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BACKGROUND: After neural precursor cells (NPCs) induced from embryonic stem cells (ESCs) have been grafted into the brain, it would still keep some potency of proliferation and differentiation, strong plasticity and integration into the host neural tissues, which would help to observe the therapeutic effect of PD.OBJECTIVE: To observe the differentiation of mouse ESCs into NPCs and the therapeutic effect of NPCs after being transplanted on the behavior of Parkinson disease (PD) rats.DESIGN: Randomly and controlled animal experiment.SETTING: Staff Room of physiology and Staff Room of Neurobiology, Depayment of Basic Medical Sciences, the Third Military Medical University of Chinese PLA.MATERIALS: Totally 50 healthy adult Wistar rats were chosen and randomly divided into experimental group (n=45) and control group (n=5).METHODS: ① 5 μ L (2 g/L)6-hydroxydopamine (6-OHDA) was injected into substantia nigra pars compacta and ventral tegmental area two points in the experimental group to prepare PD rats, and normal saline with the dosage of 5 μL per point was injected into the rats in the control group.Behavioral test began at 1 week after operation to measure successful rate of model establishing, once a week for 7 consecutive weeks. ② Totally 20 successful PD rat models were chosen to perform corpus striatum NPCs with the dosage of 2 μL [the count of cell suspension was (5-8)×106/μL].The other 5 rats were given 2 μL normal saline at corpus striatum as normal saline control group.MAIN OUTCOME MEASURES: ① Successful rate of PD model. ②Effect of NPCs transplantation on the rotation times of PD models. ③ Distribution of transplanted NPCs in vivo, and survival and differentiation.RESULTS: ①6 weeks later, totally 33 of 45 rats in the experimental group achieve the standard of PD model . ② About 85% of mouse ESCs were differentiated into Nestin-positive NPCs 5 days after the embryoid bodies formed in the bacterial dishes and cultured in the N2 serum-free medium. ③The rotation times of the PD rats was significantly decreased after the intracerebral transplantation of NPCs as compared with normal control group. Most of the NPCs grafted into striatum of PD rats were survived, and some were differentiated into TH-positive neurons.CONCLUSION: The mouse ESCs-derived NPCs could be transplanted into striatum of PD rats, and then differentiated into TH-positive neurons,which leads to the obvious decrease of rotation times.
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Objective To clarify the effective location of propofol in central nervous system (CNS). Methods Forty-two Wistar rats were ramdomized into control group,50 mg/kg propofol,100 mg/kg propofol,150 mg/kg propofol,tail shearing,propofol followed by tail shearing (n=7 in each group). The NOS expressions in the CNS were recorded by NADPH-d histochemistry after anesthesia by intraperitoneal injection of propofol. Results Rather widely stained NOS positive neurons were observed in the control group. In propofol groups,the NOS expressions were decreased significantly as compared with the control group,mainly located in ACB,LS,Pe,VLG,Den,SO,SCh,AVPO,Sol,SuM,BL,PV,LHb and Icj,showing a negative dose-effect relation with propofol. Conclusion Propofol has the determined sites of action in CNS and the decrease of NO synthesis by the inhibition of NOS may play a role in propofol-induced general anesthesia.
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Objective To clarify the effective location of propofol in central nervous system (CNS) by detection of the c-jun expression after propofol-induced anesthesia in rats. Methods Wistar rats were randomly divided into 6 groups: normal control (C), low-dose propofol group (50 mg/kg, P 1), middle-dose propofol group (100 mg/kg, P 2), high-dose propofol group (150 mg/kg, P 3), stimulation with tail broken group (S 1), and propofol + stimulation with tail broken group (S 2). The expressions of nucleoprotein JUN in the CNS were detected by immunohistochemisty. Results Rather weakly stained nucleoprotein JUN positive neurons were observed in the supraoptic nucleus, lateral septal nucleus, and lateral habenular nucleus in the control group. In groups P 1, P 2, and P 3, the expressions of nucleoprotein JUN were increased significantly as compared with those in the control group. The expressions were mainly located in the accumbent nucleus, lateral septal nucleus, periventricular hypothalamic nucleus, ventral lateral geniculalaten nucleus, dorsal lateral geniculate nucleus, supraoptic nucleus, suprachiasmatic nucleus, anteroventral preoptic nucleus, nucleus of the solitary tract, supramammillary nucleus, basolateral amygdaloid nucleus, paraventricular thalamic nucleus, lateral habenula nucleus, and islands of Calleja. The expressed positive neuron number was positively correlated with the doses of propofol. Conclusion Propofol anesthesia has the determined sites of action in rat CNS.
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Objective To evaluate the therapeutical effect of dopaminergic neurons induced by transplantation on Parkinson's disease (PD) rats. Methods Mesencephalic nerve stem cells (NSCs) were induced by striatal extracts to differentiate into tyroxine hydroxylase (TH) positive dopaminergic neurons. The differentiated cells were transplanted into the striatum of PD rats. The survived cells were detected by TH immunocytochemical staining. The therapeutical effect was observed using apomorphine induced rotation. Results Mesencephalic NSCs could be induced to differentiate into dopaminergic neurons which could survive in the host for long time after cell transplantation, and could improve the apomorphine induced rotation. Conclusion The induced mesencephalic NSCs have the obvious therapeutical effect on PD.
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Objective To observe the expression changes of noggin mRNA and BMP4 mRNA in the hippocampus and frontal cortex of rats at different stages. Methods The expressions of noggin mRNA and BMP4 mRNA were analyzed by the method of reverse transcriptase polymerase chain reaction (RT PCR).Results It was revealed that the level of noggin mRNA in the frontal cortex decreased significantly in P1W rats but high level of BMP4 mRNA was detected in P1M and P3M rats. The expressions of noggin mRNA and BMP4 mRNA in the hippocampus showed the opposite expression pattern. The peak of noggin mRNA expression in the hippocampus was found in E13 and E16 rats. The expression of noggin mRNA decreased gradually but that of BMP4 mRNA in hippocampus increased gradually during the developmental stage. The peak of the expression of BMP4 mRNA was found in P1M rats. Conclusion There are expressions of noggin mRNA and BMP4 mRNA in the frontal cortex and hippocampus in rats at different developmental stages. The expression level is closely correlated with the developmental age. This indicates that noggin and BMP4 play important roles in the development of rat frontal cortex and hippocampus.
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Objective To investigate the effect of noggin on BrdU labeled cells in the adult rat hippocampus. Methods The expressions of noggin and bone morphogenetic protein 4 (BMP4) in rat hippocampus were detected using in situ hybridization histochemistry (ISHH) and reverse transcription polymerase chain reaction (RT PCR). By using antisense technique combined with bromodeoxyuridine!(BrdU) labeling, the effect of noggin on hippocampal neurogenesis in adult rats was explored. Results The number of noggin mRNA positive cells in the adult rat hippocampus decreased significantly after treatment with antisense noggin but no change was found in the number of BMP4 mRNA positive cells. In addition, the number of BrdU labeled cells decreased significantly in the adult rat hippocampus after treatment with antisense noggin, but the sense noggin had no such effect. Conclusion Noggin can promote proliferation of neural precursor cells in adult rat hippocampus.
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Objective To investigate the effect of estrogen on the pain score,c-Fos and substance P expressions in the dorsal horn of the spinal cord in the mice following formalin stimulation.Methods Fifteen C57/BL6 mice were randomized to 3 groups: control group(intact mice without estrogen treatment),OVX+V group(ovariectomized mice given vehicle) and OVX+E group(ovariectomized mice with subcutaneous injection of 2 ?g/d 17?-estradiol for 10 d).Pain score was used to assay the role of estrogen in affecting pain threshold in the mice following formalin injected into the right hind paw,and expressions of c-Fos and substance P in the dorsal horn of spinal cord(L3 to L5) in 2 h after injection of formalin was tested with immunohistochemisty to evaluate neuron activity and pain afferent fibers.Results Pain score was increased in ovariectomized mice following formalin stimulation,which was inhibited by estrogen especially in the early stage of secondary phase.The number of c-Fos-like immunoreactivity neuron(FLIN,P
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Objective To investigate the expression of neuron-specific enolase(NSE) and bone morphogenic protein 4(BMP4) in different hippocampal areas of pentylenetetrazol(PTZ) kindled epilepsy rats and explore their relationship with the pathogenesis of epilepsy and brain injury.Methods Fifty male SD rats were divided into experimental group(n=40) and control group(n=10).The rats in experimental group were kindled into epilepsy by chemical method,and according to the kindling process,subdivided into four groups(grade Ⅰ,Ⅲ,Ⅳ,Ⅴ).Immunohistochemistry,in situ hybridization labeled with Dig-oligonucleotide probe and the image analyzing system were used to observe the expressions of NSE and BMP4 in rat hippocampus.Results In PTZ kindled epilepsy rats,the number of cells positive for NSE and BMP4 was increased in many regions of hippocampal formation.Compared with control group,the expressions of NSE and BMP4 in CA3 and DG was elevated obviously in the grade Ⅲ group and grade Ⅳ group(P
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Objective To examine the expression of BMP4 in CNS of the developing rat. Methods In situ hybridization histochemistry(ISHH) was carried out on tissue sections using specific digoxigenin\|labeled oligonucleotide probe. Results It showed that BMP4 mRNA positive cells were located mainly in cerebellum and olfactory at E16.Strong positive signal was seen in hypoglossal nucleus,and moderate signal also seen in spinocerebellar tract and spinal lemniscus at P1\|2.The number of BMP4 mRNA positive cells was increased in the frontal cortex,parietal cortex,and hippocampus subiculum at P1W.The peak of BMP4 expression was in cortex and periamydaloid cortex.Widely distributed BMP4 mRNA positive cells were detected in cortex and hippocampus of rats at P1M,strong positive signal was observed in temporal CNS at P3M,strong positive signal was observed in hippocampus,temporal corex and periamydaloid cortex,lateral nucleus of thalamus and paraventricular nucleus of hypothalamus.BMP4 mRNA positive cells were also found in corex,hippocampus,hypothalamus and thalamus at P18M.Conclusion\ These results indicated that BMP4 could play an important role in CNS development of rats.
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Objective To examine the expression of Noggin in CNS of the developing rat. Methods In situ hybridization histochemistry(ISHH) was performed using digoxigenin-labeled cRNA as probes. Results It was revealed that densely and deeply stained noggin positive cells were detected in cortex,hippocampus,cerebellum,and nucleus of hypothalamus and thalamus in embryonic day(E)16 rats.The number of noggin positive cells was increased in the thalamus and medulla oblongata at postnatal day(P)1-2,whereas decreased in the hippocampus and cortex.The number of noggin positive cells was decreased significantly in brain at 1 week postnatal(P1W),and began to increase at P2W,especially in the cortex and hippocamps.Strong positive signal can be detected in the frontal cortex,parietal cortex,cingulated cortex,hippocampus,olfactory and cerebellum at 1 month postnatal(P1M).The expression of noggin begins to decline at P3M,only sparse noggin positive cells can be seen in CNS at P18M.Furthermore,there is no noggin positive cells seen in the spinal cord of rats during development.Conclusion Our results indicated that noggin could play an important role in CNS development of rats.
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AIM: To observe the effect of adrenocorticotropic hormone(ACTH) on the levels of brain-derived neurotrophic factor (BDNF), trkB and corticotropin-releasing hormone(CRH) in the hippocampus of arthritic rats.METHODS: The BDNF immunoreactivity (IR) and CRH-positive neurons were stained with immunohistochemistry and in situ hybridization methods, respectively.RESULTS: The BDNF-IR, CRH mRNA-positive neurons in the contralateral hippocampus of the arthritic rats were increased significantly, which was decreased markedly by intraperitoneal injection of ACTH. However, the effect of ACTH was attenuated after adrenalectomy (ADX).CONCLUSION: These results suggest that BDNF and CRH in the hippocampus of arthritic rats were involved in the modulation of chronic pain, ACTH produced its analgesic effect by inhibiting the increase in BDNF and CRF level. Adrenal is critical to the analgesic action of ACTH.
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AIM: To observe the effect of altitude hypoxia on glutamate, aspartate and nitric oxide synthase (NOS) in the rat hypothalamus. METHODS: Using altitude hypoxia model, amino acid analysis system and the NADPH-d histochemistry, we determined the content of glutamate, aspartate and the number of NADPH-d neurons in the rat hypothalamus. RESULTS: After altitude hypoxia, the contents of glutamate, aspartate in the hypothalamus of rats were increased significantly, densely and deeply stained NADPH-d neurons were seen in hypothalamic paraventricular nucleus (PVN)and supraoptic nucleus(SON). If rat were pretreated with the NMDA receptor blockers Ketamine (ip,40 mg/Kg)or AP-V(i.c.v, 10 ?g) , the number of NADPH-d neurons in the rat hypothalamic PVN and SON was markedly less than that in corresponding altitude hypoxia group. CONCLUSION: NMDA receptor may take part in the expression of hypothalamic NOS induced by altitude hypoxia.
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AIM: To observe the survive, differentiation and therapeutic effect of neural precursor cells (NPCs) differentiated from mouse embryonic stem cells (ESc) when transplanted in the frontal cortex of Alzheimer's disease (AD) rats. METHODS: NPCs were induced from mouse ESc with serum-free methods. The differentiation of transplanted NPCs was observed with immunohistochemistry methods and memory of rats was evaluated with Morris water maze test. RESULTS: About 85% of mouse ESc were differentiated into NPCs 5 days after the embryoid bodies cultured in the N2 medium. 4 and 6 weeks after transplantation, the memory impairment of AD rats was relieved, most of the grafted NPCs were kept undifferentiated and proliferated in clone shape, neuron-like long processes was observed. CONCLUSIONS: The NPCs derived from ESc survive and differentiate into neurons after grafted into the frontal cortex of AD rats, which produces therapeutic effects on AD.
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AIM:To observe the effect of altitude hypoxia on ?-aminobutyric (GABA) content and prepro-somatostatin mRNA (PPS-mRNA) in the rat hypothalamus. METHODS: Using altitude hypoxia model,in situ hybridization and amino acid analyzer, the number of PPS-mRNA and GABA content in rat hypothalamus was determined. RESULTS:After altitude hypoxia, the contents of GABA in hypothalamus and the number of PPS-mRNA neurons in periventricular nucleus (PeVN), paraventricular nucleus (PaVN) and arcuate nucleus (ArcN) increased significiantly. Bicuculline, a GABA receptor antagonits, could enhance PPS-mRNA expression evoked by altitude hypoxia, but had no effect on GABA content. CONCLUSION: Altitude hypoxia can induce neurotransmitter imbalance of hypothalamus.
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Objective To exam the expression and the distribution difference between melatonin membrane receptor subtype Mel 1a and Mel 1b in the central nervous system of rats. Methods In situ hybridization technique was used. Results (1)The Mel 1a mRNA positive cells were mainly detected in the hippocampus,cerebral cortex,supraoptic nucleus,paraventricular nucleus,suprachiasmatic nucleus,inferior olivary nucleus,cortex and fastigial nucleus of cerebellar,ventral horn of the spinal cord,facial nerve nucleus,gigantocellular reticular nucleus,striatum cortex and trigeminal nerve nucleus,etc.(2)The Mel 1b mRNA positive cells were mainly observed in the cerebellar cortex,fastigial nucleus,global nucleus,emboliform nucleus of the medullaris cerebelli,hippocampus,cerebral cortex,ventral horn of the spinal cord,supraoptic nucleus and suprachiasmatic nucleus.Conclusion\ Mel 1a mRNA positive neurons were abundant and distributed widely in the CNS,while Mel 1b mRNA\|positive neurons distributed comparatively localized.However,the hippocampus and the cortex were two regions which were rich in both Mel 1a and Mel 1b mRNA positive neurons.\;[