Altered fronto-striatal functions in the Gdi1 -null mouse model of X-linked Intellectual Disability

RAB-GDP dissociation inhibitor 1 (GDI1) loss-of-function mutations are responsible for a form of non-specific X-linked Intellectual Disability (XLID) where the only clinical feature is cognitive impairment. GDI1 patients are impaired in specific aspects of executive functions and conditioned response, which are controlled by fronto-striatal circuitries. Previous molecular and behavioral characterization of the Gdi1-null mouse revealed alterations in the total number/distribution of hippocampal and cortical synaptic vesicles as well as hippocampal short-term synaptic plasticity, and memory deficits. In this study, we employed cognitive protocols with high translational validity to human condition that target the functionality of cortico-striatal circuitry such as attention and stimulus selection ability with progressive degree of complexity. We previously showed that Gdi1-null mice are impaired in some hippocampus-dependent forms of associative learning assessed by aversive procedures. Here, using appetitive-conditioning procedures we further investigated associative learning deficits sustained by the fronto-striatal system. We report that Gdi1-null mice are impaired in attention and associative learning processes, which are a key part of the cognitive impairment observed in XLID patients

Forebrain Deletion of αGDI in Adult Mice Worsens the Pre-Synaptic Deficit at Cortico-Lateral Amygdala Synaptic Connections

The GDI1 gene encodes αGDI, which retrieves inactive GDP-bound RAB from membranes to form a cytosolic pool awaiting vesicular release. Mutations in GDI1 are responsible for X-linked Intellectual Disability. Characterization of the Gdi1-null mice has revealed alterations in the total number and distribution of hippocampal and cortical synaptic vesicles, hippocampal short-term synaptic plasticity and specific short-term memory deficits in adult mice, which are possibly caused by alterations of different synaptic vesicle recycling pathways controlled by several RAB GTPases. However, interpretation of these studies is complicated by the complete ablation of Gdi1 in all cells in the brain throughout development. In this study, we generated conditionally gene-targeted mice in which the knockout of Gdi1 is restricted to the forebrain, hippocampus, cortex and amygdala and occurs only during postnatal development. Adult mutant mice reproduce the short-term memory deficit previously reported in Gdi1-null mice. Surprisingly, the delayed ablation of Gdi1 worsens the pre-synaptic phenotype at cortico-amygdala synaptic connections compared to Gdi1-null mice. These results suggest a pivotal role of αGDI via specific RAB GTPases acting specifically in forebrain regions at the pre-synaptic sites involved in memory formation

Cognitive impairment in Gdi1-deficient mice is associated with altered synaptic vesicle pools and short-term synaptic plasticity, and can be corrected by appropriate learning training

The GDI1 gene, responsible in human for X-linked non-specific mental retardation, encodes αGDI, a regulatory protein common to all GTPases of the Rab family. Its alteration, leading to membrane accumulation of different Rab GTPases, may affect multiple steps in neuronal intracellular traffic. Using electron microscopy and electrophysiology, we now report that lack of αGDI impairs several steps in synaptic vesicle (SV) biogenesis and recycling in the hippocampus. Alteration of the SV reserve pool (RP) and a 50% reduction in the total number of SV in adult synapses may be dependent on a defective endosomal-dependent recycling and may lead to the observed alterations in short-term plasticity. As predicted by the synaptic characteristics of the mutant mice, the short-term memory deficit, observed when using fear-conditioning protocols with short intervals between trials, disappeared when the Gdi1 mutants were allowed to have longer intervals between sessions. Likewise, previously observed deficits in radial maze learning could be corrected by providing less challenging pre-training. This implies that an intact RP of SVs is necessary for memory processing under challenging conditions in mice. The possibility to correct the learning deficit in mice may have clinical implication for future studies in huma

Deletion of the mental retardation gene Gdi1 impairs associative memory and alters social behavior in mice

Non-specific mental retardation (NSMR) is a common human disorder characterized by mental handicap as the only clinical symptom. Among the recently identified MR genes is GDI1, which encodes αGdi, one of the proteins controlling the activity of the small GTPases of the Rab family in vesicle fusion and intracellular trafficking. We report the cognitive and behavioral characterization of mice carrying a deletion of Gdi1. The Gdi1-deficient mice are fertile and anatomically normal. They appear normal also in many tasks to assess spatial and episodic memory and emotional behavior. Gdi1-deficient mice are impaired in tasks requiring formation of short-term temporal associations, suggesting a defect in short-term memory. In addition, they show lowered aggression and altered social behavior. In mice, as in humans, lack of Gdi1 spares most central nervous system functions and preferentially impairs only a few forebrain functions required to form temporal associations. The general similarity to human mental retardation is striking, and suggests that the Gdi1 mutants may provide insights into the human defect and into the molecular mechanisms important for development of cognitive function

A CTNNA3 compound heterozygous deletion implicates a role for αT-catenin in susceptibility to autism spectrum disorder

BACKGROUND: Autism spectrum disorder (ASD) is a highly heritable, neurodevelopmental condition showing extreme genetic heterogeneity. While it is well established that rare genetic variation, both de novo and inherited, plays an important role in ASD risk, recent studies also support a rare recessive contribution. METHODS: We identified a compound heterozygous deletion intersecting the CTNNA3 gene, encoding αT-catenin, in a proband with ASD and moderate intellectual disability. The deletion breakpoints were mapped at base-pair resolution, and segregation analysis was performed. We compared the frequency of CTNNA3 exonic deletions in 2,147 ASD cases from the Autism Genome Project (AGP) study versus the frequency in 6,639 controls. Western blot analysis was performed to get a quantitative characterisation of Ctnna3 expression during early brain development in mouse. RESULTS: The CTNNA3 compound heterozygous deletion includes a coding exon, leading to a putative frameshift and premature stop codon. Segregation analysis in the family showed that the unaffected sister is heterozygote for the deletion, having only inherited the paternal deletion. While the frequency of CTNNA3 exonic deletions is not significantly different between ASD cases and controls, no homozygous or compound heterozygous exonic deletions were found in a sample of over 6,000 controls. Expression analysis of Ctnna3 in the mouse cortex and hippocampus (P0-P90) provided support for its role in the early stage of brain development. CONCLUSION: The finding of a rare compound heterozygous CTNNA3 exonic deletion segregating with ASD, the absence of CTNNA3 homozygous exonic deletions in controls and the high expression of Ctnna3 in both brain areas analysed implicate CTNNA3 in ASD susceptibility

Loss of Either Rac1 or Rac3 GTPase Differentially Affects the Behavior of Mutant Mice and the Development of Functional GABAergic Networks

Rac GTPases regulate the development of cortical/hippocampal GABAergic interneurons by affecting the early development and migration of GABAergic precursors. We have addressed the function of Rac1 and Rac3 proteins during the late maturation of hippocampal interneurons. We observed specific phenotypic differences between conditional Rac1 and full Rac3 knockout mice. Rac1 deletion caused greater generalized hyperactivity and cognitive impairment compared with Rac3 deletion. This phenotype matched with a more evident functional impairment of the inhibitory circuits in Rac1 mutants, showing higher excitability and reduced spontaneous inhibitory currents in the CA hippocampal pyramidal neurons. Morphological analysis confirmed a differential modification of the inhibitory circuits: deletion of either Rac caused a similar reduction of parvalbumin-positive inhibitory terminals in the pyramidal layer. Intriguingly, cannabinoid receptor-1-positive terminals were strongly increased only in the CA1 of Rac1-depleted mice. This increase may underlie the stronger electrophysiological defects in this mutant. Accordingly, incubation with an antagonist for cannabinoid receptors partially rescued the reduction of spontaneous inhibitory currents in the pyramidal cells of Rac1 mutants. Our results show that Rac1 and Rac3 have independent roles in the formation of GABAergic circuits, as highlighted by the differential effects of their deletion on the late maturation of specific populations of interneurons

Neural precursor cells tune striatal connectivity through the release of IGFBPL1

The adult brain retains over life endogenous neural stem/precursor cells (eNPCs) within the subventricular zone (SVZ). Whether or not these cells exert physiological functions is still unclear. In the present work, we provide evidence that SVZ-eNPCs tune structural, electrophysiological, and behavioural aspects of striatal function via secretion of insulin-like growth factor binding protein-like 1 (IGFBPL1). In mice, selective ablation of SVZ-eNPCs or selective abrogation of IGFBPL1 determined an impairment of striatal medium spiny neuron morphology, a higher failure rate in GABAergic transmission mediated by fast-spiking interneurons, and striatum-related behavioural dysfunctions. We also found IGFBPL1 expression in the human SVZ, foetal and induced-pluripotent stem cell-derived NPCs. Finally, we found a significant correlation between SVZ damage, reduction of striatum volume, and impairment of information processing speed in neurological patients. Our results highlight the physiological role of adult SVZ-eNPCs in supporting cognitive functions by regulating striatal neuronal activity

Short-term plasticity at Cortico-LA synapses is perturbed in absence of αGDI.

<p>(<b>A</b>) Iterative stimulations (20 Hz/4 sec) were applied at Cortico-LA synapses in <i>Gdi1</i> WT and <i>Gdi1</i>-null mice. Scale bars: 50 (up) and 100 (bottom) pA and 250 msec. (<b>B</b>) The pronounced synaptic depression in <i>Gdi1</i>-null mice is well illustrated by the extraction of the last EPSCs of the train response. Scale bars: 50 (up) and 100 (bottom) pA and 10 msec. (<b>C</b>) Mean EPSC amplitude at a given stimulation for each genotype. (<b>D</b>) Cumulative plot based on the same data, allowing to visualize the deficit in refilling rate at <i>Gdi1</i>-null synapses (see material and methods section for further details). (<b>E₁</b>) Ready releasable pool size (RRP) and refilling rate (<b>E₂</b>) were calculated in <i>Gdi1</i>-null and <i>Gdi1</i> WT preparations and expressed as % of initial EPSC size. Number of recorded cells is indicated. **p<0,01.</p

Immunofluorescence analysis of <i>Gdi1^lox</i>, <i>Gdi1^flox/Y</i> and <i>Gdi1</i>-null mouse brain regions.

<p>Immunofluorescence analysis of 15 µm coronal sections of <i>Gdi1^lox/Y</i>. <i>Gdi1^flox/Y</i> and <i>Gdi1</i>-null. (<b>A</b>) Low magnification coronal section from <i>Gdi1^lox/Y</i> brain indicating the regions of interest reported on the right: DG (dentate gyrus, <b>B-B″</b> and <b>F-F″</b>), CA1 (hippocampal CA1 region, <b>C-C″</b> and <b>G-G″</b>), LA (lateral amygdala, <b>D-D″</b> and <b>H-H″</b>) and MGm (medial geniculate nucleus, <b>E-E″</b> and <b>I-I″</b>). (<b>A</b>) Scale bar: 1 mm. (<b>B–I″</b>) Scale bar: 0.015 mm.</p

Brain region- and age-specific down regulation of αGDI on <i>Gdi1^flox/Y</i> mice.

<p>Two to eight week old male mice were analysed. (<b>A</b>) Protein lysates were prepared from the indicated tissues (Hip: hippocampus, CC: cerebral cortex, CbC: cerebellum, OB: olfactory bulb, T: thalamus, H: hypothalamus and BLA: amygdala) at the indicated age (PND: post natal days), fractionated on 10% SDS–PAGE gels and analyzed with a commercial anti-GDI antibody. β-tubulin was used as loading control. (<b>B</b>) Quantitative analysis of residual αGDI normalized by β-tubulin in the hippocampus and cerebral cortex. Protein levels were quantified by measuring the intensity of the western blot signal with the Image Quant system. Values are expressed as mean ± SD from three independent animals at each age. Grey squares are <i>Gdi1^lox/Y</i> animals and black squares are <i>Gdi1^flox/Y</i>. *p<0.05, **p<0.01.</p