EGB on high functional activity in central neural system so as to provide the experimental evidence on clinical application of EGB in treatment of cognitive disturbance.DESIGN: Randomized controlled experiment was designed.SETTING: Department of Geriatrics of Psychiatric Hospital affiliated to Tongji Medical College of Huazhong University of Science and Technology,Department of Pathophysiology in Tongji Medical College of Huazhong University of Science and Technology and Department of Neurology in Union Hospital affiliated to Jinan Medical College of Huazhong University of Science and Technology.MATERIALS: The experiment was performed in Basic Department of Tongji Medical College of Huazhong University of Science and Technology in June 2002. Forty Wistar rats were employed and randomized into 5groups, named as normal control of aged rats (normal group), model group,EGB 75 mg/kg group, EGB 150 mg/kg group and EGB 500 mg/kg group, 8 rats in each one.METHODS: Scopolamine was used to induce disturbance of learning and memory in aged rats to simulate the model of senile dementia animals. In normal and model groups, physiological saline of same volume was used for gastric perfusion and in every EGB group, EGB of 75, 150 and 500 mg/kg was used for gastric perfusion successively, 50-400 g/time, continuously for 5 days. On the 6th day, water maze and evading-dark-room tests were performed. During the testing, the medical perfusion stopped. The assay methods of behavioral training of learning and memory, such as experiment with water maze and evading-dark-room test, and biochemical assay were used to observe the changes in learning and memory and in acetylcholine (Ach) and protein contents in cerebral hippocampus before and after medication.MAIN OUTCOME MEASURES: ① Time required in maze test of rats in each group. ② Mistakes in maze test of rats in each group. ③ Time required and mistakes in evading-dark-room test of rats in each group. ④Contents of Ach and protein in cerebral hippocampus of rats in each group.RFSULTS: Except that 1 rat was died without definite reason in EGB 150 mg/kg group and 1 rat was escaped in either EGB 75 mg/kg or 500 mg/kg group during gastric perfusion, terminally, 37 rats entered result analysis.① The time required and mistakes in maze test in every EGB group were less remarkably than model group (P＜0.05 or P＜0.01). The time required and mistakes in maze test in model group were higher remarkably than normal group (P＜0.01). ② In learning of passive escaping in evading-darkroom test, the duration of learning for the rats in EGB 500, 150, 75 mg/kg groups was shorter remarkably than that in model group [(156.78±25.97),(172.66±13.56), (198.54±17.12), (208.34±28.56) s, P ＜ 0.05 or P＜0.01].The mistakes of electric shock in EGB 500, 150, 75 mg/kg groups were less remarkably than model group [(3.41±0.26), (6.97±0.35), (7.23±0.62),(8.38±0.92) times, P＜0.01]. The times of electric shock in EGB 500 mg/kg group was less significantly than 150 mg/kg group (P＜0.01) and that in 150 mg/kg was less remarkably than 75 mg/kg group (P＜0.05). ③ Hippocampal Ach content in modeled rats in EGB 500, 150, 75 mg/kg groups was higher than that in model group [(421.89±36.32), (387.45±32.76),(380.17±41.25), (365.28±11.42) μg/g, P＜0.05 or P＜0.01]. Hippocampal Ach content in 500 mg/kg group was higher significantly than 150 and 75 mg/kg groups (P＜0.01). In addition, compared with normal group,protein content in hippocampus in rats with disturbance of learning and memory induced by scopolamine in model group was reduced significantly [(41.75±3.82), (95.13±6.34) mg/kg, P ＜ 0.01]. After administrated with EGB,even though the protein content in hippocampus was increased in experimental rats after modeling, the difference was not significant (P＞0.05).CONCLUSION: EGB improves significantly learning and memory in experimental animal in dose-dependence and increases significantly Ach content in hippocampus.

By using Fura-2/AM, the effects of magnesium (Mg2+) on the glutamate-induced increase of intracellular free calcium ([Ca2+]i) in the cultured hippocampal neurons and the features were investigated by integrated photoelectric detecting system. The experiments were designed to three groups (The drug was spit to the cells for 20 s): Group A receiving 1 x 10(-5) mol/L glutamate; Group B receiving 1 x 10(-5) mol/L glutamate and 1 x 10(-5) mol/L Mg2+ simultaneously; Group C receiving 1 x 10(-5) mol/L glutamate again after [Ca2+]i in group B back to the baseline. The results showed that in group A, [Ca2+]i was obviously increased. In group B, the changes in [Ca+] i and the peak value were significantly decreased. Moreover, the elevation of Phase 1 was slowed down and Phase 2 was shortened to some extent, and the plateau phase between them was relatively prolonged. In group C, calcium oscillation similar to that in group A occurred, but both the Phase 1 and Phase 2 were shortened and the delta[Ca2+]i was slightly decreased. It was suggested that Mg2+ could quickly inhibit the rise of [Ca2+]i induced by glutamate in the cultured hippocampal neurons in rats.
Animals ; Animals, Newborn ; Biological Transport, Active ; drug effects ; Calcium ; metabolism ; Cells, Cultured ; Fura-2 ; pharmacology ; Glutamates ; pharmacology ; Hippocampus ; cytology ; metabolism ; Magnesium ; pharmacology ; Neurons ; cytology ; metabolism ; Rats ; Rats, Sprague-Dawley

Mutations in the parkin gene have recently been identified in familial and isolated patients with early-onset Parkinson disease (PD) and that subregions between exon 2 and 4 of the parkin gene are hot spots of deletive mutations. To study the distribution of deletions in the parkin gene among variant subset patients with PD in China, and to explore the role of parkin gene in the pathogenesis of PD, 63 patients were divided into early onset and later onset groups. Exons 1-12 were amplified by PCR, templated by the genomic DNA of patients, and then the deletion distribution detected by agarose electrophoresis. Four patients were found to be carrier of exon deletions in 63 patients with PD. The location of the deletion was on exon 2 (1 case), exon 3 (2 cases) and exon 4 (1 case). All patients were belong to the group of early onset PD. The results showed that parkin gene deletion on exon 2, exon 3 and exon 4 found in Chinese population contributes partly to early onset PD.
Adult ; Aged ; Exons ; genetics ; Female ; Gene Deletion ; Humans ; Male ; Middle Aged ; Parkinson Disease ; genetics ; Point Mutation ; Ubiquitin-Protein Ligases ; genetics

Mutations in the parkin gene have recently been identified in familial and isolated patients with early-onset Parkinson disease (PD) and that subregions between exon 2 and 4 of the parkin gene are hot spots of deletive mutations. To study the distribution of deletions in the parkin gene among variant subset patients with PD in China, and to explore the role of parkin gene in the pathogenesis of PD, 63 patients were divided into early onset and later onset groups. Exons 1-12 were amplified by PCR, templated by the genomic DNA of patients, and then the deletion distribution detected by agarose electrophoresis. Four patients were found to be carrier of exon deletions in 63 patients with PD. The location of the deletion was on exon 2 (1 case), exon 3 (2 cases) and exon 4 (1 case). All patients were belong to the group of early onset PD. The results showed that parkin gene deletion on exon 2, exon 3 and exon 4 found in Chinese population contributes partly to early onset PD.
Exons/*genetics ; *Gene Deletion ; Parkinson Disease/*genetics ; Point Mutation ; Ubiquitin-Protein Ligases/*genetics

To study the effect of glutamate on the intracellular calcium signal of pure cultured rat astrocytes and the role of NMDA and AMPA receptors in the procedure, the change of calcium signal was investigated by monitoring the fluctuation of intracellular Ca2+ concentration ([Ca2+]i) on the basis of Fura-2 single cell fluorescent ratio (F345/F380). The changes in the effect of glutamate on the intracellular calcium signal were observed after blockage of NMDA and (or) AMPA receptors. It was found that L-glutamate could induce an increased [Ca2+]i in most of the cells in concentration- and time-dependent manner. D-(-)-2-amino-5-phosphonopentanoic acid (D-AP-5, a selective antagonist of the NMDA receptor) and 6-cyano-7-nitroquinoxaline-2, 3-dione (CNQX, a selective antagonist of the AMPA receptor) could abolish the effects of NMDA and AMPA respectively. The treatment of D-AP-5 and CNQX simultaneously or respectively could attenuate the effect of L-glutamate at varying degrees. All these indicated that glutamate could modulate intracellular Ca2+ of pure cultured rat astrocytes through different pathways. The activation of NMDA and AMPA receptors took part in the complex mechanisms.
Animals ; Animals, Newborn ; Astrocytes ; cytology ; metabolism ; Biological Transport ; Calcium ; metabolism ; Calcium Channel Blockers ; pharmacology ; Cells, Cultured ; Cytosol ; metabolism ; Glutamic Acid ; pharmacology ; Hippocampus ; cytology ; metabolism ; Rats ; Receptors, AMPA ; metabolism ; Receptors, N-Methyl-D-Aspartate ; metabolism ; Synaptic Transmission

OBJECTIVETo investigate the distribution of possible novel mutation of parkin gene in variant subset patients with Parkinson's disease (PD) in China and to explore the role of parkin gene in the pathogenesis of PD.

METHODSSeventy patients were divided into early onset and late-onset groups, and 70 healthy subjects were included as controls. Genomic DNA from 70 normal controls and from those of PD patients were extracted from peripheral blood leukocytes with standard procedures. Mutations of parkin gene (exons 1-12) in all subjects mentioned above were screened by PCR-SSCP. And in the samples with abnormal SSCP result, further sequencing was performed to confirm the mutation and its location.

RESULTSA new missense mutation (Gly284Arg) on exon 7 was found in a sample, while 4 samples from all subjects exhibited abnormal mobility shift on SSCP electrophoresis. All mentioned DNA variants were sourced from the samples of the patients with early-onset PD.

CONCLUSIONPoint mutation in parkin gene also contributes partly to the development of early-onset Parkinson's disease in Chinese population.

Adult ; Age of Onset ; Aged ; Base Sequence ; China ; DNA ; chemistry ; genetics ; DNA Mutational Analysis ; Female ; Humans ; Ligases ; genetics ; Male ; Middle Aged ; Mutation, Missense ; Parkinson Disease ; genetics ; Point Mutation ; Polymerase Chain Reaction ; Polymorphism, Single-Stranded Conformational ; Ubiquitin-Protein Ligases

Objective To evaluate the effect of mild hypothermia on amino acids and free radicals in rats during cerebral ischemia and reperfusion. Methods Sixty-three Wistar rats were used in this experiment. Zea-Longa' s method was employed to establish the middle cerebral artery occlusion model in 56 rats, which were then randomized into a normal temperature group (n=28) and a mild hypothermia group (n=28). The other 7 rats underwent sham operation and served as control. The rats in the former two groups were observed at 4 time points (3 hours of cerebral ischemia, 1, 2 and 3 hours reperfusion after 3 hours of ischemia) with regard to the dynamic changes of various amino acids and free radicals in the cortex of the rats. Results Compared with the sham operation group, the concentration of SOD and GSH-PX in the ischemic cortex of normal temperature group and mild hypothermia group decreased significantly (P