Possible Role of Protein CPG15 in Hippocampal Mossy Fiber Sprouting Under Conditions of Pentylenetetrazole Kindling

We examined changes in expression of the candidate plasticity-related gene 15 (CPG15) in the dentate gyrus (DG) and hippocampal CA3 region in the pentylenetetrazole (PTZ) kindling model and investigated the role of this gene in the phenomenon of mossy fiber sprouting (MFS). Experimental rats were divided into the control and PTZ groups. The epileptic model was created by intraperitoneal PTZ injection, while control rats were injected with saline. At days 3, 7, 14, 28, and 42 after the first PTZ injection, Timm staining was scored in the CA3 hippocampal area, and a product of CPG15 (protein CPG15) was labeled in the DG stratum granulosum and in the CA3 area using immunohistochemistry. The Timm scores in the CA3 region increased gradually from day 3 and were significantly higher than those in the control within the subsequent period. The level of CPG15 protein in the DG and CA3 area decreased gradually until day 14 and returned to the normal level at day 28. The results obtained indicate, for the first time, that CPG15 may be involved in the process of MFS. Understanding the molecular mechanisms underlying this phenomenon may lead to successful therapeutic interventions that limit epileptogenesis.Ми досліджували зміни експресії продукту гена CPG15 у зубчастій звивині (ЗЗ) та зоні CA3 гіпокампа в моделі пентилентетразолового (ПТЗ-) кіндлінгу та можливу роль цього гена у феномені спрутингу моховитих волокон (СМВ). Піддослідні щури були поділені на групи контролю та ПТЗ-кіндлінгу. Модель епілепсії створювали за допомогою внутрішньоочеревинних ін’єкцій пентилентетразолу (ПТЗ); контрольним щурам ін’єкували фізіологічний розчин. На третю, сьому, 14-ту, 28-му та 42-гу добу після першої ін’єкції ПТЗ оцінювали забарвлення, за Тіммом, у зоні CA3. Локалізацію протеїну CPG15 у stratum granulosum ЗЗ та зоні CA3 гіпокампа визначали з використанням імуногістохімічної методики. Інтенсивність забарвлення, за Тіммом, у зоні CA3 поступово збільшувалася починаючи з третьої доби та була вірогідно вищою, ніж така в контролі, протягом усього наступного періоду. Рівень протеїну CPG15 у ЗЗ та полі CA3 поступово зменшувався до 14-ї доби та повертався до нормальних значень на 28-му добу. Отримані результати вперше вказують на те, що CPG15 може бути залученим у процес СМВ. Зрозуміння молекулярних механізмів, на яких базується цей феномен, може призвести до розробки успішних терапевтичних заходів, котрі обмежували б епілептогенез

Cloning and Phylogenetic Analysis of NMDA Receptor Subunits NR1, NR2A and NR2B in Xenopus laevis Tadpoles

N-methyl-d-aspartate receptors (NMDARs) play an important role in many aspects of nervous system function such as synaptic plasticity and neuronal development. NMDARs are heteromers consisting of an obligate NR1 and most commonly one or two kinds of NR2 subunits. While the receptors have been well characterized in some vertebrate and invertebrate systems, information about NMDARs in Xenopus laevis brain is incomplete. Here we provide biochemical evidence that the NR1, NR2A and NR2B subunits of NMDARs are expressed in the central nervous system of X. laevis tadpoles. The NR1-4a/b splice variants appear to be the predominant isoforms while the NR1-3a/b variants appear to be expressed at low levels. We cloned the X. laevis NR2A and NR2B subunits and provide a detailed annotation of their functional domains in comparison with NR2A and NR2B proteins from 10 and 13 other species, respectively. Both NR2A and NR2B proteins are remarkably well conserved between species, consistent with the importance of NMDARs in nervous system function

ERK/MAPK Is Essential for Endogenous Neuroprotection in SCN2.2 Cells

Glutamate (Glu) is essential to central nervous system function; however excessive Glu release leads to neurodegenerative disease. Strategies to protect neurons are underdeveloped, in part due to a limited understanding of natural neuroprotective mechanisms, such as those present in the suprachiasmatic nucleus (SCN). This study tests the hypothesis that activation of ERK/MAPK provides essential protection to the SCN after exposure to excessive Glu using the SCN2.2 cells as a model.Immortalized SCN2.2 cells (derived from SCN) and GT1-7 cells (neurons from the neighboring hypothalamus) were treated with 10 mM Glu in the presence or absence of the ERK/MAPK inhibitor PD98059. Cell death was assessed by Live/Dead assay, MTS assay and TUNEL. Caspase 3 activity was also measured. Activation of MAPK family members was determined by immunoblot. Bcl2, neuritin and Bid mRNA (by quantitative-PCR) and protein levels (by immunoblot) were also measured.As expected Glu treatment increased caspase 3 activity and cell death in the GT1-7 cells, but Glu alone did not induce cell death or affect caspase 3 activity in the SCN2.2 cells. However, pretreatment with PD98059 increased caspase 3 activity and resulted in cell death after Glu treatment in SCN2.2 cells. This effect was dependent on NMDA receptor activation. Glu treatment in the SCN2.2 cells resulted in sustained activation of the anti-apoptotic pERK/MAPK, without affecting the pro-apoptotic p-p38/MAPK. In contrast, Glu exposure in GT1-7 cells caused an increase in p-p38/MAPK and a decrease in pERK/MAPK. Bcl2-protein increased in SCN2.2 cells following Glu treatment, but not in GT1-7 cells; bid mRNA and cleaved-Bid protein increased in GT1-7, but not SCN2.2 cells.Facilitation of ERK activation and inhibition of caspase activation promotes resistance to Glu excitotoxicity in SCN2.2 cells.Further research will explore ERK/MAPK as a key molecule in the prevention of neurodegenerative processes

Therapy Development for Spinal Muscular Atrophy in SMN Independent Targets

Spinal muscular atrophy (SMA) is an autosomal recessive neurodegenerative disorder, leading to progressive muscle weakness, atrophy, and sometimes premature death. SMA is caused by mutation or deletion of the survival motor neuron-1 (SMN1) gene. An effective treatment does not presently exist. Since the severity of the SMA phenotype is inversely correlated with expression levels of SMN, the SMN-encoded protein, SMN is the most important therapeutic target for development of an effective treatment for SMA. In recent years, numerous SMN independent targets and therapeutic strategies have been demonstrated to have potential roles in SMA treatment. For example, some neurotrophic, antiapoptotic, and myotrophic factors are able to promote survival of motor neurons or improve muscle strength shown in SMA mouse models or clinical trials. Plastin-3, cpg15, and a Rho-kinase inhibitor regulate axonal dynamics and might reduce the influences of SMN depletion in disarrangement of neuromuscular junction. Stem cell transplantation in SMA model mice resulted in improvement of motor behaviors and extension of survival, likely from trophic support. Although most therapies are still under investigation, these nonclassical treatments might provide an adjunctive method for future SMA therapy

Insulin receptor signaling in the development of neuronal structure and function

Sensory experience plays a crucial role in regulating neuronal shape and in developing synaptic contacts during brain formation. These features are required for a neuron to receive, integrate, and transmit signals within the neuronal network so that animals can adapt to the constant changing environment. Insulin receptor signaling, which has been extensively studied in peripheral organ systems such as liver, muscle and adipocyte, has recently been shown to play important roles in the central nervous system. Here we review the current understanding of the underlying mechanisms that regulate structural and functional aspects of circuit development, particularly with respect to the role of insulin receptor signaling in synaptic function and the development of dendritic arbor morphology. The potential link between insulin receptor signaling malfunction and neurological disorders will also be discussed

Optic nerve crush induces spatial and temporal gene expression patterns in retina and optic nerve of BALB/cJ mice

BACKGROUND: Central nervous system (CNS) trauma and neurodegenerative disorders trigger a cascade of cellular and molecular events resulting in neuronal apoptosis and regenerative failure. The pathogenic mechanisms and gene expression changes associated with these detrimental events can be effectively studied using a rodent optic nerve crush (ONC) model. The purpose of this study was to use a mouse ONC model to: (a) evaluate changes in retina and optic nerve (ON) gene expression, (b) identify neurodegenerative pathogenic pathways and (c) discover potential new therapeutic targets. RESULTS: Only 54% of total neurons survived in the ganglion cell layer (GCL) 28 days post crush. Using Bayesian Estimation of Temporal Regulation (BETR) gene expression analysis, we identified significantly altered expression of 1,723 and 2,110 genes in the retina and ON, respectively. Meta-analysis of altered gene expression (≥1.5, ≤-1.5, p < 0.05) using Partek and DAVID demonstrated 28 up and 20 down-regulated retinal gene clusters and 57 up and 41 down-regulated optic nerve clusters. Regulated gene clusters included regenerative change, synaptic plasticity, axonogenesis, neuron projection, and neuron differentiation. Expression of selected genes (Vsnl1, Syt1, Synpr and Nrn1) from retinal and ON neuronal clusters were quantitatively and qualitatively examined for their relation to axonal neurodegeneration by immunohistochemistry and qRT-PCR. CONCLUSION: A number of detrimental gene expression changes occur that contribute to trauma-induced neurodegeneration after injury to ON axons. Nrn1 (synaptic plasticity gene), Synpr and Syt1 (synaptic vesicle fusion genes), and Vsnl1 (neuron differentiation associated gene) were a few of the potentially unique genes identified that were down-regulated spatially and temporally in our rodent ONC model. Bioinformatic meta-analysis identified significant tissue-specific and time-dependent gene clusters associated with regenerative changes, synaptic plasticity, axonogenesis, neuron projection, and neuron differentiation. These ONC induced neuronal loss and regenerative failure associated clusters can be extrapolated to changes occurring in other forms of CNS trauma or in clinical neurodegenerative pathological settings. In conclusion, this study identified potential therapeutic targets to address two key mechanisms of CNS trauma and neurodegeneration: neuronal loss and regenerative failure

Androgen regulation of axon growth and neurite extension in motoneurons

Androgens act on the CNS to affect motor function through interaction with a widespread distribution of intracellular androgen receptors (AR). This review highlights our work on androgens and process outgrowth in motoneurons, both in vitro and in vivo. The actions of androgens on motoneurons involve the generation of novel neuronal interactions that are mediated by the induction of androgen-dependent neurite or axonal outgrowth. Here, we summarize the experimental evidence for the androgenic regulation of the extension and regeneration of motoneuron neurites in vitro using cultured immortalized motoneurons, and axons in vivo using the hamster facial nerve crush paradigm. We place particular emphasis on the relevance of these effects to SBMA and peripheral nerve injurie

Activity-regulated genes as mediators of neural circuit plasticity

Modifications of neuronal circuits allow the brain to adapt and change with experience. This plasticity manifests during development and throughout life, and can be remarkably long lasting. Evidence has linked activity-regulated gene expression to the long-term structural and electrophysiological adaptations that take place during developmental critical periods, learning and memory, and alterations to sensory map representations in the adult. In all these cases, the cellular response to neuronal activity integrates multiple tightly coordinated mechanisms to precisely orchestrate long-lasting, functional and structural changes in brain circuits. Experience-dependent plasticity is triggered when neuronal excitation activates cellular signaling pathways from the synapse to the nucleus that initiate new programs of gene expression. The protein products of activity-regulated genes then work via a diverse array of cellular mechanisms to modify neuronal functional properties. Synaptic strengthening or weakening can reweight existing circuit connections, while structural changes including synapse addition and elimination create new connections. Posttranscriptional regulatory mechanisms, often also dependent on activity, further modulate activity-regulated gene transcript and protein function. Thus, activity-regulated genes implement varied forms of structural and functional plasticity to fine-tune brain circuit wiring.National Institutes of Health (U.S.). Ruth L. Kirschstein National Research Service Award (F31 NS069510)RO1 EY01189

Cellular and molecular analysis of neuronal structure plasticity in the mammalian cortex

Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Brain and Cognitive Sciences, 2006.This electronic version was submitted by the student author. The certified thesis is available in the Institute Archives and Special Collections.Includes bibliographical references (p. 87-100).Despite decades of evidence for functional plasticity in the adult brain, the role of structural plasticity in its manifestation remains unclear. cpg15 is an activity-regulated gene encoding a membrane-bound ligand that coordinately regulates growth of apposing dendritic and axonal arbors and the maturation of their synapses. Here we compare cpg15 expression during normal development of the rat visual system, with that seen in response to dark rearing, monocular retinal action potential blockade, or monocular deprivation. Our results show that: (1) cpg15 expression in visual cortex correlates with the electrophysiologically mapped critical period for development of eye-specific preference in the primary visual cortex. (2) Dark rearing elevates adult levels of expression. (3) A component of cpg15 expression is activity-dependent after the peak of the critical period. (4) At the peak of the critical period, monocular deprivation decreases cpg15 expression more than monocular TTX blockade. And (5) cpg15 expression is robust and regulated by light in the superficial layers of the adult visual cortex.(cont.) This suggests that cpg15 is an excellent molecular marker for the visual system's capacity for plasticity and predicts that neural remodeling normally occurs in the extragranular layers of the adult visual cortex. To examine the extent of neuronal remodeling that occurs in the brain on a daily basis, we used a multi-photon based microscopy system for chronic in vivo imaging and reconstruction of entire neurons in the superficial layers of the rodent cerebral cortex. Here, we show the first unambiguous evidence of dendrite growth and remodeling in adult neurons. Over a period of months, neurons could be seen extending and retracting existing branches, and in rare cases adding new branch tips. Neurons exhibiting dynamic arbor rearrangements were GABA positive non-pyramidal interneurons, while pyramidal cells remained stable. These results are consistent with the idea that dendritic structural remodeling is a substrate for adult plasticity and suggest that circuit rearrangement in the adult cortex is restricted by cell type-specific rules.by Wei-Chung Allen Lee.Ph.D