CREB信号途径介导OPs致认知功能障碍的作用机制
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摘要
目前农药已经成为人们提高农作物产量和日常生活杀虫的必需品,有机磷杀虫剂(Organophosphorus pesticides,OPs)因其经济、高效的特点,逐渐成为目前应用最多的杀虫剂。OPs在全球范围内的大量使用,势必对环境造成持续的污染,这种长期、低剂量的OPs暴露对人类健康的威胁值得关注。
流行病学调查显示OPs暴露人群出现学习记忆等认知功能障碍,且学习记忆能力的下降出现在OPs暴露后的一段时间。而目前关于OPs长期低剂量染毒对动物认知功能影响的试验研究并不多见,其研究结果并不一致,且大多数体内研究水平还停留在对现象的描述阶段。因此,长期、低剂量OPs暴露对中枢神经系统认知功能的作用机制尚需进一步研究。
毒死蜱(Chlorpyrifos,CPF)是一种广谱、高效、低毒的OPs,由于其严重的发育毒性,美国已经在2002年禁止CPF在家庭杀虫中应用,但目前CPF仍然大量应用于全世界的农作物杀虫,是全球使用量最多的OPs之一。CPF是我国2007年以后推荐的对硫磷、甲胺磷等禁用高毒OPs的替代品之一,也是我国农副产品中经常检测出的主要残留农药和超标农药,因此研究CPF长期低剂量暴露对认知功能的影响具有重要意义。
由于海马体在学习记忆,尤其是空间学习记忆功能中的重要作用,海马体已经成为研究空间学习记忆这一高级认知功能的重要手段。本研究以CPF为受试物,选择不引起动物出现有机磷全身中毒症状的低剂量(1、5、10mg/kg),对雄性Wistar大鼠施以连续12周的灌胃染毒,围绕海马在空间学习记忆中的作用,利用Morris水迷宫、自发行为和抓力测试检测CPF低剂量重复染毒对动物中枢神经系统认知功能的影响,尤其是空间学习记忆功能的影响,采用免疫组化、免疫荧光和蛋白印记等方法检测相关信号通路的变化,探讨CPF低剂量重复染毒致认知功能障碍的可能作用机制。
经过12周CPF连续染毒,动物的体重、摄食量没有明显改变,试验过程中也没有观察到流涎、肌肉振颤、腹泻等明显的有机磷中毒症状,而Morris水迷宫测试结果显示染毒动物到达平台的时间(潜伏期)明显延长,而游泳速度也出现降低趋势;为探讨潜伏期的延长原因,进一步对动物进行连续3天的可视平台训练,发现染毒动物的潜伏期和游泳速度与对照组相比没有明显改变,训练后立即利用Morris水迷宫再次进行测试,发现动物的潜伏期和游泳速度均没有明显改变,上述结果表明CPF低剂量重复染毒能够诱发大鼠长时程空间记忆障碍,而对动物的学习和短时程空间记忆能力没有影响。而自发行为和抓力测试结果进一步表明CPF染毒后大鼠运动能力没有明显的影响,而动物海马的AChE活性检测结果显示10mg/kg剂量组出现轻度的AChE抑制(30%),而1、5mg/kg剂量组没有明显改变,暗示CPF致长时程空间记忆障碍存在着非胆碱能因素。
海马组织病理切片检查发现CPF染毒动物海马的基本结构没有明显改变,但是细胞计量分析却表明海马区神经细胞出现减少。为进一步分析海马区神经细胞的变化,利用特异的标志物对海马区几种主要的细胞进行检测。NeuN是只在成熟神经元的胞核中表达的转录因子,western blot结果显示染毒动物海马的NeuN蛋白表达产物不仅没有减少,反而增加,而免疫组化结果则进一步提示海马齿状回(DG)区NeuN阳性细胞出现增加趋势。β-3 tubulin(TUJ1)和NF68是新生神经元特异标志物,western blot和免疫组化均证明CPF染毒动物海马的β-3 tubulin、NF68表达明显降低,而BrdU(神经前体细胞)、GFAP(星形胶质细胞)的检测也同样出现剂量依赖式减少。上述结果提示:CPF低剂量重复染毒能够引起海马区细胞构成的改变,主要表现为新生神经元减少,而成熟的神经元却没有明显变化。可见,海马区神经细胞的减少并非是CPF的直接毒性作用,可能跟信号途径的调控有关。而TUNEL免疫组化的凋亡细胞检测结果并未发现染毒动物海马凋亡细胞明显增多,凋亡相关蛋白Bax/Bcl-2、p53以及caspase-3的检测结果也未见明显改变,说明凋亡并非细胞减少的主要原因,上述结果表明:CPF重复染毒影响了海马的神经发生(neurogenesis)。大量文献确证成年动物的海马齿状回(DG区)终身存在着神经发生,而这些新生神经细胞在海马相关的空间记忆,尤其是长时程记忆中起着非常重要的作用。可见,CPF低剂量重复染毒后动物海马新生神经细胞的大量减少可能是其出现长时程空间记忆障碍的重要原因。
成年动物神经发生的调控是目前神经生物学研究的热点,最新研究结果表明cAMP反应单元结合蛋白(CREB)在成年动物的记忆存储分配(Memory Allocation)中具有非常重要的作用,磷酸化的CREB可以活化其下游基因,促使其转录,从而诱导海马DG区神经前体细胞的神经分化和促进新生神经元的存活与增殖,CREB活性的降低则导致新生神经元的减少。而CREB信号通路的激活也是长时程学习记忆能够发生的关键,在学习记忆发生过程中,CREB的磷酸化蛋白表达增加。为探讨CPF低剂量重复染毒后致长时程空间记忆障碍的机制,对海马CREB的磷酸化进行检测,发现染毒组动物海马CREB的磷酸化蛋白表达明显降低,而总CREB却没有明显改变。
CREB信号通路的激活主要受钙/钙调素依赖性蛋白激酶Ⅱ(CaMKⅡ)、蛋白质激酶A(PKA)、细胞外调节蛋白激酶(ERK)等激酶的调控。CaMKⅡ在空间学习与记忆和突触可塑性中发挥着重要的作用,被认为是记忆的分子开关;腺苷酸环化酶(cAMP)依赖性蛋白激酶途径是研究最早的学习记忆相关通路,活化的PKA可直接激活下游CREB而调控学习记忆功能。ERK信号通路在中枢神经系统中具有调控细胞增殖分化和学习记忆等多种功能,与脑内长时程增强(LTP)的形成以及空间学习记忆功能有重要的联系。本研究中,检测CREB上游的主要调控激酶发现ERK的磷酸化蛋白表达也显著降低,而PKA、p-CaMKⅡ蛋白表达则没有明显改变。由于ERK和CREB均在空间记忆和长时程记忆中起重要作用,表明ERK-CREB信号途径可能是CPF重复染毒致长时程空间记忆障碍的重要调控通路。
综上所述,CPF低剂量重复染毒能够抑制ERK-CREB信号途径的磷酸化,从而影响海马DG区神经发生,导致海马新生神经细胞减少,最终诱发长时程空间记忆障碍。
Organophosphorus insecticides (OPs) are the most widely used group of pesticides in the world. Because of their relatively low cost, OPs have been used in agriculture, homes, gardens, and in veterinary practice. The use of OPs around the world continues to produce detectable levels of OPs in the environment, thereby posing an ongoing risk for low-level exposure. Chronic exposures to OPs, especially the low-level envireonmental exposures, require greater attention.
The epidemicology studies had reported memory impairments and behavioral alterations after long-term exposures to OPs, and the decrease cognitive function appeared not immediately after the OPs exposure but monthes later. However, there are a few animals studies available that focused on the mechanisam of cognitive impairments on chronic low-level OPs exposure. Some studies in which animals were used as experimental subjects had contradictory results on the cognitive and behavioral effects of OPs exposure. Therefor, the results and mechanisam of chronic low-level OPs exposure need further study.
Chlorpyrifos (O,O’-dithyl-O-3,5,6-trichloro-2-pyrydyl phosphorothionate, CPF) is one of the most widely used OPs throughout the world. Despite the EPA (U.S. Environmental Protection Agency.2002) has entered into an agreement with the registrant to begin terminating residential CPF indoor uses since 2000, it is still one of the most widely used insecticides. With the elimination of acephatemet, methyl parathion, parathion, monocrotophos and phosphamidon in agriculture since 2007, CPF had been recommend as the low-toxic OPs replacement in agriculture in China.These days, CPF has become one of the major pesticides detected in farm products in China.
In the present study, male rats were administrated CPF through intragastric for 12 weeks to evaluate the neurobehavioral impacts of CPF at the subthreshold doses(doses that do not produce overt signs of cholinergic toxicity, e.g. excessive salivation, urination, defecation, muscle fasiculations and so on). The neurobehavioral changes (especially the spatial memory) were studied and the and mechanisam of chronic low-level OPs exposure was explored.
There were no effects on weight gain and food consumption detected during the 12 weeks exposures to CPF, and the daily dosages of 1.0, 5.0 and 10.0 mg/kg CPF for 12 weeks did not elicit any other signs of OPs toxicity, such as excessive salivation, urination, defecation, muscle fasiculations etc. However, the animals exhibited obvious neurobehavioral changes (especially the long-term spatial memory) after 12 weeks repeated exposures to CPF. The increased latency in the Morris water maze during the treated animals indicated that the chronic low-level CPF exposures can impair the long-term spatial memory. As there was no difference between the CPF treated groups and the vehicle treated group on the results from grip strength effects of CPF, and there were no effects on short-term spatial memory or learning. What’s more, only the highest dosage (10mg/kg) had significant inhibition effects on AChE activity, the exhibition of impaired long-term spatial memory may exist non- cholinergic reasons.
The hippocampus has become the most important method to study the higher cognitive function, since its special function in the spatial memory. In the present study, we found significant decrease of cell numbers in the regions of hippocampus, although the hippocampal regions from the CPF treated groups did not exhibit any obvious architectural difference from the vehicle control group on gross observation. However, the immunohistochemisty study exhibited that NeuN (mature neuron marker) positive cells in the dentate gyrus(DG)increased, and the western blot of the NeuN protein from the whole hippocampi also increased significantly. The results from immunohistochemisty and western blot study showed thatβ-3 tubulin(TUJ1)and NF68 (adult new born neuron marker) decreased significantly and the marker of neuron precursor cells, BrdU, also exhibited significant decrease in a dose-dependent manner. The phenomenon discussed above illustrated that the repeated exposures of CPF can affect the constitute of cells in hipocampus, in another words, repeated exposures of CPF lead to the decrease of the adult new born neurons, but have no effect on the mature neurons. The hurt to brain(hypoxia, infection,poisoning,et al.) often exhibited glial cell hyperplasia under normal circumstances, while the GFAP protein expression protein from the whole hippocampi and the GFAP positive cells in the dentate gyrus(DG)decreased simultaneously. The changes that discussed above indicated the decrease of the cell in DG may not the direct toxicity effect from CPF, but the regulation result of some signal pathway. What’s more, the TUNEL positive cells did not show any increase in the treated animals, and apoptosis related regulator , Bax/Bcl-2、p53 and caspase-3 did not show any change after the repeated exposures to CPF. It means the apoptosis could not be the main reason of the cells decrease in the DG. The changes of the cell constitute in the DG indicate the repeated CPF exposures can affect the neurogensis of the hippocampus.
More and more studies had proved that the DG region of the adult hippocampus can have neurogensis all the life, and the adult new born neurons have important function in the hippocampus related spatial memory, especially the long-term memory. The decrease of the new born neurons can result in the impairment of long-term memory. Then the significant decrease of the adult new born neurons in the DG region may be the main reason of the impaired long-term spatial memory in the CPF treated animals.
The regulation of the adult neurogensis is the research focus of neurobiology studies, and the latest results indicated that the cyclic AMP responsive element binding protein (CREB) have an important function during the Memory Allocation. The phosphorylated CREB can activate the dowmstream gene transcription, which can induce the differentiation of the hippocampal precursor cells and promote the survival and proliferation of the adult new born neurons. The decrease of the phosphorylated CREB can lead to deficiency of the adult new born neurons. As CREB is one of the most prominent molecules involved in learning and memory processes. Evidence suggests that CREB may play a crucial role in hippocampal plasticity and consolidation of long-term memory, since CREB activates the transcription of genes required for long-term synaptic changes involved in memory formation. Animals with CREB protein mutations exhibit learning and memory deficits. While, the activation of CREB pathway can be the regulating results of Ca2+/calmodulin-dependent kinaseⅡ(CaMKⅡ)、protein phosphakinase A (PKA)、extracellular regulated protein kinases (ERK). CaMKⅡhas been regarded as the molecular switch during the leaning and memory function. The cAMP related protein kinases pathway was the first memory related signal pathway studied, and the PKA can active CREB directly. ERK, as the upstream kinases of CREB pathway have important function during the neuron differentiation and long-term potentiation(LTP).
In the present study, the expression protein level of phosphorylated CREB and ERK1/2 in hippocampus from the CPF treated rats decreased significantly, but the CPF had no effects on the expression of PKA and phosphorylated CaMKⅡ. The dose-dependent decrease indicated that the inhibition of the ERK-CREB Signaling may be one of the important regulating pathway during the CPF caused spatial long-term memory impairments.
In summary, the present study identified that repeated exposure to subthreshold doses of CPF can lead to the inhibition of ERK-CREB signaling, which regulated the neruogenesis of the hippocampal DG and induced the deficiency of adult new born neurons. The decreased young neurons lead to the impaired long-term spatial memory.
流行病学调查显示OPs暴露人群出现学习记忆等认知功能障碍,且学习记忆能力的下降出现在OPs暴露后的一段时间。而目前关于OPs长期低剂量染毒对动物认知功能影响的试验研究并不多见,其研究结果并不一致,且大多数体内研究水平还停留在对现象的描述阶段。因此,长期、低剂量OPs暴露对中枢神经系统认知功能的作用机制尚需进一步研究。
毒死蜱(Chlorpyrifos,CPF)是一种广谱、高效、低毒的OPs,由于其严重的发育毒性,美国已经在2002年禁止CPF在家庭杀虫中应用,但目前CPF仍然大量应用于全世界的农作物杀虫,是全球使用量最多的OPs之一。CPF是我国2007年以后推荐的对硫磷、甲胺磷等禁用高毒OPs的替代品之一,也是我国农副产品中经常检测出的主要残留农药和超标农药,因此研究CPF长期低剂量暴露对认知功能的影响具有重要意义。
由于海马体在学习记忆,尤其是空间学习记忆功能中的重要作用,海马体已经成为研究空间学习记忆这一高级认知功能的重要手段。本研究以CPF为受试物,选择不引起动物出现有机磷全身中毒症状的低剂量(1、5、10mg/kg),对雄性Wistar大鼠施以连续12周的灌胃染毒,围绕海马在空间学习记忆中的作用,利用Morris水迷宫、自发行为和抓力测试检测CPF低剂量重复染毒对动物中枢神经系统认知功能的影响,尤其是空间学习记忆功能的影响,采用免疫组化、免疫荧光和蛋白印记等方法检测相关信号通路的变化,探讨CPF低剂量重复染毒致认知功能障碍的可能作用机制。
经过12周CPF连续染毒,动物的体重、摄食量没有明显改变,试验过程中也没有观察到流涎、肌肉振颤、腹泻等明显的有机磷中毒症状,而Morris水迷宫测试结果显示染毒动物到达平台的时间(潜伏期)明显延长,而游泳速度也出现降低趋势;为探讨潜伏期的延长原因,进一步对动物进行连续3天的可视平台训练,发现染毒动物的潜伏期和游泳速度与对照组相比没有明显改变,训练后立即利用Morris水迷宫再次进行测试,发现动物的潜伏期和游泳速度均没有明显改变,上述结果表明CPF低剂量重复染毒能够诱发大鼠长时程空间记忆障碍,而对动物的学习和短时程空间记忆能力没有影响。而自发行为和抓力测试结果进一步表明CPF染毒后大鼠运动能力没有明显的影响,而动物海马的AChE活性检测结果显示10mg/kg剂量组出现轻度的AChE抑制(30%),而1、5mg/kg剂量组没有明显改变,暗示CPF致长时程空间记忆障碍存在着非胆碱能因素。
海马组织病理切片检查发现CPF染毒动物海马的基本结构没有明显改变,但是细胞计量分析却表明海马区神经细胞出现减少。为进一步分析海马区神经细胞的变化,利用特异的标志物对海马区几种主要的细胞进行检测。NeuN是只在成熟神经元的胞核中表达的转录因子,western blot结果显示染毒动物海马的NeuN蛋白表达产物不仅没有减少,反而增加,而免疫组化结果则进一步提示海马齿状回(DG)区NeuN阳性细胞出现增加趋势。β-3 tubulin(TUJ1)和NF68是新生神经元特异标志物,western blot和免疫组化均证明CPF染毒动物海马的β-3 tubulin、NF68表达明显降低,而BrdU(神经前体细胞)、GFAP(星形胶质细胞)的检测也同样出现剂量依赖式减少。上述结果提示:CPF低剂量重复染毒能够引起海马区细胞构成的改变,主要表现为新生神经元减少,而成熟的神经元却没有明显变化。可见,海马区神经细胞的减少并非是CPF的直接毒性作用,可能跟信号途径的调控有关。而TUNEL免疫组化的凋亡细胞检测结果并未发现染毒动物海马凋亡细胞明显增多,凋亡相关蛋白Bax/Bcl-2、p53以及caspase-3的检测结果也未见明显改变,说明凋亡并非细胞减少的主要原因,上述结果表明:CPF重复染毒影响了海马的神经发生(neurogenesis)。大量文献确证成年动物的海马齿状回(DG区)终身存在着神经发生,而这些新生神经细胞在海马相关的空间记忆,尤其是长时程记忆中起着非常重要的作用。可见,CPF低剂量重复染毒后动物海马新生神经细胞的大量减少可能是其出现长时程空间记忆障碍的重要原因。
成年动物神经发生的调控是目前神经生物学研究的热点,最新研究结果表明cAMP反应单元结合蛋白(CREB)在成年动物的记忆存储分配(Memory Allocation)中具有非常重要的作用,磷酸化的CREB可以活化其下游基因,促使其转录,从而诱导海马DG区神经前体细胞的神经分化和促进新生神经元的存活与增殖,CREB活性的降低则导致新生神经元的减少。而CREB信号通路的激活也是长时程学习记忆能够发生的关键,在学习记忆发生过程中,CREB的磷酸化蛋白表达增加。为探讨CPF低剂量重复染毒后致长时程空间记忆障碍的机制,对海马CREB的磷酸化进行检测,发现染毒组动物海马CREB的磷酸化蛋白表达明显降低,而总CREB却没有明显改变。
CREB信号通路的激活主要受钙/钙调素依赖性蛋白激酶Ⅱ(CaMKⅡ)、蛋白质激酶A(PKA)、细胞外调节蛋白激酶(ERK)等激酶的调控。CaMKⅡ在空间学习与记忆和突触可塑性中发挥着重要的作用,被认为是记忆的分子开关;腺苷酸环化酶(cAMP)依赖性蛋白激酶途径是研究最早的学习记忆相关通路,活化的PKA可直接激活下游CREB而调控学习记忆功能。ERK信号通路在中枢神经系统中具有调控细胞增殖分化和学习记忆等多种功能,与脑内长时程增强(LTP)的形成以及空间学习记忆功能有重要的联系。本研究中,检测CREB上游的主要调控激酶发现ERK的磷酸化蛋白表达也显著降低,而PKA、p-CaMKⅡ蛋白表达则没有明显改变。由于ERK和CREB均在空间记忆和长时程记忆中起重要作用,表明ERK-CREB信号途径可能是CPF重复染毒致长时程空间记忆障碍的重要调控通路。
综上所述,CPF低剂量重复染毒能够抑制ERK-CREB信号途径的磷酸化,从而影响海马DG区神经发生,导致海马新生神经细胞减少,最终诱发长时程空间记忆障碍。
Organophosphorus insecticides (OPs) are the most widely used group of pesticides in the world. Because of their relatively low cost, OPs have been used in agriculture, homes, gardens, and in veterinary practice. The use of OPs around the world continues to produce detectable levels of OPs in the environment, thereby posing an ongoing risk for low-level exposure. Chronic exposures to OPs, especially the low-level envireonmental exposures, require greater attention.
The epidemicology studies had reported memory impairments and behavioral alterations after long-term exposures to OPs, and the decrease cognitive function appeared not immediately after the OPs exposure but monthes later. However, there are a few animals studies available that focused on the mechanisam of cognitive impairments on chronic low-level OPs exposure. Some studies in which animals were used as experimental subjects had contradictory results on the cognitive and behavioral effects of OPs exposure. Therefor, the results and mechanisam of chronic low-level OPs exposure need further study.
Chlorpyrifos (O,O’-dithyl-O-3,5,6-trichloro-2-pyrydyl phosphorothionate, CPF) is one of the most widely used OPs throughout the world. Despite the EPA (U.S. Environmental Protection Agency.2002) has entered into an agreement with the registrant to begin terminating residential CPF indoor uses since 2000, it is still one of the most widely used insecticides. With the elimination of acephatemet, methyl parathion, parathion, monocrotophos and phosphamidon in agriculture since 2007, CPF had been recommend as the low-toxic OPs replacement in agriculture in China.These days, CPF has become one of the major pesticides detected in farm products in China.
In the present study, male rats were administrated CPF through intragastric for 12 weeks to evaluate the neurobehavioral impacts of CPF at the subthreshold doses(doses that do not produce overt signs of cholinergic toxicity, e.g. excessive salivation, urination, defecation, muscle fasiculations and so on). The neurobehavioral changes (especially the spatial memory) were studied and the and mechanisam of chronic low-level OPs exposure was explored.
There were no effects on weight gain and food consumption detected during the 12 weeks exposures to CPF, and the daily dosages of 1.0, 5.0 and 10.0 mg/kg CPF for 12 weeks did not elicit any other signs of OPs toxicity, such as excessive salivation, urination, defecation, muscle fasiculations etc. However, the animals exhibited obvious neurobehavioral changes (especially the long-term spatial memory) after 12 weeks repeated exposures to CPF. The increased latency in the Morris water maze during the treated animals indicated that the chronic low-level CPF exposures can impair the long-term spatial memory. As there was no difference between the CPF treated groups and the vehicle treated group on the results from grip strength effects of CPF, and there were no effects on short-term spatial memory or learning. What’s more, only the highest dosage (10mg/kg) had significant inhibition effects on AChE activity, the exhibition of impaired long-term spatial memory may exist non- cholinergic reasons.
The hippocampus has become the most important method to study the higher cognitive function, since its special function in the spatial memory. In the present study, we found significant decrease of cell numbers in the regions of hippocampus, although the hippocampal regions from the CPF treated groups did not exhibit any obvious architectural difference from the vehicle control group on gross observation. However, the immunohistochemisty study exhibited that NeuN (mature neuron marker) positive cells in the dentate gyrus(DG)increased, and the western blot of the NeuN protein from the whole hippocampi also increased significantly. The results from immunohistochemisty and western blot study showed thatβ-3 tubulin(TUJ1)and NF68 (adult new born neuron marker) decreased significantly and the marker of neuron precursor cells, BrdU, also exhibited significant decrease in a dose-dependent manner. The phenomenon discussed above illustrated that the repeated exposures of CPF can affect the constitute of cells in hipocampus, in another words, repeated exposures of CPF lead to the decrease of the adult new born neurons, but have no effect on the mature neurons. The hurt to brain(hypoxia, infection,poisoning,et al.) often exhibited glial cell hyperplasia under normal circumstances, while the GFAP protein expression protein from the whole hippocampi and the GFAP positive cells in the dentate gyrus(DG)decreased simultaneously. The changes that discussed above indicated the decrease of the cell in DG may not the direct toxicity effect from CPF, but the regulation result of some signal pathway. What’s more, the TUNEL positive cells did not show any increase in the treated animals, and apoptosis related regulator , Bax/Bcl-2、p53 and caspase-3 did not show any change after the repeated exposures to CPF. It means the apoptosis could not be the main reason of the cells decrease in the DG. The changes of the cell constitute in the DG indicate the repeated CPF exposures can affect the neurogensis of the hippocampus.
More and more studies had proved that the DG region of the adult hippocampus can have neurogensis all the life, and the adult new born neurons have important function in the hippocampus related spatial memory, especially the long-term memory. The decrease of the new born neurons can result in the impairment of long-term memory. Then the significant decrease of the adult new born neurons in the DG region may be the main reason of the impaired long-term spatial memory in the CPF treated animals.
The regulation of the adult neurogensis is the research focus of neurobiology studies, and the latest results indicated that the cyclic AMP responsive element binding protein (CREB) have an important function during the Memory Allocation. The phosphorylated CREB can activate the dowmstream gene transcription, which can induce the differentiation of the hippocampal precursor cells and promote the survival and proliferation of the adult new born neurons. The decrease of the phosphorylated CREB can lead to deficiency of the adult new born neurons. As CREB is one of the most prominent molecules involved in learning and memory processes. Evidence suggests that CREB may play a crucial role in hippocampal plasticity and consolidation of long-term memory, since CREB activates the transcription of genes required for long-term synaptic changes involved in memory formation. Animals with CREB protein mutations exhibit learning and memory deficits. While, the activation of CREB pathway can be the regulating results of Ca2+/calmodulin-dependent kinaseⅡ(CaMKⅡ)、protein phosphakinase A (PKA)、extracellular regulated protein kinases (ERK). CaMKⅡhas been regarded as the molecular switch during the leaning and memory function. The cAMP related protein kinases pathway was the first memory related signal pathway studied, and the PKA can active CREB directly. ERK, as the upstream kinases of CREB pathway have important function during the neuron differentiation and long-term potentiation(LTP).
In the present study, the expression protein level of phosphorylated CREB and ERK1/2 in hippocampus from the CPF treated rats decreased significantly, but the CPF had no effects on the expression of PKA and phosphorylated CaMKⅡ. The dose-dependent decrease indicated that the inhibition of the ERK-CREB Signaling may be one of the important regulating pathway during the CPF caused spatial long-term memory impairments.
In summary, the present study identified that repeated exposure to subthreshold doses of CPF can lead to the inhibition of ERK-CREB signaling, which regulated the neruogenesis of the hippocampal DG and induced the deficiency of adult new born neurons. The decreased young neurons lead to the impaired long-term spatial memory.
引文
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