Participation of CaMKII in neuronal plasticity and memory formation

1. The unique biochemical properties of Ca²⁺/calmodulin (CaM)-dependent protein kinase II have made this enzyme one of the paradigmatic models of the forever searched memory molecule.2. In particular, the central participation of CaMKII as a sensor of the Ca²⁺ signals generated by activation of NMDA receptors after the induction of long-term plastic changes, has encouraged the use of pharmacological, genetic, biochemical, and imaging tools to unveil the role of this kinase in the acquisition, consolidation, and expression of different types of memories.3. Here we review some of the more exciting discoveries related to the mechanisms involved in CaMKII activation and synaptic plasticity.     

艾灸对缺氧缺血性脑损伤新生小鼠的治疗作用*

目的: 探讨艾灸对缺氧缺血性脑损伤新生小鼠行为学表现、脑组织形态结构的影响及作用机制。方法: 将106只出生7 d小鼠随机分为三组:假手术组(23只)、模型组(46只)和艾灸组(37只)。采用左侧颈总动脉结扎后再置于37℃密闭舱内进行低氧处理(氧气浓度为8%,100 min),制备新生儿缺氧缺血性脑病动物模型。艾灸组同模型组,并于造模后2 h开始艾灸“大椎”进行治疗,以后每日1次,每次35 min,连续治疗4 d。采用行为学测试评价小鼠的行为学表现;HE染色观察小鼠脑组织形态结构;Western blot技术检测小鼠脑组织超氧化物歧化酶2(SOD2)蛋白表达;比色法测定小鼠脑组织丙二醛(MDA)含量。结果: 假手术组小鼠行为表现正常,脑组织细胞排列致密整齐,脑组织SOD2蛋白表达量和MDA含量正常。与假手术组相比,模型组小鼠翻正反射、趋地反射、悬崖躲避试验时间延长(P＜0.05),抓力试验时间缩短(P＜0.05);脑组织细胞大量坏死脱落;脑组织SOD2蛋白表达量明显减少(P＜0.05)、MDA含量增加。与模型组相比,艾灸组小鼠翻正反射、趋地反射、悬崖躲避试验时间缩短(P＜0.05),抓力试验时间增长(P＜0.05);脑组织细胞排列较致密、整齐;脑组织SOD2蛋白表达量增多(P＜0.05)、MDA含量降低(P＜0.05)。结论: 艾灸能减轻缺氧缺血性脑病新生小鼠脑损伤、改善行为学表现,这可能与其增加脑组织SOD2蛋白的表达、降低MDA含量,从而提高抗氧化应激能力有关。     

Differential roles of CaMKII isoforms in phase separation with NMDA receptors and in synaptic plasticity

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Dynamics underlying synaptic gain between pairs of cortical pyramidal neurons

Changes in connectivity between pairs of neurons can serve as a substrate for information storage and for experience-dependent changes in neuronal circuitry. Early in development, synaptic contacts form and break, but how these dynamics influence the connectivity between pairs of neurons is not known. Here we used time-lapse imaging to examine the synaptic interactions between pairs of cultured cortical pyramidal neurons, and found that the axon-dendrite contacts between each neuronal pair were composed of both a relatively stable and a more labile population. Under basal conditions, loss and gain of contacts within this labile population was well balanced and there was little net change in connectivity. Selectively increasing the levels of activated CaMKII in the postsynaptic neuron increased connectivity between pairs of neurons by increasing the rate of gain of new contacts without affecting the probability of contact loss, or the proportion of stable and labile contacts, and this increase required Calcium/calmodulin binding to CaMKII. Our data suggest that activating CaMKII can increase synaptic connectivity through a CaM-dependent increase in contact formation, followed by stabilization of a constant fraction of new contacts.     

SDF-1/CXCR4轴在缺氧缺血性脑损伤中的研究进展

干细胞在许多组织器官显示巨大的细胞分化潜能,其治疗缺血缺氧性疾病成为当前研究的热点。已知局部缺血可诱导干细胞的动员,并能感受组织损伤而定向迁移到损伤区并进行分化。具有趋化因子受体4（CXC chemokine receptor 4,CXCR4）的干细胞迁移到高表达间质细胞来源的因子-1（stromal cell-derived factor-1,SDF-1）的组织区域,这种细胞的迁移运动能被CXCR4拈抗剂所阻断或通过CXCR4的过表达增强迁移的运动。SDF-1-CXCR4轴是体内各种类型的干细胞迁移及细胞在骨髓的滞留和归巢中的重要调节物质。本文就缺氧缺血性脑损伤的骨髓间质干细胞（bone marrow stromal cell,BMSC）治疗,SDF- 1-CXCR4轴在MSCs动员和损伤、修复中的作用作一综述。     

Membrane traffic during axon development

Brain formation requires the establishment of complex neural circuits between a diverse array of neuronal subtypes in an intricate and ever changing microenvironment and yet with a large degree of specificity and reproducibility. In the last three decades, mounting evidence has established that neuronal development relies on the coordinated regulation of gene expression, cytoskeletal dynamics, and membrane trafficking. Membrane trafficking has been considered important in that it brings new membrane and proteins to the plasma membrane of developing neurons and because it also generates and maintains the polarized distribution of proteins into neuronal subdomains. More recently, accumulating evidence suggests that membrane trafficking may have an even more active role during development by regulating the distribution and degree of activation of a wide variety of proteins located in plasma membrane subdomains and endosomes. In this article the evidence supporting the different roles of membrane trafficking during axonal development, particularly focusing on the role of SNAREs and Rabs was reviewed. © 2016 Wiley Periodicals, Inc. Develop Neurobiol 76: 1185–1200, 2016     

Synapse formation and remodeling

Synapses are specialized structures that mediate information flow between neurons and target cells,and thus are the basis for neuronal system to execute various functions,including learning and memory.There are around 1011 neurons in the human brain,with each neuron receiving thousands of synaptic inputs,either excitatory or inhibitory.A synapse is an asymmetric structure that is composed of pre-synaptic axon terminals,synaptic cleft,and postsynaptic compartments.Synapse formation involves a number of cell ...     

脊椎动物脑发育中糖皮质激素受体功能的研究进展

糖皮质激素受体（glucocorticoid reccptor,GR）广泛分布在脊椎动物中枢神经系统的多个组织区域中,而且结构及功能保守。在与激素结合的状态下,受体能够特异性地与靶基因的启动子结合影响基因的表达,或通过激活G蛋白偶联的信号途径引起神经递质的释放。外界环境刺激和外源糖皮质激素暴露都能改变GR在脑中的表达,并对神经的发育及功能产生影响,同时也对学习、记忆以及情感等高级神经活动和行为起到重要的作用。该文对脊椎动物糖皮质激素受体的结构和在脑中的分布,以及对神经发育和功能的影响及其中的分子机制的最新研究进展进行综述。     

CaMKII regulates pericyte loss in the retina of early diabetic mouse

Inducible nitric oxide synthase (iNOS) is an essential mediator in diabetic vascular lesions and known to be regulated by activation of Ca²⁺/calmodulin-dependent protein kinase II (CaMKII). The aim of this study was to investigate whether CaMKII affects iNOS-mediated pericyte death in the retina of diabetic mice with early stage disease. Total- and phospho-CaMKII, iNOS, and active caspase-3 protein levels were assessed by Western blotting, and CaMKII activity was measured by kinase assay. iNOS-related pericyte death was assessed by double immunofluorescent staining for iNOS and α-smooth muscle actin, followed by the TUNEL assay. Autocamtide-2-related inhibitory peptide (AIP), a specific inhibitor of CaMKII, was injected into the right vitreous 2 days before sacrifice of mice, to examine the effect of CaMKII inactivation in diabetic retinas. The levels of total- and phospho-CaMKII, iNOS, and active caspase-3 protein, and CaMKII activity were significantly increased in the diabetic retinas compared with those of control retinas. Furthermore, TUNEL-positive signals colocalized with iNOS-immunoreactive pericytes in the same retinas. However, inactivation of CaMKII by AIP treatment inhibited all these changes, which was accompanied by less pericyte loss. Our results demonstrate that CaMKII contributes to iNOS-related death of pericytes in the diabetic retina and that inactivation of this enzyme may be a potential treatment for retinal vascular lesion.     

Stimulation of autophagy by rapamycin protects neurons from remote degeneration after acute focal brain damage

Autophagy is the evolutionarily conserved degradation and recycling of cellular constituents. In mammals, autophagy is implicated in the pathogenesis of many neurodegenerative diseases. However, its involvement in acute brain damage is unknown. This study addresses the function of autophagy in neurodegeneration that has been induced by acute focal cerebellar lesions. We provide morphological, ultrastructural, and biochemical evidence that lesions in a cerebellar hemisphere activate autophagy in axotomized precerebellar neurons. Through time course analyses of the apoptotic cascade, we determined mitochondrial dysfunction to be the early trigger of degeneration. Further, the stimulation of autophagy by rapamycin and the employment of mice with impaired autophagic responses allowed us to demonstrate that autophagy protects from damage promoting functional recovery. These findings have therapeutic significance, demonstrating the potential of pro-autophagy treatments for acute brain pathologies, such as stroke and brain trauma.     

Notch: from neural development to neurological disorders

Notch is an integral membrane protein that functions as receptor for ligands such as jagged and delta that are associated with the surface of neighboring cells. Upon ligand binding, notch is proteolytically cleaved within its transmembrane domain by presenilin‐1 (the enzymatic component of the γ‐secretase complex) resulting in the release of a notch intracellular domain which translocates to the nucleus where it regulates gene expression. Notch signaling plays multiple roles in the development of the CNS including regulating neural stem cell (NSC) proliferation, survival, self‐renewal and differentiation. Notch is also present in post‐mitotic neurons in the adult CNS wherein its activation influences structural and functional plasticity including processes involved in learning and memory. Recent findings suggest that notch signaling in neurons, glia, and NSCs may be involved in pathological changes that occur in disorders such as stroke, Alzheimer’s disease and CNS tumors. Studies of animal models suggest the potential of agents that target notch signaling as therapeutic interventions for several different CNS disorders.     

Targeted electroporation in Xenopus tadpoles in vivo--from single cells to the entire brain

Electroporation is becoming more popular as a technique for transfecting neurons within intact tissues. One of the advantages of electroporation over other transfection techniques is the ability to precisely target an area for transfection. Here we highlight this advantage by describing methods to restrict transfection to either a single cell, clusters of cells, or to include large portions of the brain of the intact Xenopus tadpole. Electroporation is also an effective means of gene delivery in the retina. We have developed these techniques to examine the effects of regulated gene expression on various neuronal properties, including structural plasticity and synaptic transmission. Restriction of transfection to individual cells aids in imaging of neuronal morphology, while bulk cell transfection allows examination of the affects of gene expression on populations of cells by biochemical assays, imaging, and electrophysiological recording.     

Impaired autophagy flux is associated with neuronal cell death after traumatic brain injury

Dysregulation of autophagy contributes to neuronal cell death in several neurodegenerative and lysosomal storage diseases. Markers of autophagy are also increased after traumatic brain injury (TBI), but its mechanisms and function are not known. Following controlled cortical impact (CCI) brain injury in GFP-Lc3 (green fluorescent protein-LC3) transgenic mice, we observed accumulation of autophagosomes in ipsilateral cortex and hippocampus between 1 and 7 d. This accumulation was not due to increased initiation of autophagy but rather to a decrease in clearance of autophagosomes, as reflected by accumulation of the autophagic substrate SQSTM1/p62 (sequestosome 1). This was confirmed by ex vivo studies, which demonstrated impaired autophagic flux in brain slices from injured as compared to control animals. Increased SQSTM1 peaked at d 1–3 but resolved by d 7, suggesting that the defect in autophagy flux is temporary. The early impairment of autophagy is at least in part caused by lysosomal dysfunction, as evidenced by lower protein levels and enzymatic activity of CTSD (cathepsin D). Furthermore, immediately after injury both autophagosomes and SQSTM1 accumulated predominantly in neurons. This was accompanied by appearance of SQSTM1 and ubiquitin-positive puncta in the affected cells, suggesting that, similar to the situation observed in neurodegenerative diseases, impaired autophagy may contribute to neuronal injury. Consistently, GFP-LC3 and SQSTM1 colocalized with markers of both caspase-dependent and caspase-independent cell death in neuronal cells proximal to the injury site. Taken together, our data indicated for the first time that autophagic clearance is impaired early after TBI due to lysosomal dysfunction, and correlates with neuronal cell death.     

Impaired autophagy flux is associated with neuronal cell death after traumatic brain injury

Dysregulation of autophagy contributes to neuronal cell death in several neurodegenerative and lysosomal storage diseases. Markers of autophagy are also increased after traumatic brain injury (TBI), but its mechanisms and function are not known. Following controlled cortical impact (CCI) brain injury in GFP-Lc3 (green fluorescent protein-LC3) transgenic mice, we observed accumulation of autophagosomes in ipsilateral cortex and hippocampus between 1 and 7 d. This accumulation was not due to increased initiation of autophagy but rather to a decrease in clearance of autophagosomes, as reflected by accumulation of the autophagic substrate SQSTM1/p62 (sequestosome 1). This was confirmed by ex vivo studies, which demonstrated impaired autophagic flux in brain slices from injured as compared to control animals. Increased SQSTM1 peaked at d 1–3 but resolved by d 7, suggesting that the defect in autophagy flux is temporary. The early impairment of autophagy is at least in part caused by lysosomal dysfunction, as evidenced by lower protein levels and enzymatic activity of CTSD (cathepsin D). Furthermore, immediately after injury both autophagosomes and SQSTM1 accumulated predominantly in neurons. This was accompanied by appearance of SQSTM1 and ubiquitin-positive puncta in the affected cells, suggesting that, similar to the situation observed in neurodegenerative diseases, impaired autophagy may contribute to neuronal injury. Consistently, GFP-LC3 and SQSTM1 colocalized with markers of both caspase-dependent and caspase-independent cell death in neuronal cells proximal to the injury site. Taken together, our data indicated for the first time that autophagic clearance is impaired early after TBI due to lysosomal dysfunction, and correlates with neuronal cell death.     

Chronic psychosocial stress enhances long-term depression in a subthreshold amyloid-beta rat model of Alzheimer's disease

In addition to genetic aspects, environmental factors such as stress may also play a critical role in the etiology of the late onset, sporadic Alzheimer's disease (AD). The present study examined the effect of chronic psychosocial stress in a sub-threshold Aβ (subAβ) rat model of AD on long-term depression by two techniques: electrophysiological recordings of synaptic plasticity in anesthetized rats, and immunoblot analysis of memory- and AD-related signaling molecules. Chronic psychosocial stress was induced using a rat intruder model. The subAβ rat model of AD, which was intended to represent outwardly normal individuals with a pre-disposition to AD, was induced by continuous infusion of 160 pmol/day Aβ???? via a 14-day i.c.v. osmotic pump. Results from electrophysiological recordings showed that long-term depression evoked in stress/subAβ animals was significantly enhanced compared with that in animals exposed to stress or subAβ infusion alone. Molecular analysis of various signaling molecules 1 h after induction of long-term depression revealed an increase in the levels of calcineurin and phosphorylated CaMKII in groups exposed to stress compared with other groups. The levels of the brain-derived neurotrophic factor (BDNF) were significantly decreased in stress/subAβ animals but not in stress or subAβ animals. In addition, the levels of beta-site amyloid precursor protein cleaving enzyme were markedly increased in stress/subAβ. These findings suggest that chronic stress may accelerate the impairment of synaptic plasticity and consequently cognition in individuals 'at-risk' for AD.     

Biochemical signaling networks decode temporal patterns of synaptic input

Synapses exhibit a wide repertoire of responses to different temporal patterns of synaptic input. Many of these responses are expressed as short and long-term changes in synaptic strength. Electrical properties of channels and calcium buildup can account for rapid aspects of pattern decoding, but it is not clear how more complex input patterns, especially those lasting over many minutes, could be discriminated. This paper shows that a network of signaling pathways can discriminate between complex input patterns lasting tens of minutes, and can give rise to distinct combinatorial patterns of biochemical signaling activity in pathways involved in synaptic change. Regulatory signaling input can alter and even reverse the strengths of responses to input patterns. Thus the synaptic signaling network may function as a temporal decoder that transforms patterns from the time domain into the domain of chemical signaling. This may underlie different synaptic responses to different stimulus patterns.Supplementary material to this paper is available in electronic form at http://dx.doi.org/10.1023/A:1019644427655     

Effects of streptozotocin-diabetes on the hippocampal NMDA receptor complex in rats

In animal models of diabetes mellitus, such as the streptozotocin-diabetic rat (STZ-rat), spatial learning impairments develop in parallel with a reduced expression of long-term potentiation (LTP) and enhanced expression of long-term depression (LTD) in the hippocampus. This study examined the time course of the effects of STZ-diabetes and insulin treatment on the hippocampal post-synaptic glutamate N-methyl-D-aspartate (NMDA) receptor complex and other key proteins regulating hippocampal synaptic transmission in the post-synaptic density (PSD) fraction. In addition, the functional properties of the NMDA-receptor complex were examined. One month of STZ-diabetes did not affect the NMDA receptor complex. In contrast, 4 months after induction of diabetes NR2B subunit immunoreactivity, CaMKII and Tyr-dependent phosphorylation of the NR2A/B subunits of the NMDA receptor were reduced and alphaCaMKII autophosphorylation and its association to the NMDA receptor complex were impaired in STZ-rats compared with age-matched controls. Likewise, NMDA currents in hippocampal pyramidal neurones measured by intracellular recording were reduced in STZ-rats. Insulin treatment prevented the reduction in kinase activities, NR2B expression levels, CaMKII-NMDA receptor association and NMDA currents. These findings strengthen the hypothesis that altered post-synaptic glutamatergic transmission is related to deficits in learning and plasticity in this animal model.