Inhibitory neurons play important roles in a number of brain functions. They are composed of GABAergic neurons and glycinergic neurons, and vesicular GABA transporter (VGAT) is specifically expressed in these neurons. Since the inhibitory neurons are scattered around in the CNS, it is difficult to identify these cells in living brain preparations. The glutamate decarboxylase (GAD) 67-GFP knock-in mouse has been widely used for the identification of GABAergic neurons, but their GAD67 expression was decreased compared to the wild-type mice. To overcome such a problem and to highlight the function and morphology of inhibitory neurons, we generated four lines of VGAT-Venus transgenic mice (lines #04, #29, #39 and #49) expressing Venus fluorescent protein under the control of mouse VGAT promoter. We found higher expression level of Venus transcripts and proteins as well as brighter fluorescent signal in line #39 mouse brains, compared to brains of other lines examined. By Western blots and spectrofluorometric measurements of forebrain, the line #39 mouse showed stronger GFP immunoreactivity and brighter fluorescent intensity than the GAD67-GFP knock-in mouse. In addition, Venus was present not only in somata, but also in neurites in the line #39 mouse by histological studies. In situ hybridization analysis showed that the expression pattern of Venus in the line #39 mouse was similar to that of endogenous VGAT. Double immunostaining analysis in line #39 mouse showed that Venus-expressing cells are primarily immunoreactive for GABA in cerebral cortex, hippocampus and cerebellar cortex and for GABA or glycine in dorsal cochlear nucleus. These results demonstrate that the VGAT-Venus line #39 mouse should be useful for studies on function and morphology of inhibitory neurons in the CNS.

The hippocampus is a multifaceted, complex brain structure considered to be the learning center. The use of primary hippocampal cell cultures has uncovered important cellular mechanisms involved in overall physiological function. Yet, the use of primary culture is inherently difficult, and the lack of immortalized cell lines from the murine hippocampus for mechanistic studies at the molecular level is evident. We have immortalized cell lines from embryonic (E18) and adult-derived hippocampal primary cell culture using retroviral infection of SV40 T-antigen. Four clonal embryonic lines, mHippoE-2, mHippoE-5, mHippoE-14, mHippoE-18, and one mixed adult line, mHippoA-mix, exhibited neuronal morphologies with neurite extensions and expression of neuronal markers, with unique gene expression profiles. We used these cell models to study the neuroprotective effects of 17beta-estradiol (E2) on glutamate-induced neurotoxicity. The cell lines express a relevant array of genes and receptors suggested to play a role in neuroprotection, including estrogen receptors ERalpha, ERbeta, and GPR30. We find that pretreatment with E2 (10 or 100 nM) for 24 h significantly reduced cell death induced by glutamate mHippoE-14 and mHippoE-18 cells, but not the mHippoA-mix. Using 24 h pretreatment with the specific estrogen receptor (ER) agonists, 4,4',4''-(4-propyl-[1H]-pyrazole-1,3,5-triyl)trisphenol (PPT) and diarylpropionitrile, 2,3-bis(4-Hydroxyphenyl)-propionitrile (DPN), we linked the E2-mediated neuroprotection to ERalpha, but only in the mHippoE-18 cells. Since E2 activated both PI3K/Akt and STAT3 signaling pathways, we also tested whether the membrane-bound E2 receptor GPR30 was involved in its neuroprotective action. Pretreatment with the GPR30 agonist G-1 (10 and 100 nM) for 1 h, but not 24 h, significantly attenuated cell death in both mHippoE-14 and mHippoE-18 cells. The use of specific ER antagonist ICI 182780 and GPR30 antagonist G-15 linked these effects to both ER and GPR30 receptors. This is the first evidence that GPR30 may play a role in the protective effects of estrogen in hippocampal neurons.

The cerebellum contains more neurons than all other brain regions combined and these cells exhibit complex circuit development and dendritic elaboration during the postnatal period. Neural development, cellular morphogenesis, and synaptic plasticity are dependent on the dynamic regulation of the actin cytoskeleton by actin-binding proteins. The identification of the actin filament interactome, including proteins developmentally regulated in the postnatal cerebellum, could help define important regulators of actin cytoskeletal dynamics in developing cerebellar neurons. Affinity purification of cerebellar proteins on F-actin columns, combined with mass spectrometry, in total, 434 actin filament-associated proteins in postnatal rat cerebellum (P7) were identified. Furthermore, semi-quantitative RT-PCR was performed to screening postnatal developmentally regulated genes involved in actin dynamics and membrane trafficking in rat cerebellum (P0-P56). As the result, nine genes encoding members of the cerebellar F-actin interactome were developmentally regulated in the transcriptional level and at least five of them exhibited a similar pattern at the protein expression level by Western blot analysis. Further fluorescent immunohistochemical observations demonstrated that the actin-associated proteins Lethal(2) giant larvae protein homolog 1 (LLGL1) and metastasis suppressor 1 (MTSS1) were specifically upregulated in granule neurons and Purkinje cells during morphogenesis of axons and dendrites. This work defines a provisional actin filament interactome in rat postnatal cerebellum and identifies several candidate proteins that may be involved in the postnatal development of the cerebellum.

Huntington's disease is a progressive, inherited neurodegenerative disorder characterized by the loss of subsets of neurons primarily in the striatum. In this study, we assessed the neuroprotective effect of lithium against striatal lesion formation in a rat model of Huntington's disease in which quinolinic acid was unilaterally infused into the striatum. For this purpose, we used a dopamine receptor autoradiography and glutamic acid decarboxylase mRNA in situ hybridization analysis, methods previously shown to be adequate for quantitative analysis of the excitotoxin-induced striatal lesion size. Here we demonstrated that subcutaneous injections of LiCl for 16 days prior to quinolinic acid infusion considerably reduced the size of quinolinic acid-induced striatal lesion. Furthermore, these lithium pre-treatments also decreased the number of striatal neurons labeled with the terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling assay. Immunohistochemistry and western blotting demonstrated that lithium-elicited neuroprotection was associated with an increase in Bcl-2 protein levels. Our results raise the possibility that lithium may be considered as a neuroprotective agent in treatment of neurodegenerative diseases such as Huntington's disease.

Mas-related G-protein-coupled receptor subtype C (MrgC) may play an important role in pain sensation. However, the distribution of MrgC receptors in different subpopulations of rodent dorsal root ganglion (DRG) neurons has not been clearly demonstrated owing to a lack of MrgC-selective antibody. It is also unclear whether peripheral nerve injury induces different time-dependent changes in MrgC expression in injured and uninjured DRG neurons. Here we showed that MrgC immunoreactivity is distributed in both IB4-positive (non-peptidergic) and calcitonin gene-related peptide-positive (peptidergic) DRG neurons in mice and rats. Importantly, the MrgC mRNA level and MrgC immunoreactivity were both decreased in the injured L5 DRG compared to corresponding levels in the contralateral (uninjured) DRG in rats on days 14 and 30 after an L5 spinal nerve ligation. In contrast, mRNA and protein levels of MrgC were increased in the adjacent uninjured L4 DRG. Thus, nerve injury may induce temporal changes in MrgC expression that differ between injured and uninjured DRG neurons. In animal behavior tests, chronic constriction injury of the sciatic nerve induced mechanical pain hypersensitivity in wild-type mice and Mrg-clusterΔ(-/-) mice (Mrg KO). However, the duration of mechanical hypersensitivity was longer in the Mrg KO mice than in their wild-type littermates, indicating that activation of Mrgs may constitute an endogenous mechanism that inhibits the maintenance of neuropathic pain in mice. These findings extend our knowledge about the distribution of MrgC in rodent DRG neurons and the regulation of its expression by nerve injury.

Ginseng serves as a potential candidate for the treatment of aging-related memory decline or memory loss. However, the related mechanism is not fully understood. In this study, we applied an intraperitoneal injection of ginsenoside Rg1, an active compound from ginseng in middle-aged mice and detected memory improvement and the underlying mechanisms. Our results showed that a period of 30-day administration of ginsenoside Rg1 enhanced long-term memory in the middle-aged animals. Consistent with the memory improvement, ginsenoside Rg1 administration facilitated weak theta-burst stimulation (TBS)-induced long-term potentiation (LTP) in acute hippocampal slices from middle-aged animals. Ginsenoside Rg1 administration increased the dendritic apical spine numbers and area in the CA1 region. In addition, ginsenoside Rg1 administration up-regulated the expression of hippocampal p-AKT, brain-derived neurotrophic factor (BDNF), proBDNF and glutamate receptor 1 (GluR1), but not p-ERK. Interestingly, the phosphatase and tensin homolog deleted on chromosome ten (PTEN) inhibitor (bpV) mimicked the ginsenoside Rg1 effects, including increasing p-AKT expression, promoting hippocampal basal synaptic transmission, LTP and memory. Taken together, our data suggest that ginsenoside Rg1 treatment improves memory in middle-aged mice possibly through regulating the PI3K/AKT pathway, altering apical spines and facilitating hippocampal LTP.

Vascular dementia (VD), defined as a loss of memory and cognitive function resulting from vascular lesions in the brain, is the second-most-common cause of dementia in the elderly, after Alzheimer's disease. In recent years, research has focused on the pathogenesis of VD, and mitochondrial bioenergetic deficits have been suggested to contribute to VD onset. To further investigate the role of mitochondria in VD, we used a rat model of VD, which involved permanent bilateral occlusion of the common carotid arteries (with a 1-week interval between artery occlusion to avoid an abrupt reduction in cerebral blood flow) leading to chronic cerebral hypoperfusion. Prior to occlusion, male Wistar rats underwent 7 days of Morris water maze training. Only animals that could swim and passed the Morris water maze test were chosen for the study. After 5 days of Morris water maze training, mitochondria from the hippocampi of rats, which were randomly selected from animals that could complete the Morris water maze test, were isolated for functional assessment. Mitochondria isolated from the hippocampi of rats from the ischemia group had decreased pyruvate dehydrogenase protein levels, and increased oxidative stress, as manifested by increased hydrogen peroxide production. The ischemia group mitochondria also exhibited decreased respiration coupled to decreased expression and activity of the electron transport chain complex IV (cytochrome c oxidase). These results indicate that the mitochondrial oxidative metabolism is inhibited in the hippocampi of rats following chronic ischemia-induced VD. As the mitochondrial oxidative metabolism deficits, namely mitochondrial bioenergetic deficits directly affect the functions of neurons, it may contribute to VD onset.

Restrained eaters (REs) characterized by less efficient response inhibition are at risk for future onset of binge eating and bulimic pathology. Previous imaging studies investigating REs have been based on task-related functional magnetic resonance imaging (fMRI) and little is known about resting-state neural activity underlying restrained eating. To illuminate this issue, we investigated resting-state fMRI differences between REs (n=22) and unrestrained eaters (UREs) (n=30) using regional homogeneity (ReHo) analysis, which measures the temporal synchronization of spontaneous fluctuations. Samples were equated on body mass index (BMI) and caloric deprivation levels (i.e., 14±2.1h since last evening meal) before undergoing fMRI. Correlation analyses were performed between the ReHo index of identified regions and response inhibition based on stop-signal reaction time (SSRT) within each sample. Compared with UREs, REs showed more ReHo in brain regions associated with food reward (i.e., orbitofrontal cortex (OFC), dorsal-lateral prefrontal cortex (dlPFC)), attention (i.e., lingual gyrus, cuneus, inferior parietal lobule) and somatosensory functioning (i.e., paracentral lobule, anterior insula). In addition, ReHo values for the left dlPFC and left anterior insula, respectively, were negatively and positively correlated with SSRT among REs but not UREs. In concert with previous studies, these results suggest altered local synchronization may help to explain why dieting to maintain or lose weight often fails or increases risk for binge eating among REs.

Cocaine- and amphetamine-regulated transcript (CART) is a neuropeptide that plays neuroprotective roles in cerebral ischemia and reperfusion (I/R) injury in animal models or oxygen and glucose deprivation (OGD) in cultured neurons. Recent data suggest that intranasal CART treatment facilitates neuroregeneration in stroke brain. However, little is known about the effects of post-treatment with CART during the neuronal recovery after OGD and reoxygenation in cultured primary cortical neurons. The present study was to investigate the role of CART treated after OGD injury in neurons. Primary mouse cortical neurons were subjected to OGD and then treated with CART. Our data show that post-treatment with CART reduced the neuronal apoptosis caused by OGD injury. In addition, CART repaired OGD-impaired cortical neurons by increasing the expression of growth-associated protein 43 (GAP43), which promotes neurite outgrowth. This effect depends on pleiotrophin (PTN) as siRNA-mediated PTN knockdown totally abolished the increase in CART-stimulated GAP43 protein levels. In summary, our findings demonstrate that CART repairs the neuronal injury after OGD by facilitating neurite outgrowth through PTN-dependent pathway. The role for CART in neurite outgrowth makes it a new potential therapeutic agent for the treatment of neurodegenerative diseases.

Depression is a common symptom in Parkinson's disease (PD), but its pathophysiology remains unclear. Several lines of studies have revealed that the prelimbic (PrL) sub-region of the ventral medial prefrontal cortex and 5-HT1A receptors are involved in the regulation of depression. In this study, we examined whether complete unilateral lesions of the medial forebrain bundle (MFB) using 6-hydroxydopamine in rats are able to induce depressive-like behaviors, the role of PrL 5-HT1A receptors in the regulation of these behaviors, and co-localization of 5-HT1A receptor and neuronal glutamate transporter EAAC1-immunoreactive (EAAC1-ir) neurons in the PrL. The MFB lesions induced depressive-like responses as measured by the sucrose preference and forced swim tests when compared to sham-operated rats. The intra-PrL injection of 5HT1A receptor agonist 8-OH-DPAT (50, 100, and 500ng/rat) increased sucrose consumption, and decreased immobility time in both sham-operated and the lesioned rats, indicating the induction of antidepressant effects. Furthermore, the intra-PrL injection of 5HT1A receptor antagonist WAY-100635 (60, 120, and 240ng/rat) showed a decrease in sucrose consumption, and an increase in immobility time, indicating the induction of depressive-like responses. However, the effective doses in the lesioned rats were higher than those in sham-operated rats, which attribute to down-regulation of 5-HT1A receptor expression on EAAC1-ir neurons in the PrL of the lesioned rats. These findings suggest that unilateral lesions of the MFB in rats may induce depressive-like behaviors, and 5-HT1A receptors of the PrL play an important role in the regulation of these behaviors.

Neuronal differentiation is a critical developmental process that determines accurate synaptic connection and circuit wiring. A wide variety of naturally occurring compounds have been shown as promising drug leads for the generation and differentiation of neurons. Here we report that a diarylheptanoid from the plant Alpinia officinarum, 7-(4-hydroxyphenyl)-1-phenyl-4E-hepten-3-one (Cpd 1), exhibited potent activities in neuronal differentiation and neurite outgrowth. Cpd 1 induced differentiation of neuroblastoma Neuro-2a cells into a neuron-like morphology, and accelerated the establishment of axon-dendrite polarization of cultured hippocampal neurons. Moreover, Cpd 1 promoted neurite extension in both Neuro-2a cells and neurons. We showed that the effects of Cpd 1 on neuronal differentiation and neurite growth were specifically dependent on the activation of extracellular signal-regulated kinases (ERKs) and phosphoinositide 3-kinase (PI3K)-Akt signaling pathways. Importantly, intraperitoneal administration of Cpd 1 promoted the differentiation of new-born progenitor cells into mature neurons in the adult hippocampal dentate gyrus. Collectively, this study identifies a naturally occurring diarylheptanoid with beneficial effects on neuronal differentiation and neurite outgrowth in vitro and in vivo.

The elongation of neuron is highly dependent on membrane trafficking. Brefeldin A (BFA)-inhibited guanine nucleotide-exchange protein 1 (BIG1) functions in the membrane trafficking between the Golgi apparatus and the plasma membrane. BFA, an uncompetitive inhibitor of BIG1 can inhibit neurite outgrowth and polarity development. In this study, we aimed to define the possible role of BIG1 in neurite development and to further investigate the potential mechanism. By immunostaining, we found that BIG1 was extensively colocalized with synaptophysin, a marker for synaptic vesicles in soma and partly in neurites. The amount of both protein and mRNA of BIG1 were up-regulated during rat brain development. BIG1 depletion significantly decreased the neurite length and inhibited the phosphorylation of phosphatidylinositide 3-kinase (PI3K) and protein kinase B (AKT). Inhibition of BIG1 guanine nucleotide-exchange factor (GEF) activity by BFA or overexpression of the dominant-negative BIG1 reduced PI3K and AKT phosphorylation, indicating regulatory effects of BIG1 on PI3K-AKT signaling pathway is dependent on its GEF activity. BIG1 siRNA or BFA treatment also significantly reduced extracellular signal-regulated kinase (ERK) phosphorylation. Overexpression of wild-type BIG1 significantly increased ERK phosphorylation, but the dominant-negative BIG1 had no effect on ERK phosphorylation, indicating the involvement of BIG1 in ERK signaling regulation may not be dependent on its GEF activity. Our result identified a novel function of BIG1 in neurite development. The newly recognized function integrates the function of BIG1 in membrane trafficking with the activation of PI3K-AKT and ERK signaling pathways which are critical in neurite development.

Matrix metalloproteinases (MMPs) are a family of zinc-dependent endopeptidases that can be released or activated in a neuronal activity dependent manner. Although pathologically elevated levels of MMPs may be synaptotoxic, physiologically appropriate levels of MMPs may instead enhance synaptic transmission. MMP inhibitors can block long term potentiation (LTP), and at least one family member can affect an increase in the volume of dendritic spines. While the mechanism by which MMPs affect these changes is not completely understood, one possibility is that the cleavage of specific synaptic cell adhesion molecules plays a role. In the present study, we have examined the ability of neuronal activity to stimulate rapid MMP dependent shedding of the intercellular adhesion molecule-5 (ICAM-5), a synaptic adhesion molecule that is thought to inhibit the maturation and enlargement of dendritic spines. Since such cleavage would likely occur within minutes if it were relevant to a process such as LTP, we focused on post stimulus time points of 30 min or less. We show that NMDA can stimulate rapid shedding of ICAM-5 from cortical neurons in dissociated cell cultures and that such shedding is diminished by pretreatment of cultures with inhibitors that target MMP-3 and -9, proteases thought to influence synaptic plasticity. Additional studies suggest that MMP mediated cleavage of ICAM-5 occurs at amino acid 780, so that the major portion of the ectodomain is released. Since reductions in ICAM-5 have been linked to changes in dendritic spine morphology that are associated with LTP, we also examined the possibility that MMP dependent ICAM-5 shedding occurs following high frequency tetanic stimulation of murine hippocampal slices. Results show that the shedding of ICAM-5 occurs in association with LTP, and that both LTP and the associated ICAM-5 shedding are reduced when slices are pretreated with an MMP inhibitor. Together, these findings suggest that neuronal activity is linked to the shedding of a molecule that may inhibit dendritic spine enlargement and that MMPs can affect this change. While further studies will be necessary to determine the extent to which cleavage of ICAM-5 in particular contributes to MMP dependent LTP, our data support an emerging body of literature suggesting that MMPs are critical mediators of synaptic plasticity.

The present study sought to investigate if p53 mediates autophagy activation and mitochondria dysfunction in primary striatal neurons in kainic acid (KA)-induced excitotoxicity. The excitotoxic model of primary striatal neurons was established with KA. The levels of p53, microtubule-associated protein 1 light chain 3 (LC3), Beclin1, and p62 were examined by Western blot and immunostaining. Autophagy activation was also determined with electron microscope. To evaluate the contribution of p53 to autophagy activation and mitochondria dysfunction in KA-induced excitotoxicity, the protein levels of LC3, Beclin1, and p62, the mitochondrial transmembrane potential and the mitochondrial Reactive oxygen species (ROS) after pretreatment with the p53 inhibitor pifithrin-alpha (PFT-α) and the autophagy inhibitor 3-methyladenine (3-MA) were analyzed. Excitotoxic neuronal injury was induced after KA treatment as demonstrated by increases in lactate dehydrogenase (LDH) leakage and was significantly inhibited by PFT-α. Western blot and immunostaining showed that the induction of p53 protein occurred in the cytosol and the nucleus. Increases in autophagic proteins LC3 and Beclin1 were observed, whereas the protein levels of p62 decreased after KA treatment. Electron microscope analysis showed increased autophagosomes in the cytoplasm. The changes in LC3, Beclin1, and p62 levels were blocked by PFT-α, PFT-μ, 3-MA, and E64d but not Z-DEVD-FMK. JC-1 staining showed the depolarization of mitochondrial membrane potential after excitotoxic insult. Mito-tracker and RedoxSensor Red CC-1 staining showed an increased production of mitochondrial ROS after excitotoxic insult. These effects were significantly suppressed after pretreatment with PFT-α and 3-MA. This study suggests that p53 mediates KA-induced autophagy activation and mitochondrial dysfunction in striatal neurons.

The dyslexia-associated gene DCDC2 is a member of the DCX family of genes known to play roles in neurogenesis, neuronal migration, and differentiation. Here we report the first phenotypic analysis of a Dcdc2 knockout mouse. Comparisons between Dcdc2 knockout mice and wild-type (wt) littermates revealed no significant differences in neuronal migration, neocortical lamination, neuronal cilliogenesis or dendritic differentiation. Considering previous studies showing genetic interactions and potential functional redundancy among members of the DCX family, we tested whether decreasing Dcx expression by RNAi would differentially impair neurodevelopment in Dcdc2 knockouts and wild-type mice. Consistent with this hypothesis, we found that deficits in neuronal migration, and dendritic growth caused by RNAi of Dcx were more severe in Dcdc2 knockouts than in wild-type mice with the same transfection. These results indicate that Dcdc2 is not required for neurogenesis, neuronal migration or differentiation in mice, but may have partial functional redundancy with Dcx.

Epileptogenesis following traumatic brain injury (TBI) is likely due to a combination of increased excitability, disinhibition, and increased excitatory connectivity via aberrant axon sprouting. Targeting these pathways could be beneficial in the prevention and treatment of posttraumatic epilepsy. Here, we tested this possibility using the novel anticonvulsant (R)-N-benzyl 2-acetamido-3-methoxypropionamide ((R)-lacosamide [LCM]), which acts on both voltage-gated sodium channels and collapsin response mediator protein 2 (CRMP2), an axonal growth/guidance protein. LCM inhibited CRMP2-mediated neurite outgrowth, an effect phenocopied by CRMP2 knockdown. Mutation of LCM-binding sites in CRMP2 reduced the neurite inhibitory effect of LCM by ∼8-fold. LCM also reduced CRMP2-mediated tubulin polymerization. Thus, LCM selectively impairs CRMP2-mediated microtubule polymerization, which underlies its neurite outgrowth and branching. To determine whether LCM inhibits axon sprouting in vivo, LCM was injected into rats subjected to partial cortical isolation, an animal model of posttraumatic epileptogenesis that exhibits axon sprouting in cortical pyramidal neurons. Two weeks following injury, excitatory synaptic connectivity of cortical layer V pyramidal neurons was mapped using patch clamp recordings and laser scanning photostimulation of caged glutamate. In comparison with injured control animals, there was a significant decrease in the map size of excitatory synaptic connectivity in LCM-treated rats, suggesting that LCM treatment prevented enhanced excitatory synaptic connectivity due to posttraumatic axon sprouting. These findings suggest, for the first time, that LCM's mode of action involves interactions with CRMP2 to inhibit posttraumatic axon sprouting.

Embryonic knockdown of candidate dyslexia susceptibility gene (CDSG) homologs in cerebral cortical progenitor cells in the rat results in acute disturbances of neocortical migration. In the current report we investigated the effects of embryonic knockdown and overexpression of the homolog of DCDC2, one of the CDSGs, on the postnatal organization of the cerebral cortex. Using a within-litter design, we transfected cells in rat embryo neocortical ventricular zone around embryonic day (E) 15 with either 1) small hairpin RNA (shRNA) vectors targeting Dcdc2, 2) a DCDC2 overexpression construct, 3) Dcdc2 shRNA along with DCDC2 overexpression construct, 4) an overexpression construct composed of the C terminal domain of DCDC2, or 5) an overexpression construct composed of the DCX terminal domain of DCDC2. RNAi of Dcdc2 resulted in pockets of heterotopic neurons in the periventricular region. Approximately 25% of the transfected brains had hippocampal pyramidal cell migration anomalies. Dcdc2 shRNA-transfected neurons migrated in a bimodal pattern, with approximately 7% of the neurons migrating a short distance from the ventricular zone, and another 30% migrating past their expected lamina. Rats transfected with Dcdc2 shRNA along with the DCDC2 overexpression construct rescued the periventricular heterotopia phenotype, but did not affect the percentage of transfected neurons that migrate past their expected laminar location. There were no malformations associated with any of the overexpression constructs, nor was there a significant laminar disruption of migration. These results support the claim that knockdown of Dcdc2 expression results in neuronal migration disorders similar to those seen in the brains of dyslexics.

The transcription factor Otx1 is specifically expressed in layer V pyramidal cells (L5PCs) in the cerebral cortex. Otx1 null mutant mice have a defect in the developmental axon pruning of L5PCs and show epileptic seizures. However, the role of Otx1 in electrophysiology, morphology and synaptology of the cortical neurons has not been fully investigated. This study examines the influences of Otx1 on neuronal properties of L5PCs by loss- and gain-of-function approaches. Mice with an Otx1-null mutation had decreased structural measurements of basal dendrites in L5PCs. In contrast, the size of basal dendrites was increased in the Otx1-over-expressed pyramidal cells (PCs) in L2/3 where the gene normally does not express. PCs showed burst and non-burst firing patterns of action potentials. The proportion of burst firing neurons was reduced in the Otx1 mutant but increased in the neurons over-expressing Otx1. Although the burst firing population decreased, the proportion of those bursting neurons with a low threshold increased in the Otx1 mutant mice. Moreover, excitatory facilitating synaptic connections formed between L5PCs were predominant in the Otx1 mutant mice, which greatly contrasted with the predominant depressing synaptic connections in the controls. Taken together, it suggests an enhanced activity of neuronal network in the cortex of Otx1 mutant mice. These data indicate that the Otx1 expression is essential for the normal development of dendritic morphology, intrinsic electrophysiology and synaptic dynamics of L5PCs. This study provides new insights into molecular mechanisms underlying the spatial and temporal regulation of neuronal and synaptic properties of L5PCs, and improves our understanding on the generation of epileptic seizures.