Zsuzsanna Callaerts-Vegh

Vesicular Glutamate Transporter VGLUT2 Expression Levels Control Quantal Size and Neuropathic Pain

Uptake of l-glutamate into synaptic vesicles is mediated by vesicular glutamate transporters (VGLUTs). Three transporters (VGLUT1–VGLUT3) are expressed in the mammalian CNS, with partial overlapping expression patterns, and VGLUT2 is the most abundantly expressed paralog in the thalamus, midbrain, and brainstem. Previous studies have shown that VGLUT1 is necessary for glutamatergic transmission in the hippocampus, but the role of VGLUT2 in excitatory transmission is unexplored in glutamatergic neurons and in vivo. We examined the electrophysiological and behavioral consequences of loss of either one or both alleles of VGLUT2. We show that targeted deletion of VGLUT2 in mice causes perinatal lethality and a 95% reduction in evoked glutamatergic responses in thalamic neurons, although hippocampal synapses function normally. Behavioral analysis of heterozygous VGLUT2 mice showed unchanged motor function, learning and memory, acute nociception, and inflammatory pain, but acquisition of neuropathic pain, maintenance of conditioned taste aversion, and defensive marble burying were all impaired. Reduction or loss of VGLUT2 in heterozygous and homozygous VGLUT2 knock-outs led to a graded reduction in the amplitude of the postsynaptic response to single-vesicle fusion in thalamic neurons, indicating that the vesicular VGLUT content is critically important for quantal size and demonstrating that VGLUT2-mediated reduction of excitatory drive affects specific forms of sensory processing.

Concomitant deficits in working memory and fear extinction are functionally dissociated from reduced anxiety in metabotropic glutamate receptor 7-deficient mice.

Metabotropic glutamate receptor 7 (mGluR7), a receptor with a distinct brain distribution and a putative role in anxiety, emotional responding, and spatial working memory, could be an interesting therapeutic target for fear and anxiety disorders. mGluR7-deficient (mGluR7−/−) mice showed essentially normal performance in tests for neuromotor and exploratory activity and passive avoidance learning but prominent anxiolytic behavior in two anxiety tests. They showed a delayed learning curve during the acquisition of the hidden-platform water maze, and three interspersed probe trials indicated that mGluR7−/− mice were slower to acquire spatial information. Working memory in the water maze task and the radial arm maze was impaired in mGluR7−/− mice compared with mGluR7+/+. mGluR7−/− mice also displayed a higher resistance to extinction of fear-elicited response suppression in a conditioned emotional response protocol. In a non-fear-based water maze protocol, mGluR7−/− mice displayed similar delayed extinction. These observed behavioral changes are probably not attributable to changes in AMPA or NMDA receptor function because expression levels of AMPA and NMDA receptors were unaltered. Extinction of conditioned fear is an active and context-dependent form of inhibitory learning and an experimental model for therapeutic fear reduction. It appears to depend on glutamatergic and higher-level brain functions similar to those involved in spatial working memory but functionally dissociated from those that mediate constitutional responses in anxiety tests.

Low hippocampal PI(4,5)P2 contributes to reduced cognition in old mice as a result of loss of MARCKS

Cognitive and motor performances decline during aging. Although it is clear that such signs reflect synaptic compromise, the underlying mechanisms have not been defined. We found that the levels and activity of the synaptic plasticity modulators phosphatidylinositol-(4,5)-bisphosphate (PI(4,5)P2) and phospholipase Cγ (PLCγ) were substantially reduced in hippocampal synaptic membranes from old mice. In addition, these membranes contained reduced levels of the PI(4,5)P2-clustering molecule myristoylated alanine-rich C kinase substrate (MARCKS). Consistent with a cause-effect relationship, raising MARCKS levels in the brain of old mice led to increased synaptic membrane clustering of PI(4,5)P2 and to PLCγ activation. MARCKS overexpression in the hippocampus of old mice or intraventricular perfusion of MARCKS peptide resulted in enhanced long-term potentiation and improved memory. These results reveal one of the mechanisms involved in brain dysfunction during aging.

TBP as a candidate gene for mental retardation in patients with subtelomeric 6q deletions

Monozygotic twin brothers with a subtelomeric 6q deletion presented with mental retardation, microcephaly, seizures, an enlarged cisterna magna, dimpling at elbows, a high arched palate and a thin upper lip. The same subtelomeric deletion was detected in the mother of the patients, presenting with a milder phenotype. We narrowed down the breakpoint to a region of approximately 100u2009kb and estimated the size of the terminal deletion to be 1.2u2009Mb. This region contains four known and seven putative genes. Comparison of the deletion with other reported patients showed TBP was the most plausible candidate gene for the mental retardation in this syndrome. We verified that the TBP gene expression was halved in our patients using real-time PCR. Cognitive and behavioural tests performed on previously described heterozygous tbp mice suggested that TBP is potentially involved in cognitive development.

Notch3 Arg170Cys Knock-In Mice Display Pathologic and Clinical Features of the Neurovascular Disorder Cerebral Autosomal Dominant Arteriopathy With Subcortical Infarcts and Leukoencephalopathy

Objective—Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) is an adult-onset neurovascular disorder caused by stereotyped mutations in the NOTCH3 receptor. Elucidation of its pathobiology is still incomplete and remains a challenge, in part because the available preclinical mouse models to date do not reproduce the full spectrum of CADASIL pathology and clinical disease. Methods and Results—Here, we report a novel knock-in mouse with Arg170Cys substitution in murine Notch3, corresponding to the prevalent Arg169Cys substitution in CADASIL. The Notch3Arg170Cys mice displayed late-onset, dominant CADASIL arteriopathy with typical granular osmiophilic material deposition and developed brain histopathology including thrombosis, microbleeds, gliosis, and microinfarction. Furthermore, Notch3Arg170Cys mice experienced neurological symptoms with motor defects such as staggering gait and limb paresis. Conclusion—This model, for the first time, phenocopies the arteriopathy and the histopathologic as well as clinical features of CADASIL and may offer novel opportunities to investigate disease pathogenesis.

The Early Developmental Gene Semaphorin 5c Contributes to Olfactory Behavior in Adult Drosophila

Behaviors are complex traits influenced by multiple pleiotropic genes. Understanding the mechanisms that give rise to complex behaviors requires an understanding of how variation in transcriptional regulation shapes nervous system development and how variation in brain structure influences an organisms ability to respond to its environment. To begin to address this problem, we used olfactory behavior in Drosophila melanogaster as a model and showed that a hypomorphic transposon-mediated mutation of the early developmental gene Semaphorin-5c (Sema-5c) results in aberrant behavioral responses to the repellant odorant benzaldehyde. We fine mapped this effect to the Sema-5c locus using deficiency mapping, phenotypic reversion through P-element excision, and transgenic rescue. Morphometric analysis of this Sema-5c allele reveals subtle neuroanatomical changes in the brain with a reduction in the size of the ellipsoid body. High-density oligonucleotide expression microarrays identified 50 probe sets with altered transcriptional regulation in the Sema-5c background and quantitative complementation tests identified epistatic interactions between nine of these coregulated genes and the transposon-disrupted Sema-5c gene. Our results demonstrate how hypomorphic mutation of an early developmental gene results in genomewide transcriptional consequences and alterations in brain structure accompanied by profound impairment of adult behavior.

Tetraspanin 6: A novel regulator of hippocampal synaptic transmission and long term plasticity

Tetraspanins (Tspan) are transmembrane proteins with important scaffold and signalling functions. Deletions of Tetraspanin 6 (Tspan6) gene, a member of the tetraspanin family, have been reported in patients with Epilepsy Female-restricted with Mental Retardation (EFMR). Interestingly, mutations in Tspan7, highly homologous to Tspan6, are associated with X-linked intellectual disability, suggesting that these two proteins are important for cognition. Considering recent evidences showing that Tspan7 plays a key role in synapse development and AMPAR trafficking, we initiated the study of Tspan6 in synaptic function using a Tspan6 knock out mouse model. Here we report that hippocampal field recordings from Tspan6 knock out mice show an enhanced basal synaptic transmission and impaired long term potentiation (LTP). A normal paired-pulse facilitation response suggests that Tspan6 affects the properties of the postsynaptic rather than the presynaptic terminal. However, no changes in spine morphology or postsynaptic markers could be detected in Tspan6 KO mice compared with wild types. In addition, Tspan6 KO mice show normal locomotor behaviour and no defects in hippocampus-dependent memory tests.

Improved spatial learning is associated with increased hippocampal but not prefrontal long-term potentiation in mGluR4 knockout mice

Although much information about metabotropic glutamate receptors (mGluRs) and their role in normal and pathologic brain function has been accumulated during the last decades, the role of group III mGluRs is still scarcely documented. Here, we examined mGluR4 knockout mice for types of behavior and synaptic plasticity that depend on either the hippocampus or the prefrontal cortex (PFC). We found improved spatial short‐ and long‐term memory in the radial arm maze, which was accompanied by enhanced long‐term potentiation (LTP) in hippocampal CA1 region. In contrast, LTP in the PFC was unchanged when compared with wild‐type controls. Changes in paired‐pulse facilitation that became overt in the presence of the GABAA antagonist picrotoxin indicated a function of mGluR4 in maintaining the excitation/inhibition balance, which is of crucial importance for information processing in the brain and the deterioration of these processes in neuropsychological disorders such as autism, epilepsy and schizophrenia.

Correction: Tetraspanin 6: A novel regulator of hippocampal synaptic transmission and long term plasticity.

[This corrects the article DOI: 10.1371/journal.pone.0171968.].