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We designed and fabricated a novel high throughput brain-on-chip with three dimensional structure with the aim to simulate the in vivo three-dimensional growth environment for brain tissues. The chip consists of a porous filter and 3D brain cell particles, and is loaded into a conventional 96-well plate for use. The filter and the particle molds were fabricated by using computer modeling, 3D printing of positive mold and agarose-PDMS double reversal mold. The 3D cell particles were made by pouring and solidifying a suspension of mouse embryonic brain cells with sodium alginate into a cell particle mold, and then cutting the resulting hydrogel into pieces. The loaded brain-on-chip was used to determine the neurotoxicity of pesticides. The cell particles were exposed to 0, 10, 30, 50, 100 and 200 µmol/L of chlorpyrifos or imidacloprid, separated conveniently from the medium by removing the porous filter after cultivation. Subsequently, cell proliferation, acetylcholinesterase activity and lactate dehydrogenase release were determined for toxicity evaluation. The embryonic brain cells were able to grow and proliferate normally in the hydrogel particles loaded into the filter in a 96-well plate. Pesticide neurotoxicity test showed that both chlorpyrifos and imidacloprid presented dose-dependent inhibition on cell growth and proliferation. Moreover, the pesticides showed inhibition on acetylcholinesterase activity and increase release of lactate dehydrogenase. However, the effect of imidacloprid was significantly weaker than that of chlorpyrifos. In conclusion, a novel brain-on-chip was developed in this study, which can be used to efficiently assess the drug neurotoxicity, pharmacodynamics, and disease mechanism by combining with a microtiterplate reader.
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Animals , Mice , Brain , Chlorpyrifos/toxicity , Culture Media , Oligonucleotide Array Sequence Analysis , Pesticides/toxicityABSTRACT
Objective To explore the effects of survivin on radiation injury and its relation to caspase 9/6,3 activity. Methods ① IEC 6 cells were treated under hypoxic environment for 8 h and followed by reoxygenation for 24 and 48 h, and then the expression of survivin was identified by Western blotting. ② After treatment with 35 Gy 60 Co , the apoptosis rate in different groups was detected with FACS and the caspase 9/6,3 activity was tested using the caspase 9/6,3 assay kit. Results Hypoxic preconditioning could induce the expression of survivin in IEC 6 cells and survivin could decrease the caspase 3 activity, resulting in reduced apoptosis rate induced by radiation. Conclusion Survivin induced by hypoxic preconditioning can inhibit the apoptosis induced by radiation by reducing the caspase 3 activity.
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Objective To observe the time course of the changes in nitric oxide (NO) content and the activity of NO synthase (NOS) during hippocampal long term potentiation (LTP) in vitro . Methods The production and maintenance of LTP were detected by using extracelluar electrophysiological recording. NO content and NOS activity were determined by biochemical reaction. The expression of NOS mRNA was detected by in situ hybridization. Results Conditioning stimulation for 10 min induced LTP production and significant increases in NO content, NOS activity and expression of NOS mRNA. However, at 60 min after conditioning stimulation, LTP remained stably but NO content and NOS activity returned to the pre conditioning stimulation level. Moreover, the NOS mRNA was overexpressed at the early stage of LTP production. Conclusion Significant NO changes may occur at the early stage of LTP formation.
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<p><b>OBJECTIVE</b>To observe the change of the gene expression of N-methyl-D-aspartate receptors (NMDAR) subunits in rat hippocampus after scalding.</p><p><b>METHODS</b>The backs of the rats were shaved and immersed in warm water for 10 sec. to make control group (C), and the backs shaved and immersed in hot water (90 degrees C) for 10 sec to make 30% full skin scalding model as the scalding group (S). The mRNA expression of the subunits of rat hippocampus NMDAR-NMDAR1, NMDAR2A, NMDAR2B, NMDAR2D was determined with RT-PCR technique in C group and at 0.5, 1, 2 and 4 postburn hours (PBHs) in S group.</p><p><b>RESULTS</b>(1) There exhibited no obvious change of the mRNA expression of all the subunits of NMDAR at 0.5 and 1 PBH in S group when compared with that in C group. But the mRNA expression of NMDAR1 increased for 24.3% and 20.9% and that of NMDAR2A increased for 27.8% and 27.6% at 2 and 4 PBHs respectively when compared with that in C group (P < 0.05). In addition, the mRNA expression of NMDAR2B and NMDAR2D revealed no change after scalding.</p><p><b>CONCLUSION</b>The receptor channel constructed by NMDAR1/NMDAR2A demonstrated increased mRNA expression at 2 PBH, which might lead to the further opening of NMDAR after scalding which might participate in the maladjustment of HPA axis during scalding stress and the following pathophysiological changes.</p>
Subject(s)
Animals , Female , Male , Rats , Burns , Metabolism , Pathology , Disease Models, Animal , Gene Expression , Hippocampus , Metabolism , Pathology , Rats, Wistar , Receptors, N-Methyl-D-Aspartate , GeneticsABSTRACT
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 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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Objective To observe the effects of NMDA receptors and nitric oxide (NO) on neural cell adhesion molecule(NCAM) synthesis during hippocampal long term potentiation (LTP) induction in vitro . Methods LTP induction and maintenance were tested by using extracelluar electrophysiological recording. The synthesis of NCAM protein was detected by Western blotting. Results Application of conditioned stimulation induced LTP and a rise in NCAM at 10 min. NCAM protein level continued to rise while LTP remained stably at 60 min. The NMDA receptor inhibitor AP 5 and the NO synthase inhibitor N nitro arginine inhibited the LTP induction and the increase in the NCAM synthesis. Conclusion The changes in NCAM synthesis during hippocampal LTP induction in vitro may be involved in NMDA receptors and NO.
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Objective To investigate the changes of glucocorticoid receptor (GR) gene expression in the hippocampus of rats following scald burn stress and the role of N methyl D aspartate (NMDA) receptor in this change. Methods Adult male Wistar rats inflicted with 30% TBSA full thickness scalding were applied as severe scalding stress model. GR mRNA levels in the hippocampus were detected with RT PCR. Results A significant decrease of GR mRNA levels was observed in the hippocampus 2 h after the scalding stress. The decrease could be inhibited when MK 801, an NMDA receptor antagonist, was administered prior to stress, and be augmented with the administration of NMDA, an NMDA receptor agonist, but not be affected by normal saline. Conclusion NMDA receptors are involved in the scalding stress induced down regulation of GR gene expression in the rat hippocampus.
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Objective To explore the behavioral and electrophysiological characteristics of prefrontal executive control in macaca mulatas. Methods Three macaca mulatas were used for behavioral research. Neuronal activities of the prefrontal cortex were recorded in two monkeys while visual discriminated go/no go tasks were being performed. Results ① The response time was (419?18)ms, (376?26)ms and (540?21)ms in three monkeys while performing go/no go tasks; ② A total of 41 tetrodes penetrations were made in the PFC of two monkeys. A total of 92 task related neurons were sampled and categorized as 5 types: visual signal related neurons, decision making related neurons, go movement related neurons, reward related neurons and multi events related neurons; ③ When it was correct, the discharge rate of a neuron to a go signal was significantly higher than that to a no go signal during cue period; the discharge rate of a decision making neuron in a go task was significantly higher than that in a no go task. However, when it was wrong, the situation was on the contrary during delay period. Conclusions ① The prefrontal cortex contains various kinds of neurons; ② There are two types of errors in executing go/no go tasks: cue period error and delay period error.
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Objective To investigate the changes of HPA axis activity following scald stress and to elucidate if NMDA receptor is involved in this change. Methods Adult male Wistar rats were inflicted with 30% TBSA full thickness scalding burn, which was applied as severe trauma stress model. Using this model, we detected the changes of serum cortisol and ACTH concentration in scald rats pretreated with intrahippocampal microinjection of NMDA receptor antagonist MK801 or NMDA receptor 1 antisense oligodeoxynucleotide. Results Intrahippocampal microinjection of MK 801 6 ?g resulted in an significant decrease of serum cortisol and ACTH concentration at 2 h after burn, and microinjection of MK 801 12 ?g resulted in a more significant decrease of these values. In accordance with microinjection of MK 801, intrahippocampal microinjection of NMDA receptor 1 antisense oligodeoxynucleotide of 10 nmol/L and 20 nmol/L also resulted in a significant and a more significant decrease of serum cortisol and ACTH concentration after burn. Conclusion Hippocampal NMDA receptor plays an important role in over excitation of HPA axis following burn.
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Objective To study the effect of chronic low dose soman on learning and memory and long term potentiation(LTP) of hippocampal slices. Methods Rat model was established by consecutive subcutaneous injection of soman(6-10 ?g?kg 1 , s. c, sig?14) for 14 days for Morris water maze test. Long term potential of synaptic transmission was observed in CA1 region by tetanization of the Schaffer commissural pathway in rat hippocampal slices. Results In the Morris water maze, latency to find a hidden platform was longer and the times of crossing the situation of platform and the time percent of swimming in northeast obviously decreased. In the experiment on hippocampal slice of rats in vitro by microelectrode method, the generation of long term potentiation was inhibited. Conclusion Chronic low dose soman may cause an evident learning and memory disturbance and decrease hippocampal synaptic plasticity.
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Objective To investigate the characteristics of spontaneous discharges of hippocampal pyramidal cells (PC) in rats in age and memory groups with the help of nonlinear theory. Methods Rats were randomly divided into age group and memory group. Age group included aged group (16-19 months) and young group (3-4 months) whereas memory group included good memory and poor memory groups in adult. Extracellular single cell recording was performed in vivo . Results No characteristics of the rhythm of spontaneous discharges of hippocampal pyramidal cells in distribution figure of interspike interval(ISI) were found, but the loss of complexity(C) and low percentage of favored patterns(PF) were found in age and poor memory groups. Conclusion The ISI complexity and the PF of the hippocampal pyramidal cells are correlated to age and memory, suggesting that the analysis of the complexity and favored patterns may be helpful for the discovery of the characteristics of the information coding in spontaneous discharges of hippocampal pyramidal cells.
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Objective To design the automatic control system for visual discrimination Go/No Go task training applied in the research of cognitive neuroscience, neurophysiology, neuropharmacology, psychology and psychiatry. Methods Windows was used as the operation platform. The powerful graphic visualized Visual C++6.0 was used as the software for the development. Electric magnet compatible technology was applied in the hardware for the purpose of the stable system. Results This system could automatically and accurately control the whole process of visual discrimination Go/No Go task training. Conclusion The system with friendly interface is stable, reliable, easy to operate, and accurate to judge and time the animal behaviors by electric magnet compatible transducer.
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0.05). Spike of [Ca 2+ ]i of astrocytes was induced by acute morphine administration, but the spike of [Ca 2+ ]i of astrocytes pretreated with morphine could be induced only by higher concentration of morphine. The [Ca 2+ ]i response of astrocytes to morphine could be blocked by naloxone but not by MK 801(NMDA receptor antagonist). Conclusion The response of [Ca 2+ ]i in morphine induced astrocytes may play an important role in morphine tolerance.
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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.\;[