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Neurons exhibit a high energetic need, and the question arises as how they metabolically adapt to changing activity states. This is relevant for interpreting functional neuroimaging in different brain areas. Particularly, neurons with a broad firing range might exhibit metabolic adaptations. Therefore, we studied MNTB (medial nucleus of the trapezoid body) principal neurons, which generate action potentials (APs) at frequencies up to several hundred hertz. We performed the experiments in acute brainstem slices of the Mongolian gerbil (Meriones unguiculatus) at 22.5-24.5°C. Upon electrical stimulation of afferent MNTB fibres with 400 stimuli at varying frequencies, we monitored autofluorescence levels of NAD(P)H and FAD and determined the extremum amplitudes of their biphasic response. Additionally, we compared these data with alterations in O 2 concentrations measured with an electrochemical sensor. These O 2 changes are prominent since MNTB neurons rely on oxidative phosphorylation as shown by our pharmacological experiments. We calculated the O 2 consumption rate as change in O 2 concentration divided by stimulus durations, because these periods varied inversely with stimulus frequency as a result of the constant number of 400 stimuli applied. The O 2 consumption rate increased with stimulation frequency up to a constant value at 600 Hz; that is, energy demand depends on temporal characteristics of activity despite the same number of stimuli. The rates showed no correlation with peak amplitudes of NAD(P)H or FAD, whilst the values of the two molecules were linearly correlated. This points at the complexity of analysing autofluorescence imaging for quantitative metabolic studies, because these values report only relative net changes of many superimposed oxidative and reductive processes. Monitoring O 2 concentration rates is, thus, an important tool to improve the interpretation of NAD(P)H/FAD autofluorescence data, as they do not under all conditions and in all systems appropriately reflect the metabolic activity or energy demand., (© 2024 The Authors. Journal of Neurochemistry published by John Wiley & Sons Ltd on behalf of International Society for Neurochemistry.)

Microglial cells (resident macrophages) feature rapid activation in CNS disease and can acquire multiple phenotypes exerting neuroprotection or neurotoxicity. The functional impact of surveying ("resting") microglia on neural excitability and neurotransmission in physiology is widely unknown, however. We addressed this issue in male rat hippocampal slice cultures (in situ) by pharmacological microglial ablation within days and by characterizing neuronal gamma-band oscillations (30-70 Hz) that are highly sensitive to neuromodulators and disturbances in ion and energy regulation. Gamma oscillations support action potential timing and synaptic plasticity, associate with higher brain functions like perception and memory, and require precise communication between excitatory pyramidal cells and inhibitory (GABAergic) interneurons. The slice cultures featured well-preserved hippocampal cytoarchitecture and parvalbumin-positive interneuron networks, microglia with ramified morphology, and low basal levels of IL-6, TNF-α, and nitric oxide (NO). Stimulation of slice cultures with the pro-inflammatory cytokine IFN-γ or bacterial LPS serving as positive controls for microglial reactivity induced MHC-II expression and increased cytokine and NO release. Chronic exposure of slice cultures to liposome-encapsulated clodronate reduced the microglial cell population by about 96%, whereas neuronal structures, astrocyte GFAP expression, and basal levels of cytokines and NO were unchanged. Notably, the properties of gamma oscillations reflecting frequency, number and synchronization of synapse activity were regular after microglial depletion. Also, electrical stimulus-induced transients of the extracellular potassium concentration ([K + ] o ) reflecting cellular K + efflux, clearance and buffering were unchanged. This suggests that nonreactive microglia are dispensable for neuronal homeostasis and neuromodulation underlying network signaling and rhythm generation in cortical tissue., (© 2020 The Authors. Journal of Neuroscience Research published by Wiley Periodicals LLC.)

The medial entorhinal cortex (MEC) plays a pivotal role in spatial processing together with hippocampal formation. The retrosplenial cortex (RSC) is also implicated in this process, and it is thus relevant to understand how these structures interact. This requires precise knowledge of their connectivity. Projections from neurons in RSC synapse onto principal neurons in layer V of MEC and some of these neurons send axons into superficial layers of MEC. Layer V of MEC is also the main target for hippocampal efferents from the subiculum and CA1 field. The aim of this study was to assess whether the population of cells targeted by RSC projections also receives input from the hippocampal formation and to compare the distribution of synaptic contacts on target dendrites. We labeled the cells in layer V of MEC by injecting a retrograde tracer into superficial layers. At the same time, we labeled RSC and subicular projections with different anterograde tracers. 3D-reconstruction of the labeled cells and axons revealed likely synaptic contacts between presynaptic boutons of both origins and postsynaptic MEC layer V basal dendrites. Moreover, these contacts overlapped on the same dendritic segments without targeting specific domains. Our results support the notion that MEC layer V neurons that project to the superficial layers receive convergent input from both RSC and subiculum. These data thus suggest that convergent subicular and RSC information contributes to the signal that neurons in superficial layers of EC send to the hippocampal formation. [ABSTRACT FROM AUTHOR]

The hippocampal formation is critically involved in learning and memory and contains a large proportion of neurons encoding aspects of the organism's spatial surroundings. In the medial entorhinal cortex (MEC), this includes grid cells with their distinctive hexagonal firing fields as well as a host of other functionally defined cell types including head direction cells, speed cells, border cells, and object-vector cells. Such spatial coding emerges from the processing of external inputs by local microcircuits. However, it remains unclear exactly how local microcircuits and their dynamics within the MEC contribute to spatial discharge patterns. In this review we focus on recent investigations of intrinsic MEC connectivity, which have started to describe and quantify both excitatory and inhibitory wiring in the superficial layers of the MEC. Although the picture is far from complete, it appears that these layers contain robust recurrent connectivity that could sustain the attractor dynamics posited to underlie grid pattern formation. These findings pave the way to a deeper understanding of the mechanisms underlying spatial navigation and memory. [ABSTRACT FROM AUTHOR]

Objective: A growing body of evidence indicates a potential role for the gut–brain axis as a novel therapeutic target in treating seizures. The present study sought to characterize the gut microbiome in Theiler murine encephalomyelitis virus (TMEV)‐induced seizures, and to evaluate the effect of microbial metabolite S‐equol on neuronal physiology as well as TMEV‐induced neuronal hyperexcitability ex vivo. Methods: We infected C57BL/6J mice with TMEV and monitored the development of acute behavioral seizures 0–7 days postinfection (dpi). Fecal samples were collected at 5–7 dpi and processed for 16S sequencing, and bioinformatics were performed with QIIME2. Finally, we conducted whole‐cell patch‐clamp recordings in cortical neurons to investigate the effect of exogenous S‐equol on cell intrinsic properties and neuronal hyperexcitability. Results: We demonstrated that gut microbiota diversity is significantly altered in TMEV‐infected mice at 5–7 dpi, exhibiting separation in beta diversity in TMEV‐infected mice dependent on seizure phenotype, and lower abundance of genus Allobaculum in TMEV‐infected mice regardless of seizure phenotype. In contrast, we identified specific loss of S‐equol‐producing genus Adlercreutzia as a microbial hallmark of seizure phenotype following TMEV infection. Electrophysiological recordings indicated that exogenous S‐equol alters cortical neuronal physiology. We found that entorhinal cortex neurons are hyperexcitable in TMEV‐infected mice, and exogenous application of microbial‐derived S‐equol ameliorated this TMEV‐induced hyperexcitability. Significance: Our study presents the first evidence of microbial‐derived metabolite S‐equol as a potential mechanism for alteration of TMEV‐induced neuronal excitability. These findings provide new insight for the novel role of S‐equol and the gut–brain axis in epilepsy treatment. [ABSTRACT FROM AUTHOR]

Songbirds are useful vertebrate study models for vocal learning and memory. The robust nucleus of the arcopallium (RA) receives synaptic inputs from both the posterior and anterior pathways of the song control system in songbirds. Hence, RA plays an important role in the control of singing. RA receives dopaminergic (DArgic) inputs that increase the excitability of RA projection neurons (PNs). However, the effects of DA on excitatory synaptic transmission are yet to be deciphered. In this study, the effects of DA on the excitatory synaptic transmission of the PNs in the RA of adult male zebra finches were investigated using a whole-cell patch-clamp recording. We observed that DA decreased the frequency of spontaneous excitatory postsynaptic currents (sEPSCs) and miniature excitatory postsynaptic currents (mEPSCs). The effects of DA were mimicked by the D1-like DA receptor (D1R) agonist, SKF-38393, but not the D2-like DA receptor (D2R) agonist, Quinpirole. Also, the effects of DA were blocked by D1R antagonist, SCH-23390, but not the D2R antagonist, Sulpiride. These results demonstrate that DA modulates excitatory synaptic transmission by acting on D1R in the RA of adult male zebra finches. [ABSTRACT FROM AUTHOR]

Cortical regions that are damaged by insults, such as ischemia, hypoxia, and trauma, frequently generate spreading depolarization (SD). At the neuronal level, SDs entail complete breakdown of ionic gradients, persisting for seconds to minutes. It is unclear whether these transient events have a more lasting influence on neuronal function. Here, we describe electrophysiological changes in cortical neurons after recovery from hypoxia-induced SD. When examined with standard measures of neuronal excitability several hours after recovery from SD, layer 5 pyramidal neurons in brain slices from mice of either sex appear surprisingly normal. However, we here introduce an additional parameter, dynamic gain, which characterizes the bandwidth of action potential encoding by a neuron, and thereby reflects its potential efficiency in a multineuronal circuit. We find that the ability of neurons that recover from SD to track high-frequency inputs is markedly curtailed; exposure to hypoxia did not have this effect when SD was prevented pharmacologically. Staining for Ankyrin G revealed at least a fourfold decrease in the number of intact axon initial segments in post-SD slices. Since this effect, along with the effect on encoding, was blocked by an inhibitor of the Ca2+-dependent enzyme, calpain, we conclude that both effects were mediated by the SD-induced rise in intracellular Ca2+. Although effects of calpain activation were detected in the axon initial segment, changes in soma-dendritic compartments may also be involved. Whatever the precise molecular mechanism, our findings indicate that in the context of cortical circuit function, effectiveness of neurons that survive SD may be limited. [ABSTRACT FROM AUTHOR]

Abstract: Activation of neuronal nicotinic acetylcholine receptors (nAChRs) by nicotine is reported to protect brain neurons from glutamate excitotoxicity. We inquired whether a similar phenomenon can occur in the rat isolated spinal cord (or spinal slice culture) challenged by a transient (1 hr) application of kainate (a powerful glutamate receptor agonist) to induce excitotoxicity mimicking spinal injury in vitro. We recorded spinal reflexes and fictive locomotion generated by the locomotor central pattern generator before and 24 hr after applying kainate. We also monitored network activity with Ca2+ imaging and counted neurons and glia with immunohistochemical methods. In control conditions, nicotine (1 μM; 4 hr) depressed reflexes and fictive locomotion with slow recovery and no apparent neurotoxicity at 24 hr although synchronous Ca2+ transients appeared in slice cultures. Kainate nearly halved neuron numbers (while sparing glia), decreased reflexes and Ca2+ transients, and suppressed fictive locomotion. When nicotine was applied (4 hr) after washout of kainate, fictive locomotor cycles appeared 24 hr later though with low periodicity, and significant protection of neurons, including motoneurons, was observed. Nicotine applied together with kainate and maintained for further 4 hr yielded better neuroprotection, improved fictive locomotion expression and reversed the depression of Ca2+ transients. nAChR antagonists did not intensify kainate neurotoxicity and inhibited the neuroprotective effects of nicotine. These data suggest that nicotine was efficacious to limit histological and functional excitotoxic damage probably because it activated and then desensitized nAChRs on excitatory and inhibitory network neurons to prevent triggering intracellular cell death pathways. [ABSTRACT FROM AUTHOR]

Calsenilin modulates A-type potassium channels, regulates presenilin-mediated -secretase activity, and represses prodynorphin and c-fos genes expression. RhoA is involved in various cellular functions including proliferation, differentiation, migration, transcription, and regulation of the actin cytoskeleton. Although recent studies demonstrate that calsenilin can directly interact with RhoA and that RhoA inactivation is essential for neuritogenesis, it is uncertain whether there is a link between calsenilin and RhoA-regulated neuritogenesis. Here, we investigated the role of calsenilin in RhoA-regulated neuritogenesis using in vitro and in vivo systems. We found that calsenilin induced RhoA inactivation, which accompanied RhoA phosphorylation and the reduced phosphorylation levels of LIM kinase (LIMK) and cofilin. Interestingly, PC12 cells overexpressing either full-length (FL) or the caspase 3-derived C-terminal fragment (CTF) of calsenilin significantly inactivated RhoA through its interaction with RhoA and p190 Rho GTPase-activating protein (p190RhoGAP). In addition, cells expressing FL and the CTF of calsenilin had increased neurite outgrowth compared to cells expressing the N-terminal fragment (NTF) of calsenilin or vector alone. Moreover, Tat-C3 and Y27632 treatment significantly increased the percentage of neurite-bearing cells, neurite length, and the number of neurites in cells. Finally, calsenilin deficiency in the brains of calsenilin-knockout mice significantly interfered with RhoA inactivation. These findings suggest that calsenilin contributes to neuritogenesis through RhoA inactivation. [ABSTRACT FROM AUTHOR]

Objective: The contribution of recurring seizures to the progression of epileptogenesis is debated. Seizure-induced brain damage is not conclusively demonstrated either in humans or in animal models of epilepsy. We evaluated the expression of brain injury biomarkers on postsurgical brain tissue obtained from 20 patients with frequent seizures and a long history of drug-resistant focal epilepsy.Methods: The expression patterns of specific glial, neuronal, and inflammatory molecules were evaluated by immunohistochemistry in the core of type II focal cortical dysplasias (FCD-II), at the FCD boundary (perilesion), and in the adjacent normal-appearing area included in the epileptogenic region. We also analyzed surgical specimens from cryptogenic patients not presenting structural alterations at imaging.Results: Astroglial and microglial activation, reduced neuronal density, perivascular CD3-positive T-lymphocyte clustering, and fibrinogen extravasation were demonstrated in the core of FCD-II lesions. No pathological immunoreactivity was observed outside the FCD-II or in cryptogenetic specimens, where the occurrence of interictal and ictal epileptiform activity was confirmed by either stereo-electroencephalography or intraoperative electrocorticography.Interpretation: Recurrent seizures do not induce the expression of brain damage markers in nonlesional epileptogenic cortex studied in postsurgical tissue from cryptogenic and FCD patients. This evidence argues against the hypothesis that epileptiform activity per se contributes to focal brain injury, at least in the neocortical epilepsies considered here. Ann Neurol 2017;82:331-341. [ABSTRACT FROM AUTHOR]

A number of mutations in α4β2-containing (α4β2*) nicotinic acetylcholine (ACh) receptors (nAChRs) are linked to autosomal dominant nocturnal frontal lobe epilepsy (ADNFLE), including one in the β2 subunit called β2V287L. Two α4β2* subtypes with different subunit stoichiometries and ACh sensitivities co-exist in the brain, a high-sensitivity subtype with (α4)2(β2)3 subunit stoichiometry and a low-sensitivity subtype with (α4)3(β2)2 stoichiometry. The α5 nicotinic subunit also co-assembles with α4β2 to form a high-sensitivity α5α4β2 nAChR. Previous studies suggest that the β2V287L mutation suppresses low-sensitivity α4β2* nAChR expression in a knock-in mouse model and also that α5 co-expression improves the surface expression of ADNFLE mutant nAChRs in a cell line. To test these hypotheses further, we expressed mutant and wild-type (WT) nAChRs in oocytes and mammalian cell lines, and measured the effects of the β2V287L mutation on surface receptor expression and the ACh response using electrophysiology, a voltage-sensitive fluorescent dye, and superecliptic pHluorin (SEP). The β2V287L mutation reduced the EC50 values of high- and low-sensitivity α4β2 nAChRs expressed in Xenopus oocytes for ACh by a similar factor and suppressed low-sensitivity α4β2 expression. In contrast, it did not affect the EC50 of α5α4β2 nAChRs for ACh. Measurements of the ACh responses of WT and mutant nAChRs expressed in mammalian cell lines using a voltage-sensitive fluorescent dye and whole-cell patch-clamping confirm the oocyte data. They also show that, despite reducing the maximum response, β2V287L increased the α4β2 response to a sub-saturating ACh concentration (1 μM). Finally, imaging SEP-tagged α5, α4, β2, and β2V287L subunits showed that β2V287L reduced total α4β2 nAChR surface expression, increased the number of β2 subunits per α4β2 receptor, and increased surface α5α4β2 nAChR expression. Thus, the β2V287L mutation alters the subunit composition and sensitivity of α4β2 nAChRs, and increases α5α4β2 surface expression. [ABSTRACT FROM AUTHOR]

Neuroglial interactions are now recognized as essential to brain functions. Extensive research has sought to understand the modalities of such dialog by focusing on astrocytes, the most abundant glial cell type of the central nervous system. Neuron-astrocyte exchanges occur at multiple levels, at different cellular locations. With regard to information processing, regulations occurring around synapses are of particular interest as synaptic networks are thought to underlie higher brain functions. Astrocytes morphology is tremendously complex in that their processes exceedingly branch out to eventually form multitudinous fine leaflets. The latter extremities have been shown to surround many synapses, forming perisynaptic astrocytic processes, which although recognized as essential to synaptic functioning, are poorly defined elements due to their tiny size. The current review sums up the current knowledge on their molecular and structural properties as well as the functional characteristics making them good candidates for information processing units. [ABSTRACT FROM AUTHOR]

The sensitivity of neurons from the honey bee olfactory system to pyrethroid insecticides was studied using the patch-clamp technique on central ‘antennal lobe neurons’ (ALNs) in cell culture. In these neurons, the voltage-dependent sodium currents are characterized by negative potential for activation, fast kinetics of activation and inactivation, and the presence of cumulative inactivation during train of depolarizations. Perfusion of pyrethroids on these ALN neurons submitted to repetitive stimulations induced (1) an acceleration of cumulative inactivation, and (2) a marked slowing of the tail current recorded upon repolarization. Cypermethrin and permethrin accelerated cumulative inactivation of the sodium current peak in a similar manner and tetramethrin was even more effective. The slow-down of channel deactivation was markedly dependent on the type of pyrethroid. With cypermethrin, a progressive increase of the tail current amplitude along with successive stimulations reveals a traditionally described use-dependent recruitment of modified sodium channels. However, an unexpected decrease in this tail current was revealed with tetramethrin. If one considers the calculated percentage of modified channels as an index of pyrethroids effects, ALNs are significantly more susceptible to tetramethrin than to permethrin or cypermethrin for a single depolarization, but this difference attenuates with repetitive activity. Further comparison with peripheral neurons from antennae suggest that these modifications are neuron type specific. Modeling the sodium channel as a multi-state channel with fast and slow inactivation allows to underline the effects of pyrethroids on a set of rate constants connecting open and inactivated conformations, and give some insights to their specificity. Altogether, our results revealed a differential sensitivity of central olfactory neurons to pyrethroids that emphasize the ability for these compounds to impair detection and processing of information at several levels of the bees olfactory pathway. [ABSTRACT FROM AUTHOR]

Gangliosides are widely expressed sialylated glycosphingolipids with multifunctional properties in different cell types and organs. In the nervous system, they are highly enriched in both glial and neuronal membranes. Mice lacking complex gangliosides attributable to targeted ablation of the B4galnt1 gene that encodes β-1,4-N-acetylegalactosaminyltransferase 1 (GalNAc-transferase; GalNAcT-/-) develop normally before exhibiting an age-dependent neurodegenerative phenotype characterized by marked behavioral abnormalities, central and peripheral axonal degeneration, reduced myelin volume, and loss of axo-glial junction integrity. The cell biological substrates underlying this neurodegeneration and the relative contribution of either glial or neuronal gangliosides to the process are unknown. To address this, we generated neuron-specific and glial-specific GalNAcT rescue mice crossed on the global GalNAcT-/- background [GalNAcT-/--Tg(neuronal) and GalNAcT-/--Tg(glial)] and analyzed their behavioral, morphological, and electrophysiological phenotype. Complex gangliosides, as assessed by thin-layer chromatography, mass spectrometry, GalNAcT enzyme activity, and anti-ganglioside antibody (AgAb) immunohistology, were restored in both neuronal and glial GalNAcT rescue mice. Behaviorally, GalNAcT-/--Tg(neuronal) retained a normal "wild-type" (WT) phenotype throughout life, whereas GalNAcT-/--Tg(glial) resembled GalNAcT-/- mice, exhibiting progressive tremor, weakness, and ataxia with aging. Quantitative electron microscopy demonstrated that GalNAcT-/- and GalNAcT-/--Tg(glial) nerves had significantly increased rates of axon degeneration and reduced myelin volume, whereas GalNAcT-/--Tg(neuronal) and WT appeared normal. The increased invasion of the paranode with juxtaparanodal Kv1.1, characteristically seen in GalNAcT-/- and attributed to a breakdown of the axo-glial junction, was normalized in GalNAcT-/--Tg(neuronal) but remained present in GalNAcT-/--Tg(glial) mice. These results indicate that neuronal rather than glial gangliosides are critical to the age-related maintenance of nervous system integrity. [ABSTRACT FROM AUTHOR]

The subiculum, a para-hippocampal structure positioned between the cornu ammonis 1 subfield and the entorhinal cortex, has been implicated in temporal lobe epilepsy in human patients and in animal models of epilepsy. The structure is characterized by the presence of a significant population of burst firing neurons that has been shown previously to lead epileptiform activity locally. Phase transitions in epileptiform activity in neurons following a prolonged challenge with an epileptogenic stimulus has been shown in other brain structures, but not in the subiculum. Considering the importance of the subicular burst firing neurons in the propagation of epileptiform activity to the entorhinal cortex, we have explored the phenomenon of phase transitions in the burst firing neurons of the subiculum in an in vitro rat brain slice model of epileptogenesis. Whole-cell patch-clamp and extracellular field recordings revealed a distinct phenomenon in the subiculum wherein an early hyperexcitable state was followed by a late suppressed state upon continuous perfusion with epileptogenic 4-aminopyridine and magnesium-free medium. The suppressed state was characterized by inhibitory post-synaptic potentials in pyramidal excitatory neurons and bursting activity in local fast-spiking interneurons at a frequency of 0.1-0.8 Hz. The inhibitory post-synaptic potentials were mediated by GABAA receptors that coincided with excitatory synaptic inputs to attenuate action potential discharge. These inhibitory post-synaptic potentials ceased following a cut between the cornu ammonis 1 and subiculum. The suppression of epileptiform activity in the subiculum thus represents a homeostatic response towards the induced hyperexcitability. Our results suggest the importance of feedforward inhibition in exerting this homeostatic control. [ABSTRACT FROM AUTHOR]

The neural cell adhesion molecule ( NCAM) is involved in neural development and in plasticity in the adult brain. NCAM140 and NCAM180 isoforms are transmembrane proteins with cytoplasmic domains that differ only in an alternatively spliced exon in the NCAM180 isoform. Both isoforms can interact with several extracellular and cytoplasmic molecules mediating NCAM-dependent functions. Most identified intracellular interaction partners bind to both isoforms, NCAM140 and NCAM180. To identify further intracellular interaction partners specifically binding to NCAM180 the cytosolic domain of human NCAM180 was recombinantly expressed and applied onto a protein macroarray containing the protein library from human fetal brain. We identified the ubiquitin C-terminal hydrolase ( UCHL1) which has been described as a de-ubiquitinating enzyme as a potential interaction partner of NCAM180. Since NCAM180 and NCAM140 are ubiquitinated, NCAM140 was included in the subsequent experiments. A partial colocalization of both NCAM isoforms and UCHL1 was observed in primary neurons and the B35 neuroblastoma cell line. Overexpression of UCHL1 significantly decreased constitutive ubiquitination of NCAM180 and NCAM140 whereas inhibition of endogenous UCHL1 increased NCAM's ubiquitination. Furthermore, lysosomal localization of NCAM180 and NCAM140 was significantly reduced after overexpression of UCHL1 consistent with a partial colocalization of internalized NCAM with UCHL1. These data indicate that UCHL1 is a novel interaction partner of both NCAM isoforms that regulates their ubiquitination and intracellular trafficking. Structured digital abstract NCAM1 binds to UCHL1 by protein array (View interaction), Uchl1 and NCAM180 colocalize by fluorescence microscopy (View interaction), Uchl1 and NCAM1 colocalize by fluorescence microscopy (View interaction), Uchl1 and NCAM140 colocalize by fluorescence microscopy (View interaction) [ABSTRACT FROM AUTHOR]

Voltage-gated sodium channels (VGSC) are the primary mediators of electrical signal amplification and propagation in excitable cells. VGSC subtypes are diverse, with different biophysical and pharmacological properties, and varied tissue distribution. Altered VGSC expression and/or increased VGSC activity in sensory neurons is characteristic of inflammatory and neuropathic pain states. Therefore, VGSC modulators could be used in prospective analgesic compounds. VGSCs have specific binding sites for four conotoxin families: μ-, μO-, δ- and í-conotoxins. Various studies have identified that the binding site of these peptide toxins is restricted to well-defined areas or domains. To date, only the μ- and μO-family exhibit analgesic properties in animal pain models. This review will focus on conotoxins from the μ- and μO-families that act on neuronal VGSCs. Examples of how these conotoxins target various pharmacologically important neuronal ion channels, as well as potential problems with the development of drugs from conotoxins, will be discussed. [ABSTRACT FROM AUTHOR]

The amygdala is the central component of a functional brain system regulating fear and emotional behaviors. Studies of the ontogeny of fear behaviors reveal the emergence of distinct fear responses at different postnatal ages. Here, we performed a stereological analysis of the rat amygdala to characterize the cellular changes underlying its normal structural development. Distinct amygdala nuclei exhibited different patterns of postnatal development, which were largely similar to those we have previously shown in monkeys. The combined volume of the lateral, basal, and accessory basal nuclei increased by 113% from 1 to 3 weeks of age and by an additional 33% by 7 months of age. The volume of the central nucleus increased only 37% from 1 to 2 weeks of age and 38% from 2 weeks to 7 months. At 1 week of age, the medial nucleus was 77% of the 7-month-old's volume and exhibited a constant, marginal increase until 7 months. Neuron number did not differ in the amygdala from 1 week to 7 months of age. In contrast, astrocyte number decreased from 3 weeks to 2 months of age in the whole amygdala. Oligodendrocyte number increased in all amygdala nuclei from 3 weeks to 7 months of age. Our findings revealed that distinct amygdala nuclei exhibit different developmental profiles and that the rat amygdala is not fully mature for an extended period postnatally. We identified different periods of postnatal development of distinct amygdala nuclei and cellular components, which are concomitant with the ontogeny of different fear and emotional behaviors. J. Comp. Neurol. 520:3745-3763, 2012. © 2012 Wiley Periodicals, Inc. [ABSTRACT FROM AUTHOR]

The blood-brain barrier (BBB) is located within a unique anatomic interface and has functional ramifications to most of the brain and blood cells. In the past, the BBB was considered a pharmacokinetic impediment to antiepileptic drug penetration into the brain; nowadays it is becoming increasingly evident that targeting of the damaged or dysfunctional BBB may represent a therapeutic approach to reduce seizure burden. Several studies have investigated the mechanisms linking the onset and sustainment of seizures to BBB dysfunction. These studies have shown that the BBB is at the crossroad of a multifactorial pathophysiologic process that involves changes in brain milieu, altered neuroglial physiology, development of brain inflammation, leukocyte-endothelial interactions, faulty angiogenesis, and hemodynamic changes leading to energy mismatch. A number of knowledge gaps, conflicting points of view, and discordance between clinical and experimental data currently characterize this field of neuroscience. As more pieces are added to this puzzle, it is apparent that each mechanism needs to be validated in an appropriate clinical context. We now offer a BBB-centric view of seizure disorders, linking several aspects of seizures and epilepsy physiopathology to BBB dysfunction. We have reviewed the therapeutic, antiseizure effect of drugs that promote BBB repair. We also present BBB neuroimaging as a tool to correlate BBB restoration to seizure mitigation. Add-on cerebrovascular drug could be of efficacy in reducing seizure burden when used in association with neuronal antiepileptic drugs. [ABSTRACT FROM AUTHOR]

The entorhinal cortex (ERC) is critically implicated in temporal lobe epileptogenesis—the most common type of adult epilepsy. Previous studies have suggested that epileptiform discharges likely initiate in seizure-sensitive deep layers (V–VI) of the medial entorhinal area (MEA) and propagate into seizure-resistant superficial layers (II–III) and hippocampus, establishing a lamina-specific distinction between activities of deep- versus superficial-layer neurons and their seizure susceptibilities. While layer II stellate cells in MEA have been shown to be hyperexcitable and hypersynchronous in patients and animal models of temporal lobe epilepsy (TLE), the fate of neurons in the deep layers under epileptic conditions and their overall contribution to epileptogenicity of this region have remained unclear. We used whole cell recordings from slices of the ERC in normal and pilocarpine-treated epileptic rats to characterize the electrophysiological properties of neurons in this region and directly assess changes in their excitatory and inhibitory synaptic drive under epileptic conditions. We found a surprising heterogeneity with at least three major types and two subtypes of functionally distinct excitatory neurons. However, contrary to expectation, none of the major neuron types characterized showed any significant changes in their excitability, barring loss of excitatory and inhibitory inputs in a subtype of neurons whose dendrite extended into layer III, where neurons are preferentially lost during TLE. We confirmed hyperexcitability of layer II neurons in the same slices, suggesting minimal influence of deep-layer input on superficial-layer neuron excitability under epileptic conditions. These data show that deep layers of ERC contain a more diverse population of excitatory neurons than previously envisaged that appear to belie their seizure-sensitive reputation. [ABSTRACT FROM AUTHOR]

Huntington's disease (HD) is a neurodegenerative disease where GABAergic medium spiny neurons (MSNs) in the striatum degenerate. Embryonic stem cell-derived neural transplantation may provide an appropriate therapy for HD. Here we aimed to develop a suitable protocol to obtain a high percentage of functional GABAergic neurons from mouse embryonic stem cells (mESCs), and then tested their differentiation potential in vivo. The monolayer method was compared with the embryoid body and five stage method for its efficiency in generating GABAergic neurons from mESCs. All three methods yielded a similar percentage of GABAergic neurons from mESCs. Monolayer method-derived GABAergic neurons expressed the MSN marker dopamine- and cyclic AMP-regulated phosphoprotein (DARPP32). The pluripotent stem cell population could be eliminated in vitro by treating cells with puromycin and retinoic acid. Using patch-clamp recordings, the functional properties of GABAergic neurons derived from mESCs were compared to GABAergic neurons derived from primary lateral ganglionic eminence. Both types of neurons showed active membrane properties (voltage-gated Na and K currents, Na-dependent action potentials, and spontaneous postsynaptic currents) and possessed functional glutamatergic receptors and transporters. mESC-derived neural progenitors were transplanted into a mouse model of HD. Grafted cells differentiated to mature neurons expressing glutamate decarboxylase, dopamine type 1 receptors, and DARPP32. Also, neural precursors and dividing populations were found in the grafts. In summary, mESCs are able to differentiate efficiently into functional GABAergic neurons using defined in vitro conditions, and these survive and differentiate following grafting to a mouse model of HD. [ABSTRACT FROM AUTHOR]

Purpose: Despite the consistent observation that γ-aminobutyric acid A (GABAA) receptors mediate excitatory responses at perinatal stages, the role of the GABAergic system in the generation of neonatal epileptiform activity remains controversial. Therefore, we analyzed whether tonic and phasic GABAergic transmission had differential effects on neuronal excitability during early development. Methods: We performed whole cell patch-clamp and field potential recordings in the CA3 region of hippocampal slices from immature (postnatal day 4-7) rats to analyze the effect of specific antagonists and modulators of tonic and phasic GABAergic components on neuronal excitability. Key Findings: The GABAergic antagonists gabazine (3 μ m) and picrotoxin (100 μ m) induced epileptiform discharges, whereas activation of GABAA receptors attenuated epileptiform discharges. Under low-Mg2+ conditions, 100 n m gabazine and 1 μ m picrotoxin were sufficient to provoke epileptiform activity in 63.2% (n = 19) and 53.8% (n = 26) of the slices, respectively. Whole-cell patch-clamp experiments revealed that these concentrations significantly reduced the amplitude of phasic GABAergic postsynaptic currents but had no effect on tonic currents. In contrast, 1-μ m 4,5,6,7-tetrahydroisoxaz-olo[5,4-c]-pyridin-3-ol (THIP) induced a tonic current of −12 ± 2.5 pA (n = 6) and provoked epileptiform discharges in 57% (n = 21) of the slices. Significance: We conclude from these results that in the early postnatal rat hippocampus a constant phasic synaptic activity is required to control excitability and prevent epileptiform activity, whereas tonic GABAergic currents can mediate excitatory responses. Pharmacologic intervention at comparable human developmental stages should consider these ambivalent GABAergic actions. [ABSTRACT FROM AUTHOR]

Background: After focal neuronal injury the endocannabinioid system becomes activated and protects or harms neurons depending on cannabinoid derivates and receptor subtypes. Endocannabinoids (eCBs) play a central role in controlling local responses and influencing neural plasticity and survival. However, little is known about the functional relevance of eCBs in long-range projection damage as observed in stroke or spinal cord injury (SCI). Methods: In rat organotypic entorhino-hippocampal slice cultures (OHSC) as a relevant and suitable model for investigating projection fibers in the CNS we performed perforant pathway transection (PPT) and subsequently analyzed the spatial and temporal dynamics of eCB levels. This approach allows proper distinction of responses in originating neurons (entorhinal cortex), areas of deafferentiation/anterograde axonal degeneration (dentate gyrus) and putative changes in more distant but synaptically connected subfields (cornu ammonis (CA) 1 region). Results: Using LC-MS/MS, we measured a strong increase in arachidonoylethanolamide (AEA), oleoylethanolamide (OEA) and palmitoylethanolamide (PEA) levels in the denervation zone (dentate gyrus) 24 hours post lesion (hpl), whereas entorhinal cortex and CA1 region exhibited little if any changes. NAPE-PLD, responsible for biosynthesis of eCBs, was increased early, whereas FAAH, a catabolizing enzyme, was up-regulated 48hpl. Conclusion: Neuronal damage as assessed by transection of long-range projections apparently provides a strong timedependent and area-confined signal for de novo synthesis of eCB, presumably to restrict neuronal damage. The present data underlines the importance of activation of the eCB system in CNS pathologies and identifies a novel site-specific intrinsic regulation of eCBs after long-range projection damage. [ABSTRACT FROM AUTHOR]

Amyloid beta (Aβ) peptide accumulation in the brains of patients with Alzheimer's disease (AD) is closely associated with increased nerve cell death. However, many cells survive and it is important to understand the mechanisms involved in this survival response. Recent studies have shown that an anti-apoptotic mechanism in cancer cells is mediated by aerobic glycolysis, also known as the Warburg effect. One of the major regulators of aerobic glycolysis is pyruvate dehydrogenase kinase (PDK), an enzyme which represses mitochondrial respiration and forces the cell to rely heavily on glycolysis, even in the presence of oxygen. Recent neuroimaging studies have shown that the spatial distribution of aerobic glycolysis in the brains of AD patients strongly correlates with Aβ deposition. Interestingly, clonal nerve cell lines selected for resistance to Aβ exhibit increased glycolysis as a result of activation of the transcription factor hypoxia inducible factor 1. Here we show that Aβ resistant nerve cell lines upregulate Warburg effect enzymes in a manner reminiscent of cancer cells. In particular, Aβ resistant nerve cell lines showed elevated PDK1 expression in addition to an increase in lactate dehydrogenase A (LDHA) activity and lactate production when compared to control cells. In addition, mitochondrial derived reactive oxygen species (ROS) were markedly diminished in resistant but not sensitive cells. Chemically or genetically inhibiting LDHA or PDK1 resensitized resistant cells to Aβ toxicity. These findings suggest that the Warburg effect may contribute to apoptoticresistance mechanisms in the surviving neurons of the AD brain. Loss of the adaptive advantage afforded by aerobic glycolysis may exacerbate the pathophysiological processes associated with AD. [ABSTRACT FROM AUTHOR]

During the past decade experimental evidence has accumulated demonstrating that the electrical communication between neurons through gap junctions (GJs) is a necessary neural mechanism underlying oscillations and synchrony. Here we extended our earlier observations concerning the involvement of GJs in hippocampal theta production. Using trimethylamine, a GJ opener, we demonstrated a reversible increase in theta amplitude and power and an increase in the duration of theta epochs. This effect was accompanied by a decrease in the percentage of recorded theta-off cells, an increase in the percentage of recorded theta-on phasic cells, and an increase in the number of rhythmic cell discharges per theta wave. We suggest that all these findings result from an enhanced level of interneuronal excitation, mediated by an increase in the efficacy of local GJ coupling. [ABSTRACT FROM AUTHOR]

Seizures in focal epilepsies are sustained by a highly synchronous neuronal discharge that arises at restricted brain sites and subsequently spreads to large portions of the brain. Despite intense experimental research in this field, the earlier cellular events that initiate and sustain a focal seizure are still not well defined. Their identification is central to understand the pathophysiology of focal epilepsies and to develop new pharmacological therapies for drug-resistant forms of epilepsy. The prominent involvement of astrocytes in ictogenesis was recently proposed. We test here whether a cooperation between astrocytes and neurons is a prerequisite to support ictal (seizure-like) and interictal epileptiform events. Simultaneous patchclamp recording and Ca2+ imaging techniques were performed in a new in vitro model of focal seizures induced by local applications of N-methyl-D-aspartic acid (NMDA) in rat entorhinal cortex slices. We found that a Ca2+ elevation in astrocytes correlates with both the initial development and the maintenance of a focal, seizure-like discharge. A delayed astrocyte activation during ictal discharges was also observed in other models (including the whole in vitro isolated guinea pig brain) in which the site of generation of seizure activity cannot be precisely monitored. In contrast, interictal discharges were not associated with Ca2+ changes in astrocytes. Selective inhibition or stimulation of astrocyte Ca2+ signalling blocked or enhanced, respectively, ictal discharges, but did not affect interictal discharge generation. Our data reveal that neurons engage astrocytes in a recurrent excitatory loop (possibly involving gliotransmission) that promotes seizure ignition and sustains the ictal discharge. This neuron-astrocyte interaction may represent a novel target to develop effective therapeutic strategies to control seizures. [ABSTRACT FROM AUTHOR]

Astrocytes are specialized glial cells that outnumber neurons by over fivefold. They contiguously tile the entire central nervous system (CNS) and exert many essential complex functions in the healthy CNS. Astrocytes respond to all forms of CNS insults through a process referred to as reactive astrogliosis, which has become a pathological hallmark of CNS structural lesions. Substantial progress has been made recently in determining functions and mechanisms of reactive astrogliosis and in identifying roles of astrocytes in CNS disorders and pathologies. A vast molecular arsenal at the disposal of reactive astrocytes is being defined. Transgenic mouse models are dissecting specific aspects of reactive astrocytosis and glial scar formation in vivo. Astrocyte involvement in specific clinicopathological entities is being defined. It is now clear that reactive astrogliosis is not a simple all-or-none phenomenon but is a finely gradated continuum of changes that occur in context-dependent manners regulated by specific signaling events. These changes range from reversible alterations in gene expression and cell hypertrophy with preservation of cellular domains and tissue structure, to long-lasting scar formation with rearrangement of tissue structure. Increasing evidence points towards the potential of reactive astrogliosis to play either primary or contributing roles in CNS disorders via loss of normal astrocyte functions or gain of abnormal effects. This article reviews (1) astrocyte functions in healthy CNS, (2) mechanisms and functions of reactive astrogliosis and glial scar formation, and (3) ways in which reactive astrocytes may cause or contribute to specific CNS disorders and lesions . [ABSTRACT FROM AUTHOR]

Ongoing learning continuously shapes the distribution of neurons’ synaptic weights in a system with plastic synapses. Plasticity may change the weights of synapses that were active during the induction—homosynaptic changes, but also may change synapses not active during the induction—heterosynaptic changes. Here we will argue, that heterosynaptic and homosynaptic plasticity are complementary processes, and that heterosynaptic plasticity might accompany homosynaptic plasticity induced by typical pairing protocols. Synapses are not uniform in their susceptibility for plastic changes, but have predispositions to undergo potentiation or depression, or not to change. Predisposition is one of the factors determining the direction and magnitude of homo- and heterosynaptic changes. Heterosynaptic changes which take place according to predispositions for plasticity may provide a useful mechanism(s) for homeostasis of neurons’ synaptic weights and extending the lifetime of memory traces during ongoing learning in neuronal networks. [ABSTRACT FROM AUTHOR]

The present study, based on neurohistological techniques (Nissl-staining, Golgi-impregnation), focuses on the cytoarchitecture of the corticoid complex in the strawberry finch, Estrilda amandava. This complex in birds occupies the dorsolateral surface of the telencephalic pallium and remains subdivided into an intermediate corticoid area (CI) and a dorsolateral corticoid area (CDL). The CDL in the strawberry finch is a thin superficial part of the caudal pallium adjoining the medially situated hippocampal formation, whereas the CI is demarcated between the CDL and the parahippocampal area of telencephalon. Neurons of the corticoid complex are classified into three main cell groups: predominant projection neurons, local circuit neurons and stellate neurons. The spinous projection neurons send out distant projecting axons that typically extend several varicose collaterals. Most of these collaterals lie parallel to the ventricle. These neurons are subclassified into pyramidal neurons (localized only in the CI) and multipolar neurons (present in both the CI and CDL). The CDL also possesses small and medium-sized horizontal cells, which are bitufted or multipolar with smooth, moderately branching dendrites. The aspinous local circuit neurons extend short axons that ramify locally. Stellate neurons have sparse spinous dendrites and locally arborizing axons. The corticoid complex of birds corresponds to the lateral cerebral cortex of lizards and to the entorhinal cortex of mammals on the basis of neuronal morphology and bidirectional connections between adjacent areas. [ABSTRACT FROM AUTHOR]

Proteins that are released from cells consist of those in the extracellular matrix, as well as extracellular signaling and adhesion molecules. The majority of these extracellular proteins are, however, unknown. To determine their identity, we have used a proteomics approach to define proteins released from neurons, astrocytes and neural precursor cells. Using two-dimensional gels and liquid chromatography/mass spectrometry technology, it is shown that while astrocytes release a relatively small number of proteins, neurons and neuronal precursor cells release a larger number of proteins with more functional diversity. Although there is overlap between the different cell types, the exact composition of the extracellular protein pool is unique for each cell population. The various subsets of extracellular neural proteins include those involved in cellular Redox regulation and chaperones. In addition, many proteolytic enzymes are found outside of the cell. These data show that the extracellular space within the nervous system has a more diverse protein composition than previously thought. [ABSTRACT FROM AUTHOR]

Mature stellate cells of the rat medial entorhinal cortex (EC), layer II, exhibit subthreshold membrane potential oscillations (MPOs) at theta frequencies (4–12 Hz) in vitro. We find that MPOs appear between postnatal days 14 (P14) and 18 (P18) but show little further change by day 28+ (P28–P32). To identify the factors responsible, we examined the electrical responses of developing stellate cells, paying attention to two currents thought necessary for mature oscillation: the h current Ih, which provides the slow rectification required for resonance; and a persistent sodium current INaP, which provides amplification of resonance. Responses to injected current revealed that P14 cells were often nonresonant with a relatively high resistance. Densities of Ihand INaPboth rose by about 50% from P14 to P18. However, Ihlevels fell to intermediate values by P28+. Given the nonrobust trend in Ihexpression and a previously demonstrated potency of even low levels of Ihto sustain oscillation, we propose that resonance and MPOs are limited at P14 more by low levels of INaPthan of Ih. The relative importance of INaPfor the development of MPOs is supported by simulations of a conductance-based model, which also suggest that general shunt conductance may influence the precise age when MPOs appear. In addition to our physiological study, we analyzed spine densities at P14, P18, and P28+ and found a vigorous synaptogenesis across the whole period. Our data predict that functions that rely on theta rhythmicity in the hippocampal network are limited until at least P18. [ABSTRACT FROM AUTHOR]

Fas ligand (FasL)-receptor system plays an essential role in regulating cell death in the developing nervous system, and it has been implicated in neurodegenerative and inflammatory responses in the CNS. Lifeguard (LFG) is a protein highly expressed in the hippocampus and the cerebellum, and it shows a particularly interesting regulation by being up-regulated during postnatal development and in the adult. We show that over-expression of LFG protected cortical neurons from FasL-induced apoptosis and decreased caspase-activation. Reduction of endogenous LFG expression by small interfering RNA sensitized cerebellar granular neurons to FasL-induced cell death and caspase-8 activation, and also increased sensitivity of cortical neurons. In differentiated cerebellar granular neurons, protection from FasL-induced cell death could be attributed exclusively to LFG and appears to be independent of FLICE inhibitor protein. Thus, LFG is an endogenous inhibitor of FasL-mediated neuronal death and it mediates the FasL resistance of CNS differentiated neurons. Finally, we also demonstrate that LFG is detected in lipid rafts microdomains, where it may interact with Fas receptor and regulate FasL-activated signaling pathways. [ABSTRACT FROM AUTHOR]

The parasubiculum is a major component of the hippocampal formation that receives inputs from the CA1 region, anterior thalamus, and medial septum and that projects primarily to layer II of the entorhinal cortex. Hippocampal theta-frequency (4–12 Hz) electroencephalographic (EEG) activity has been correlated with sensorimotor integration, spatial navigation, and memory functions. The present study was aimed at determining if theta is also generated locally within the parasubiculum versus volume conducted from adjacent structures. In urethan-anesthetized rats, the phase-reversal of theta activity between superficial and deep layers of the parasubiculum was demonstrated using differential recordings from movable bipolar electrodes that eliminate the influence of volume-conducted activity. Parasubicular theta was abolished by atropine, and was in phase with theta in stratum radiatum/lacunosum-moleculare of the CA1 region. Whole cell current-clamp recordings in brain slices were then used to determine if parasubicular theta may be generated in part by membrane potential oscillations in layer II neurons. Membrane potential oscillations occurred in most layer II neurons, including four putative interneurons, when cells were held at near-threshold voltages using current injection. The frequency of oscillations increased from 3.2 to 6.1 Hz when bath temperature was raised from 22 to 32°C, and oscillations persisted in the presence of blockers of fast ionotropic glutamatergic and GABAergic synaptic transmission. Oscillations are therefore likely generated by intrinsic, voltage-dependent ionic conductances. These results indicate that theta field activity is generated locally within the parasubiculum and that intrinsic membrane potential oscillations, synchronized by local inhibitory inputs, may contribute to the generation of this activity. [ABSTRACT FROM AUTHOR]

Neuronal differentiation in PC12 cells induced by nerve growth factor (NGF) requires sustained activation of ERK/MAP kinase pathway (Raf–MEK–ERK cascade). Although classical Ras (H-Ras, K-Ras, and N-Ras) activated by NGF signaling induces activation of ERK pathway, the activation is transient and not sufficient for PC12 cell differentiation. Instead, it has been widely accepted that NGF signaling-mediated Rap1 activation causes sustained activation of ERK pathway. There has been no direct evidence, however, that Rap1 participates in neuronal differentiation. Here we show that NGF signaling induces sustained activation of M-Ras and subsequent sustained activation of ERK pathway and the transcription factor CREB leading to PC12 cell differentiation. Exogenously expressed constitutively active mutant of M-Ras caused neurite outgrowth in PC12 cells and activating phosphorylation of ERK, whereas activated Rap1 did not. Knockdown of endogenous M-Ras by small interfering RNAs as well as the expression of a dominant–negative mutant of M-Ras interfered with NGF-induced neuritogenesis. Since MEK inhibitors prevented M-Ras-induced neurite outgrowth, ERK pathway participates in this differentiation pathway. Furthermore, M-Ras brought about ERK pathway-mediated activating phosphorylation of CREB and the CREB-mediated transcription. In addition, a dominant–negative mutant of CREB inhibited M-Ras-induced neuritogenesis. Taken together, NGF-induced PC12 cell differentiation requires M-Ras–ERK pathway-mediated activation of CREB. M-Ras was predominantly expressed in the hippocampus and cerebellum of mouse brain and in the gray matter of the spinal cord. All these properties of M-Ras were apparently indistinguishable from those of H-Ras. However, NGF stimulation caused transient activation of classical Ras proteins but sustained activation of M-Ras as well as sustained activating phosphorylation of ERK and CREB. Therefore, M-Ras is essential for neuronal differentiation in PC12 cells by inducing sustained activation of ERK pathway. [ABSTRACT FROM AUTHOR]

Nitric oxide (NO) is thought to be involved in the central mechanism of hyperalgesia and allodynia at the spinal level. Recently, we reported that NO played an important role in the induction of long-term potentiation (LTP) of synaptic strength in spinal dorsal horn, which is believed to underlie hyperalgesia and allodynia. In this study, to elucidate the relationship of NO to LTP in spinal dorsal horn, we measured the spatiotemporal distribution of NO signal with the NO-sensitive dye, DAR-4M, and neuronal excitation with the voltage-sensitive dye, RH482, in rat spinal cord slices, elicited by dorsal root stimulation. In superficial dorsal horn, neuronal excitation evoked by C fiber-activating dorsal root stimulation was potentiated for more than 2 h after low-frequency conditioning stimulation (LFS, 240 pulses at 2 Hz for 2 min). In the same slices that exhibited LTP, NO was produced and distributed in the superficial dorsal horn during the delivery of LFS, and the amplitude of LTP and amount of NO production showed close correlation from slice to slice. LTP and production of NO were inhibited in the presence of the NO synthase inhibitors and an inhibitor of heme oxygenase, the synthetic enzyme for carbon monoxide (CO). These results suggest that production and distribution of NO is necessary for the induction of LTP in spinal dorsal horn, and that CO contributes to the LTP induction and NO production by LFS. [ABSTRACT FROM AUTHOR]

The entorhinal cortex (EC) is a key brain area controlling both hippocampal input and output via neurones in layer II and layer V, respectively. It is also a pivotal area in the generation and propagation of epilepsies involving the temporal lobe. We have previously shown that within the network of the EC, neurones in layer V are subject to powerful synaptic excitation but weak inhibition, whereas the reverse is true in layer II. The deep layers are also highly susceptible to acutely provoked epileptogenesis. Considerable evidence now points to a role of spontaneous background synaptic activity in control of neuronal, and hence network, excitability. In the present article we describe results of studies where we have compared background release of the excitatory transmitter, glutamate, and the inhibitory transmitter, GABA, in the two layers, the role of this background release in the balance of excitability, and its control by presynaptic auto- and heteroreceptors on presynaptic terminals. [ABSTRACT FROM AUTHOR]

The medial septal diagonal band area (MS/DB), made up of GABAergic and cholinergic neurones, plays an essential role in the generation and modulation of the hippocampal theta rhythm. To understand the part that the cholinergic neurones might play in this activity, we sought to determine whether postsynaptic nicotinic receptor responses can be detected in slices of the rodent MS/DB by puffing on acetylcholine (ACh). Neurones were characterized electrophysiologically into GABAergic and cholinergic neurones according to previous criteria. Responses of the MS/SB neurones to ACh were various combinations of fast depolarizations (1.5–2.5 s), fast hyperpolarizations (3–4 s) and slow depolarizations (20–30 s), the latter two being blocked by atropine. The fast depolarizations were partially or not blocked with cadmium and low calcium, tetrodotoxin, and antagonists of other ionotropic receptors, and were antagonized with 25μmmecamylamine. Pharmacological investigation of the responses showed that theα7* nicotinic receptor type is associated with cholinergic neurones and 10% of the GABAergic neurones, and that nonα7* nicotinic receptor subtypes are associated with 50% of the GABAergic neurones. Pharmacological dissection of evoked and spontaneous postsynaptic responses, however, did not provide evidence for synaptic nicotinic receptor transmission in the MS/DB. It was concluded that nicotinic receptors, although prevalent on the somatic and/or dendritic membrane compartments of neurones in the MS/DB, are on extrasynaptic sites where they presumably play a neuromodulatory role. The presence ofα7* nicotinic receptors on cholinergic neurones may also render these cells specifically vulnerable to degeneration in Alzheimer's disease. [ABSTRACT FROM AUTHOR]

Cholinergic activation of entorhinal cortex (EC) layer V neurons plays a crucial role in the medial temporal lobe memory system and in the pathophysiology of temporal lobe epilepsy. Here, we demonstrate that muscarinic activation by focal application of carbachol depolarizes EC layer V neurons and induces epileptiform activity in rat brain slices. These seizure-like bursts are associated with a somatic [Ca2+]i increase of 293 ± 82 n m and are blocked by the glutamate receptor antagonists CNQX and APV. Muscarinic activation did not directly evoke a [Ca2+]i increase, but subthreshold and suprathreshold depolarization did. Functional axon mapping revealed local axon branching as well as axon collaterals ascending to layers II and III. During blockade of ionotropic glutamatergic AMPA and NMDA receptors, carbachol depolarized layer V neurons by +7.5 ± 3.4 mV. This direct muscarinic depolarization was associated with a conductance increase of 35 ± 10.3% (+4.3 ± 1.25 nS). Intracellular buffering of [Ca2+]i changes did not block this depolarization, but prolonged action potential duration and reduced adaptation of action potential firing. The muscarinic depolarization was neither blocked by combining intracellular Ca2+-buffering (EGTA or BAPTA) with non-specific Ca2+-channel inhibition by Ni+ (1 m m), nor by Ba2+ (1 m m) nor during inhibition of the h-current by 2 m m Cs+. In whole-cell patch-clamp recording, reversal of the muscarinic current occurred at about −45 mV and −5 mV with complete substitution of intrapipette K+ with Cs+. Thus, muscarinic depolarization of EC layer V neurons appears to be primarily mediated by Ca2+-independent activation of non-specific cation channels that conduct K+ about three times as well as Na+. [ABSTRACT FROM AUTHOR]

To elucidate the cellular action of tiagabine, an inhibitor of GAT-1 GABA transporter, in the globus pallidus, whole-cell patch-clamp recordings were made from rat globus pallidus neurons in the acutely prepared brain slice. Superfusion of tiagabine significantly prolonged the decay kinetics of both action potential-dependent and -independent (tetrodotoxin-resistant) inhibitory postsynaptic currents (IPSCs) that were mediated by GABAA receptors. Furthermore, it decreased the frequency of these IPSCs. The latter effect was reversed by the GABAB receptor antagonist CGP55845, which alone had no effect, suggesting the involvement of presynaptic GABAB receptors. Thus, tiagabine could inhibit or disinhibit globus pallidus neurons by increasing the activation of the GABAA receptors and presynaptic GABAB receptors, respectively. In the behaving animal, tiagabine when injected unilaterally into the globus pallidus caused consistent ipsilateral rotation of the rats indicative of increased inhibition of globus pallidus activity. This finding could be explained by the proposition that in the presence of tiagabine, prolonged action of GABA on GABA receptors would dominate over the inhibitory effect of tiagabine on GABA release. Our findings on the electrophysiological and behavioral effects of tiagabine in globus pallidus suggest that this basal ganglia nucleus is one of the sites of action of tiagabine and provides a rationale for investigating its involvement in epilepsy. [ABSTRACT FROM AUTHOR]

Acetylcholine (ACh) is an important neurotransmitter of the CNS that binds both nicotinic and muscarinic receptors to exert its action. However, the mechanisms underlying the effects of cholinergic receptors have still not been completely elucidated. Central cholinergic neurons, mainly located in basal forebrain, send their projections to different structures including the cortex. The cortical innervation is diffuse and roughly topographic, which has prompted some authors to suspect a modulating role of ACh on the activity of the cortical network rather than a direct synaptic role. The cholinergic system is implicated in functional, behavioural and pathological states including cognitive function, nicotine addiction, Alzheimer's disease, Tourette's syndrome, epilepsies and schizophrenia. As these processes depend on the activation of glutamatergic and GABAergic systems, the cholinergic terminals must exert their effects via the modulation of excitatory and/or inhibitory neurotransmission. However, the understanding of cholinergic modulation is complex because it is the result of a mixture of positive and negative modulation, implying that there are various types, or even subtypes, of cholinergic receptors. In this review, we summarize the current knowledge on central cholinergic systems (projections and receptors) and then aim to focus on the implications for ACh in the modulation of cortical neuronal activity. [ABSTRACT FROM AUTHOR]

Provides information on neural cell lines derived from stem cells. Fundamental features of neural and pluripotent cell lines; Organization of cell lines; Information on cell lines derived by oncogene expression; Explanations on neurospheres and related cell lines.

Transcranial magnetic stimulation (TMS) was used to study visuospatial attention processing in ten healthy volunteers. In a forced choice recognition task the subjects were confronted with two symbols simultaneously presented during 120 ms at random positions, one in the left and the other in the right visual field. The subject had to identify the presented pattern out of four possible combinations and to press the corresponding response key within 2 s. Double-pulse TMS (dTMS) with a 100-ms interstimulus interval (ISI) and an intensity of 80% of the stimulator output (corresponding to 110–120% of the motor threshold) was applied by a non-focal coil over the right or left posterior parietal cortex (PPC, corresponding to P3/P4 of the international 10-20 system) at different time intervals after onset of the visual stimulus (starting at 120 ms, 270 ms and 520 ms). Double-pulse TMS over the right PPC starting at 270 ms led to a significant increase in percentage of errors in the contralateral, left visual field (median: 23% with TMS vs 13% without TMS, P=0.0025). TMS applied earlier or later showed no effect. Furthermore, no significant increase in contra- or ipsilateral percentage of errors was found when the left parietal cortex was stimulated with the same timing. These data indicate that: (1) parietal influence on visuospatial attention is mainly controlled by the right lobe since the same stimulation over the left parietal cortex had no significant effect, and (2) there is a vulnerable time window to disturb this cortical process, since dTMS had a significant effect on the percentage of errors in the contralateral visual hemifield only when applied 270 ms after visual stimulus presentation. [ABSTRACT FROM AUTHOR]

Abstract Neuronal fibres of the hippocampal formation of normal and chronic epileptic rats were investigated by fluorescent tracing methods using the pilocarpine model of limbic epilepsy. Two months after onset of spontaneous limbic seizures, hippocampal slices were prepared and maintained in vitro for 10 h. Small crystals of fluorescent dye [fluorescein (fluoro-emerald®) and tetramethylrhodamine (fluoro-ruby®)] were applied to different hippocampal regions. The main findings were: (i) in control rats there was no supragranular labelling when the mossy fibre tract was stained in stratum radiatum of area CA3. However, in epileptic rats a fibre network in the inner molecular layer of the dentate gyrus was retrogradely labelled; (ii) a retrograde innervation of area CA3 by CA1 pyramidal cells was disclosed by labelling remote CA1 neurons after dye injection into the stratum radiatum of area CA3 in chronic epileptic rats; (iii) labelling of CA1 neurons apart from the injection site within area CA1 was observed in epileptic rats but not in control animals; and (iv), a subicular-hippocampal projection was present in pilocarpine-treated rats when the tracer was injected just below the stratum pyramidale of area CA1. The findings show that fibre rearrangement in distinct regions of the epileptic hippocampal formation can occur as an aftermath of pilocarpine-induced status epilepticus. [ABSTRACT FROM AUTHOR]

Abstract: The mRNA encoding the human α5 nicotinic subunit was detected in several structures of the nervous system but appeared to be mainly expressed in cerebellum, thalamus, and the autonomic ganglia. For the first time, the α5 transcript was also detected in several non-neuronal tissues, with maximal expressions being found throughout the gastrointestinal tract, thymus, and testis. Many other extraneuronal sites expressed α5, but there were also nonexpressing organs, such as the liver, spleen, and kidney. To understand the transcriptional mechanisms controlling such a diversified expression of α5 in neuronal and nonneuronal cells, we isolated the 5′-regulatory region of the human gene and characterized its properties. Here we identify the α5 core promoter and demonstrate that the DNA regions surrounding it contain elements (with positive or negative activities) that work in a tissue-specific fashion. In particular, the segment specifying the 5′-untranslated region in neuronal cells has most of the properties of an enhancer because it activates a heterologous promoter in a position- and orientation-independent fashion. We therefore conclude that the expression of α5 relies on a highly complex promoter that uses distinct regulatory elements to comply with the different functional and developmental requirements of the various tissues and organs. [ABSTRACT FROM AUTHOR]

Acetylcholine-induced membrane currents and excitatory postsynaptic currents (EPSCs) were recorded from the neurons of rat superior cervical ganglion (SCG) using the whole-cell patch clamp and the two-electrode voltage clamp techniques, correspondingly. The EPSC decay was bi-exponential, with fast and slow components characterized by time constants 5.5 +/- 0.5 msec and 20.4 +/- 1.2 msec (mean +/- SEM; n = 23), respectively. Blocking of these currents by a series of newly synthesized bis-cationic ammonium compounds, the pentamethonium and pentaethonium derivatives, was analyzed. Blocking effects were due to a block of nicotinic acetylcholine receptor (nAChR) open channel, with mean blocker binding rate constants for the fast component three to five times higher than those for the slow component. Dimensions of a nAChR ionic channel were deduced from a relationship between blocking activity of the compounds and the size of the projections of their three-dimensional molecular models on the neuronal membrane plane. The results suggest that there are two populations of nAChRs in rat SCG neurons; while these polulations differ in the rate constants of the binding by the blocker to their open channels, they exhibit similar channel diameter, 11.8 A, at the level at which the blockers bind to the channel. [ABSTRACT FROM AUTHOR]

Although exosomes were first described in reticulocytes in 1983, many people do not realize that similar vesicles had been studied in the context of muscle and nerve, beginning in 1980. At the time of their discovery, these vesicles were named adherons, and they were found to play an important role in both cell–substrate and cell–cell adhesion. My laboratory described several molecules that are present in adherons, including heparan sulfate proteoglycans (HSPGs) and purpurin. HSPGs have since been shown to play a variety of key roles in brain physiology. Purpurin has a number of important functions in the retina, including a role in nerve cell differentiation and regeneration. In this review, I discuss the discovery of adherons and how that led to continuing studies on their role in the brain with a particular focus on HSPGs. [ABSTRACT FROM AUTHOR]

Abstract Tonic impulse flow in the septohippocampal GABAergic pathway is essential for normal cognitive functioning and is sustained, in part, by acetylcholine (ACh) that is released locally via axon collaterals of septohippocampal cholinergic neurons. Septohippocampal cholinergic neurons degenerate in Alzheimer's disease and other neurodegenerative disorders. While the importance of the muscarinic effects of ACh on septohippocampal GABAergic neurons is well recognized, the nicotinic effects of ACh remain unstudied despite the reported benefits of nicotine on cognitive functioning. In the present study, using electrophysiological recordings in a rat brain slice preparation, rapid applications of nicotine excited 90% of retrogradely labelled septohippocampal GABA-type neurons with an EC[sub 50] of 17 µm and increased the frequency of spontaneously occurring, impulse-dependent fast GABAergic and glutamatergic synaptic currents via the α4β2-nicotinic receptor. Interestingly, tetrodotoxin blocked all effects of nicotine on septohippocampal GABAergic type neurons, suggesting involvement of indirect mechanisms. We demonstrate that the effects of nicotine on septohippocampal GABA-type neurons involve recruitment of a novel, local glutamatergic circuitry as (i) Group I metabotropic glutamatergic receptor antagonists reduced the effects of nicotine; (ii) the number of nicotine responsive neurons was significantly reduced in recordings from slices that had been trimmed so as to reduce the number of glutamate-containing neurons within the slice preparation; (iii) in light and ultrastructural double immunocytochemical labelling studies vesicular glutamate 2 transporter immunoreactive terminals made synaptic contacts with parvalbumin-immunoreactive septohippocampal GABAergic neurons. The discovery of a local glutamatergic circuit within the septum may provide another avenue for restoring septohippocampal GABAergic functions in neurodegenerative disorders associated with a loss of septohippocampal cholinergic neurons. [ABSTRACT FROM AUTHOR]

Abstract Medial entorhinal cortex (EC) deep layer neurons projecting to the dentate gyrus (DG) were studied. Neurons, retrogradely-labelled with rhodamine-dextran-amine were characterized electrophysiologically with the patch clamp technique and finally labelled with biocytin. Pyramidal and nonpyramidal neurons form projections from the deep layers of the EC to the molecular layer of the DG. In addition, both classes of projection neurons send ascending axon collaterals to the superficial layers of the EC. Both classes of neurons were characterized physiologically by regular action potential firing upon depolarizing current injection. While a substantial number of pyramidal projection cells showed intrinsic membrane potential oscillations, none of the studied nonpyramidal cells exhibited oscillations. Despite the morphological similarity of bipolar and multipolar cells to those of GABAergic interneurons in the EC, their electrophysiological characteristics were similar to those of principal neurons and immunocytochemistry for GABA was negative. We conclude, that neurons of the deep layers of the medial EC projecting to the DG may function as both local circuit and projecting neurons thereby contributing to synchronization between deep layers of the EC, superficial layers of the EC and the DG. [ABSTRACT FROM AUTHOR]