An increasing number of epilepsy patients are afflicted with drug-resistant temporal lobe epilepsy (TLE) and require alternative therapeutic approaches. High-affinity glycine receptors (haGlyRs) are functionally adapted to tonic inhibition due to their response to hippocampal ambient glycine, and their synthesis is activity-dependent. Therefore, in our study, we scanned TLE hippocampectomies for expression of haGlyRs and characterized the effects mediated by these receptors using primary hippocampal neurons. Increased haGlyR expression occurred in TLE hippocampi obtained from patients with a severe course of disease. Furthermore, in TLE patients, haGlyR and potassium chloride cotransporter 2 (KCC2) expressions were inversely regulated. To examine this potential causal relationship with respect to TLE histopathology, we established a hippocampal cell culture system utilising tonic inhibition mediated by haGlyRs in response to hippocam-pal ambient glycine and in the context of a high Cl equilibrium potential, as is the case in TLE hippocampal neurons. We showed that hypoactive neurons increase their ratio between glutamatergic and GABAergic synapses, reduce their dendrite length and finally undergo excitotoxicity. Pharmacological dissection of the underlying processes revealed ionotropic glutamate and TrkB receptors as critical mediators between neuronal hypoactivity and the emergence of these TLE-characteristic histopathological signs. Moreover, our results indicate a beneficial role for KCC2, because decreasing the Cl− equilibrium potential by KCC2 expression also rescued hypoactive hippocampal neurons. Thus, our data support a causal relationship between increased haGlyR expression and the emergence of histopathological TLE-characteristic signs, and they establish a pathophysiological role for neuronal hypoactivity in the context of a high Cl− equilibrium potential.

The function of supramedullary glycine receptors (GlyRs) is still unclear. Using Wistar rat collicular slices, we demonstrate GlyR-mediated inhibition of spike discharge elicited by low glycine (10 microM). Searching for the molecular basis of this phenomenon, we identified a new GlyR isoform. GlyR alpha3(P185L), a result of cytidine 554 deamination, confers high glycine sensitivity (EC50 approximately 5 microM) to neurons and thereby promotes the generation of sustained chloride conductances associated with tonic inhibition. The level of GlyR alpha3-C554U RNA editing is sensitive to experimentally induced brain lesion, inhibition of cytidine deamination by zebularine and inhibition of mRNA transcription by actinomycin D, but not to blockade of protein synthesis by cycloheximide. Conditional regulation of GlyR alpha3(P185L) is thus likely to be part of a post-transcriptional adaptive mechanism in neurons with enhanced excitability.

Astrocytes are abundant within mature neural circuits and are involved in brain disorders. Here, we summarise our current understanding of astrocytes and Huntington’s disease (HD) with a focus on correlative and causative dysfunctions of ion homeostasis, calcium signaling, and neurotransmitter clearance, as well as on the use of transplanted astrocytes to produce therapeutic benefit in mouse models of HD. Overall, the data suggest astrocyte dysfunction may be an important contributor to the onset and progression of some HD symptoms in mice. Additional exploration of astrocytes in HD mouse models and humans is needed and may provide new therapeutic opportunities to explore in conjunction with neuronal rescue and repair strategies.

Horseradish peroxidase was injected in the somata or axons of neurons located in the intermediate and deep layers of the superior colliculus. A group of 34 neurons with physiologically identified projection in the predorsal bundle (tectobulbo-spinal neurons, TBSNs) and two commissural tecto-tectal neurons were characterized with regard to soma-dendritic profiles, axon trajectories, collateral branching, and terminations. TBSNs belong to the class of large, multipolar, wide field neurons. They send axons through the deep white layer without generating local collaterals. Prior to decussation, all TBSNs bifurcate into an ascending branch which reaches the caudal diencephalon, and a main axon descending to the medulla or spinal cord. Regularly spaced collaterals supply a variety of structures at all rostro-caudal levels. In the midbrain, preterminal and terminal ramifications are present in the medial and lateral reticular tegmentum, in the central grey (including its supraoculo-motor zone), in the nuclei of Cajal and Dark-schewitsch and in the medial aspects of the prerubral area and the fields of Forel. Rhombencephalic targets of TBSNs include the medial pontine and bulbar reticular formation, the abducens nucleus, the nucleus reticularis tegmenti pontis and the nucleus prepositus hypoglossi. An increased density of terminal ramifications was found in several brain stem regions related to the control of eye and head movements. The widespread connections of each individual TBSN suggest that neurons of this type may provide a spatio-temporal pattern of facilitation which promotes rapid orientation of eyes, head and body towards the contralateral hemifield but does not specify the details of movement to be executed.

Notch and neurotrophins control neuronal shape, but it is not known whether their signaling pathways intersect. Here we report results from hippocampal neuronal cultures that are in support of this possibility. We found that low cell density or blockade of Notch signaling by a soluble Delta-Fc ligand decreased the mRNA levels of the nuclear targets of Notch, the homologues of enhancer-of-split 1 and 5 (Hes1/5). This effect was associated with enhanced sprouting of new dendrites or dendrite branches. In contrast, high cell density or exposure of low-density cultures to NGF increased the Hes1/5 mRNA, reduced the number of primary dendrites and promoted dendrite elongation. The NGF effects on both Hes1/5 expression and dendrite morphology were prevented by p75-antibody (a p75 NTR -blocking antibody) or transfection with enhancer-of-split 6 (Hes6), a condition known to suppress Hes activity. Nuclear translocation of NF-kappaB was identified as a link between p75 NTR and Hes1/5 because it was required for the up-regulation of these two genes. The convergence of the Notch and p75 NTR signaling pathways at the level of Hes1/5 illuminates an unexpected mechanism through which a diffusible factor (NGF) could regulate dendrite growth when cell-cell interaction via Notch is not in action.

We have previously shown that dendrite morphology of cultured hippocampal neurones is controlled by Notch receptor activation or binding of nerve growth factor (NGF) to its low affinity receptor p75 NTR , i.e. processes that up-regulate the expression of the Homologue of enhancer of split 1 and 5. Thus, the increased expression of these genes decreases the number of dendrites, whereas abrogation Neurotrophins have been shown to regulate dendrite morphology in a variety of experimental models (Mcallister et al. 1995(Mcallister et al. , 1997Baker et al. 1998;Jin et al. 2003). As neurotrophins are released in an activity-dependent manner, they may be fundamental in orchestrating the structural modifications that developing and mature neuronal circuits undergo (Whitford et al. 2002). However, the signalling pathways underlying the effects of neurotrophins on dendrite morphology are not fully understood. While most studies in this area have emphasized the importance of the Trk receptors, there is evidence that nerve growth factor (NGF) regulates dendrite morphology by binding to p75 NTR , the common neurotrophin receptor (Salama-Cohen et al. 2005) Address correspondence and reprint requests to Dr A. Rodríguez-Tébar, Instituto Cajal, CSIC, Avenue. Doctor Arce, 37, 28002 Madrid, Spain. E-mail: rodriguez@cajal.csic.esAbbreviations used: DIV, days in vitro; EGFP, enhanced green fluorescent protein; E/I, ratio, ratio of excitatory to inhibitory; GAD, glutamic acid decarboxylase; Hes, homologue of enhancer of split; Mash1: mouse achaete scute homologue 1; MAP, microtubule-associated protein; NGF, nerve growth factor; Ngn3, neurogenin 3; p75-Ab, p75 NTR blocking antibody; ROI, region of interest; Syp, synaptophysin; VgluT, vesicular glutamate transporter; VIAAT, vesicular inhibitory amino acid transporter.

The extracellular concentration of the two main neurotransmitters glutamate and GABA is low but not negligible which enables a number of tonic actions. The effects of ambient GABA vary in a region-, cell-type, and age-dependent manner and can serve as indicators of disease-related alterations. Here we explored the tonic inhibitory actions of GABA in Huntington's disease (HD). HD is a devastating neurodegenerative disorder caused by a mutation in the huntingtin gene. Whole cell patch clamp recordings from striatal output neurons (SONs) in slices from adult wild type mice and two mouse models of HD (Z_Q175_KI homozygotes or R6/2 heterozygotes) revealed an HD-related reduction of the GABA(A) receptor-mediated tonic chloride current (ITonic(GABA)) along with signs of reduced GABA(B) receptor-mediated presynaptic depression of synaptic GABA release. About half of ITonic(GABA) depended on tetrodotoxin-sensitive synaptic GABA release, but the remaining current was still lower in HD. Both in WT and HD, ITonic(GABA) was more prominent during the first 4 h after preparing the slices, when astrocytes but not neurons exhibited a transient depolarization. All further tests were performed within 1–4 h in vitro. Experiments with SNAP5114, a blocker of the astrocytic GABA transporter GAT-3, suggest that in WT but not HD GAT-3 operated in the releasing mode. Application of a transportable substrate for glutamate transporters (D-aspartate 0.1–1 mM) restored the non-synaptic GABA release in slices from HD mice. ITonic(GABA) was also rescued by applying the hyperagonist gaboxadol (0.33 μM). The results lead to the hypothesis that lesion-induced astrocyte depolarization facilitates non-synaptic release of GABA through GAT-3. However, the capacity of depolarized astrocytes to provide GABA for tonic inhibition is strongly reduced in HD.

Paired pulse depression (PPD) is a common form of short‐term synaptic plasticity. The aim of this study was to characterise PPD at the level of a single inhibitory bouton. Low‐density collicular cultures were loaded with the Ca2+ indicator Oregon Green‐1, active boutons were stained with RH414, and action potentials were blocked with TTX. Evoked IPSCs (eIPSCs) and presynaptic Ca2+ transients were recorded in response to direct presynaptic depolarisation of an individual bouton. The single bouton eIPSCs had a low failure rate (< 0.1), large average quantal content (3‐6) and slow decay (τ1= 15 ms, τ2= 81 ms). The PPD of eIPSCs had two distinct components: PPDfast and PPDslow (τ= 86 ms and 2 s). PPDslow showed no dependence on extracellular Ca2+ concentration, or on the first eIPSC's failure rate or amplitude. Most probably, it reflects a release‐independent inhibition of exocytosis. PPDfast was only observed in normal or elevated Ca2+. It decreased with the failure rate and increased with the amplitude of the first eIPSC. It coincided with paired pulse depression of the presynaptic Ca2+ transients (τ= 120 ms). The decay of the latter was accelerated by EGTA, which also reduced PPDfast. Therefore, a suppressive effect of residual presynaptic Ca2+ on subsequent Ca2+ influx is considered the most likely cause of PPDfast. PPDfast may also have a postsynaptic component, because exposure to a low‐affinity GABAA receptor antagonist (TPMPA; 300 μM) counteracted PPDfast, and asynchronous IPSC amplitudes were depressed for a short interval following an eIPSC. Thus, at these synapses, PPD is produced by at least two release‐independent presynaptic mechanisms and one release‐dependent postsynaptic mechanism.