Vascular endothelial growth factor (VEGF) regulates angiogenesis, but also has important, yet poorly characterized roles in neuronal wiring. Using several genetic and in vitro approaches, we discovered a novel role for VEGF in the control of cerebellar granule cell (GC) migration from the external granule cell layer (EGL) toward the Purkinje cell layer (PCL). GCs express the VEGF receptor Flk1, and are chemoattracted by VEGF, whose levels are higher in the PCL than EGL. Lowering VEGF levels in mice in vivo or ectopic VEGF expression in the EGL ex vivo perturbs GC migration. Using GC-specific Flk1 knock-out mice, we provide for the first time in vivo evidence for a direct chemoattractive effect of VEGF on neurons via Flk1 signaling. Finally, using knock-in mice expressing single VEGF isoforms, we show that pericellular deposition of matrix-bound VEGF isoforms around PC dendrites is necessary for proper GC migration in vivo. These findings identify a previously unknown role for VEGF in neuronal migration.

Phosphorylation of CaMKII and AMPA receptor GluA1 subunit has been shown to play a major role in hippocampal-dependent long-term/reference memory (RM) and in the expression of long-term synaptic potentiation (LTP). In contrast, it has been proposed that dephosphorylation of these proteins could be involved in the opposite phenomenon of hippocampal long-term synaptic depression (LTD) and in adaptive forgetting. Adaptive forgetting allows interfering old memories to be forgotten to give new ones the opportunity to be stored in memory, and in particular in short-term/working memory (WM) that was shown to be very sensitive to proactive interference. To determine the role of CaMKII and GluA1 in adaptive forgetting, we adopted a comparative approach to assess the relative quantity and phosphorylation state of these proteins in the brain of rats trained in one of three radial maze paradigms: a RM task, a WM task involving a high level of adaptive forgetting, or a WM involving a low level of adaptive forgetting. Surprisingly, Western blot analyses revealed that training in a WM task involving a high level of adaptive forgetting specifically increased the expression of AMPA receptor GluA1 subunit and the activity of CaMKII in the dentate gyrus. These results highlight that WM with proactive interference involves mechanisms of synaptic plasticity selectively in the dentate gyrus.

Several lines of evidence have suggested that decreases in postsynaptic inhibition may have a role in epileptogenesis in cortical structures. However, other studies have suggested that GABAergic inhibition is spared, or even augmented in some forms of post-lesional epilepsy. In the studies described here, inhibitory events were recorded in two models of post-lesional chronic epileptogenesis. (i) As previously reported (D.A. Prince and G.-F. Tseng. J. Neurophysiol. 69: 1276-1291. 1993), epileptiform activity develops in slices from partially isolated rat neocortical islands 2-3 weeks after the initial in vivo lesion. In this model of post-traumatic epilepsy, large amplitude polyphasic inhibitory postsynaptic currents (IPSCs) in layer V pyramidal neurons are associated with each interictal epileptiform field potential. The frequency of spontaneous IPSCs as well as miniature IPSCs was significantly increased in neocortical slices from the epileptogenic chronically injured cortex versus controls. Immunocytochemical reactions for parvalbumin and calbindin, calcium binding proteins present in subgroups of GABAergic neurons, showed an increased staining of both neuropil and somata within the epileptogenic tissue. Immunoreactivity for glutamic acid decarboxylase (GAD) and GABA also appeared to be increased in the neuropil. (ii) Cortical microgyri resembling human malformations were produced by freeze lesions made transcranially in P0 rat cortex (K.M. Jacobs, M.J. Gutnick, and D.A. Prince. Cereb. Cortex, 6: 514-523. 1996). The boundary between the four-layered microgyrus and surrounding cortex become epileptogenic within about 2 weeks, as judged by evoked extracellular field potentials and cellular activities. Epileptogenesis in the surrounding cortex is not altered when the microgyrus itself is isolated by transcortical cuts. Patch-clamp recordings from layer V neurons in the epileptogenic zone showed that spontaneous IPSCs are larger and more dependent on glutamatergic synapses than in control neurons. The amplitudes of polysynaptic IPSCs evoked by threshold stimulation were also larger than in control cells. Although evaluation of inhibitory events in these models is still incomplete, results to date suggest that GABAergic inhibition may be enhanced in epileptogenic areas associated with chronic cortical injury. Sprouting of axonal arborizations of pyramidal cells onto interneurons, upregulation of GABAergic neurons, and perhaps sprouting of inhibitory axons that make increased numbers of contacts onto pyramidal cells may all contribute to the increased inhibitory drive. Results in these models do not support the disinhibitory hypothesis of chronic epileptogenesis.

The types of sodium channels that are expressed by neurons shape the rising phase of action potentials and influence patterns of action potential discharge. With regard to the enteric nervous system (ENS), there is uncertainty about which channels are expressed, and in particular it is unknown whether Na(v)1.7 is present. We designed specific probes for the guinea pig Na(v)1.7 alpha subunit as well as for the other tetrodotoxin (TTX)-sensitive alpha subunits (Na(v)1.1, Na(v)1.2, Na(v)1.3, and Na(v)1.6) in order to perform in situ hybridization (ISH) histochemistry on guinea pig myenteric ganglia. We established that only Na(v)1.7 mRNA and Na(v)1.3 mRNA are expressed in these ganglia. The ISH signal for Na(v)1.7 transcripts was found in seemingly all the myenteric neurons. The expression of the Na(v)1.3 alpha subunit was confirmed by immunohistochemistry in a large proportion (62%) of the myenteric neuron population. This population included enteric sensory neurons. Na(v)1.6 immunoreactivity, absent from myenteric neurons, was detected in glial cells only when a high anti-Na(v)1.6 antibody concentration was used. This suggests that the Na(v)1.6 alpha subunit and mRNA are present only at low levels, which is consistent with the fact that no Na(v)1.6 mRNA could be detected in the ENS by ISH. The fact that adult myenteric neurons are endowed with only two TTX-sensitive alpha subunits, namely, Na(v)1.3 and Na(v)1.7, emphasizes the singularity of the ENS. Both these subunits, known to have slow-inactivation kinetics, are well adapted for generating action potentials from slow excitatory postsynaptic potentials, a mode of synaptic transmission that applies to all ENS neuron types.

Local field potential (LFP) recording is a very useful electrophysiological method to study brain processes. However, this method is criticized for recording low frequency activity in a large area of extracellular space potentially contaminated by distal activity. Here, we theoretically and experimentally compare ground-referenced (RR) with differential recordings (DR). We analyze electrical activity in the rat cortex with these two methods. Compared with RR, DR reveals the importance of local phasic oscillatory activities and their coherence between cortical areas. Finally, we show that DR provides a more faithful assessment of functional connectivity caused by an increase in the signal to noise ratio, and of the delay in the propagation of information between two cortical structures. Introduction 1 LFP recording of cortical structures constitutes a powerful tool to detect functional 2 signatures of cognitive processes. However, several studies have suggested that 3 recording methods suffer of major caveats due to the recording of activity in distant 4 neural populations [1-4]. Thus, theta oscillations (6-10Hz) during active wake seem to 5 propagate from the hippocampus to the frontal cortical areas [5]. Despite these 6 important studies, LFP recording has revealed important features of cortical 7 organizations [6, 7]. For example, cortical slow wave oscillations of NREM sleep, which 8 constitute a prominent feature of this vigilance state, contribute moderately to 9 coherence between cortical areas [7]. In contrast, weak slow wave oscillations during 10 active wake contribute to a relatively high level of coherence between cortical 11 areas [6, 7]. LFPs are mainly generated by post-synaptic response to pre-synaptic 12 activity of neurons [8-11] and constitutes a natural integrator of action potentials 13coming from a given cortical region [12][13][14]. In its usual description, LFP recording 14 appears to be less local than multi-unit activity recordings. Indeed, the usual 15 PLOS 1/15 recording mode of LFP consists in implanting a single electrode in the investigated 16 cortical region and a second one in a supposed neutral site. This simple recording 17 configuration, called monopolar or referential recording (RR) mode, is well adapted to 18 evaluate a global brain state. Unlike single and multi-unit probe, the impedance of the 19 standard electrode used for LFP recording is usually low in order to record neural 20 activity of a larger area. However, this method may detect activities from distant 21 cortical areas located between the recording and the reference electrode [1, 13-19], a 22phenomenon called volume conduction. We propose here to compare monopolar or RR 23 mode to bipolar or differential recording (DR), which consists in setting a pair of 24 electrodes in the same cortical area and measuring the voltage difference between 25 them. The main historical reasons why RR is widely used [7,20] are: 1) its simplicity 26 because of the low number of wires that needs to be implanted (contributing to the 27 ...

Neuromyelitis Optica (NMO) is an autoimmune demyelinating disease of the central nervous system characterized by the presence of autoantibodies (called NMO-IgG) targeting aquaporin-4. Aquaporin-4 is expressed at the perivascular foot processes of astrocytes, in the glia limitans but also at the ependyma. Most studies have focus on studying the pathogenicity of NMO-IgG on astrocytes and NMO is now considered an astrocytopathy. However, periependymal lesions are observed in NMO suggesting that ependymal cells could also be targeted by NMO-IgG. Ependymal cells regulate CSF-parenchyma molecular exchanges, CSF flow and is a niche for subventricular neural stem cells. Our aim was to examine the effect of antibodies from NMO patients on ependymal cells. We exposed two models, i.e., primary culture of rat ependymal cells and explant cultures of rat lateral ventricular wall wholemounts, to purified IgG of NMO patients (NMO-IgG) for 24 hours. We then evaluated the treatment effect using immunolabeling, functional assays, ependymal flow analysis and bulk RNA sequencing. For each experiment, the effects were compared to purified IgG of healthy donor and to non-treated cells. We found that: i) NMO-IgG patients induced AQP4 agglomeration at the surface of ependymal cells, and induced cell enlargement in comparison to controls. In parallel, it induced an increase in gap junction connexin-43 plaque size; ii) NMO-IgG altered the orientation of ciliary basal bodies and functionally impaired cilia motility; iii) NMO-IgG activated the proliferation of subventricular neural stem cells; and iv) treatment with NMO-IgG upregulated the expression of pro-inflammatory cytokines and chemokines in the transcriptomic analysis. Our study showed that NMO-IgG can trigger an early and specific reactive phenotype in ependymal cells, with functional alterations of intercellular communication and cilia, activation of the subventricular stem cell proliferation and the secretion of pro-inflammatory cytokines. These findings suggest a key role for ependymal cells in the early phase of NMO lesion formation.

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