Mouse Model Of DYT1 Dystonia Research Articles

Ultrastructural analysis of nigrostriatal dopaminergic terminals in a knockin mouse model of DYT1 dystonia.

DYT1 dystonia is associated with decreased striatal dopamine release. In this study, we examined the possibility that ultrastructural changes of nigrostriatal dopamine terminals could contribute to this neurochemical imbalance using a serial block face/scanning electron microscope (SBF/SEM) and three-dimensional reconstruction to analyse striatal tyrosine hydroxylase-immunoreactive (TH-IR) terminals and their synapses in a DYT1(ΔE) knockin (DYT1-KI) mouse model of DYT1 dystonia. Furthermore, to study possible changes in vesicle packaging capacity of dopamine, we used transmission electron microscopy to assess the synaptic vesicle size in striatal dopamine terminals. Quantitative comparative analysis of 80 fully reconstructed TH-IR terminals in the WT and DYT1-KI mice indicate (1) no significant difference in the volume of TH-IR terminals; (2) no major change in the proportion of axo-spinous versus axo-dendritic synapses; (3) no significant change in the post-synaptic density (PSD) area of axo-dendritic synapses, while the PSDs of axo-spinous synapses were significantly smaller in DYT1-KI mice; (4) no significant change in the contact area between TH-IR terminals and dendritic shafts or spines, while the ratio of PSD area/contact area decreased significantly for both axo-dendritic and axo-spinous synapses in DYT1-KI mice; (5) no significant difference in the mitochondria volume; and (6) no significant difference in the synaptic vesicle area between the two groups. Altogether, these findings suggest that abnormal morphometric changes of nigrostriatal dopamine terminals and their post-synaptic targets are unlikely to be a major source of reduced striatal dopamine release in DYT1 dystonia.

A dystonia mouse model with motor and sequencing deficits paralleling human disease

The dystonias are a group of movement disorders characterized by involuntary twisting movements and postures. A lack of well characterized behavioral models of dystonia has impeded identification of circuit abnormalities giving rise to the disease. Most mouse behavioral assays are implemented independently of cortex, but cortical dysfunction is implicated in human dystonia. It is therefore important to identify dystonia models in which motor cortex-dependent behaviors are altered in ways relevant to human disease. The goal of this study was to characterize a cortically-dependent behavior in the recently-developed Dlx-CKO mouse model of DYT1 dystonia. Mice performed two tasks: skilled reaching and water-elicited grooming. These tests assess motor learning, dexterous skill, and innate motor sequencing. Furthermore, skilled reaching depends strongly on motor cortex, while dorsal striatum is critical for normal grooming. Dlx-CKO mice exhibited significantly lower success rates and pellet contacts compared to control mice during skilled reaching. Despite the skilled reaching impairments, Dlx-CKO mice adapt their reaching strategies. With training, they more consistently contacted the target. Grooming patterns of Dlx-CKO mice are more disorganized than in control mice, as evidenced by a higher proportion of non-chain grooming. However, when Dlx-CKO mice engage in syntactic chains, they execute them similarly to control mice. These abnormalities may provide targets for preclinical intervention trials, as well as facilitate determination of the physiologic path from torsinA dysfunction to motor phenotype.

TorsinA restoration in a mouse model identifies a critical therapeutic window for DYT1 dystonia.

In inherited neurodevelopmental diseases, pathogenic processes unique to critical periods during early brain development may preclude the effectiveness of gene modification therapies applied later in life. We explored this question in a mouse model of DYT1 dystonia, a neurodevelopmental disease caused by a loss-of-function mutation in the TOR1A gene encoding torsinA. To define the temporal requirements for torsinA in normal motor function and gene replacement therapy, we developed a mouse line enabling spatiotemporal control of the endogenous torsinA allele. Suppressing torsinA during embryogenesis caused dystonia-mimicking behavioral and neuropathological phenotypes. Suppressing torsinA during adulthood, however, elicited no discernible abnormalities, establishing an essential requirement for torsinA during a developmental critical period. The developing CNS exhibited a parallel "therapeutic critical period" for torsinA repletion. Although restoring torsinA in juvenile DYT1 mice rescued motor phenotypes, there was no benefit from adult torsinA repletion. These data establish a unique requirement for torsinA in the developing nervous system and demonstrate that the critical period genetic insult provokes permanent pathophysiology mechanistically delinked from torsinA function. These findings imply that to be effective, torsinA-based therapeutic strategies must be employed early in the course of DYT1 dystonia.

The abnormal firing of Purkinje cells in the knockin mouse model of DYT1 dystonia

DYT1 dystonia is an inherited movement disorder caused by a heterozygous trinucleotide (GAG) deletion in DYT1/TOR1A, coding for torsinA. Growing evidence suggests that the cerebellum plays a role in the pathogenesis of dystonia. Brain imaging of both DYT1 dystonia patients and animal models show abnormal activity in the cerebellum. The cerebellum-specific knockdown of torsinA in adult mice leads to dystonia-like behavior. Dyt1 ΔGAG heterozygous knock-in mouse model exhibits impaired corticostriatal long-term depression, abnormal muscle co-contraction, and motor deficits. We and others previously reported altered dendritic structures in Purkinje cells in Dyt1 knock-in mouse models. However, whether there are any electrophysiological alterations of the Purkinje cells in Dyt1 knock-in mice is not known. We used the patch-clamp recording in brain slices and in acutely dissociated Purkinje cells to identify specific alterations of Purkinje cells firing. We found abnormal firing of non-tonic type of Purkinje cells in the Dyt1 knock-in mice. Furthermore, the large-conductance calcium-activated potassium (BK) current and the BK channel protein levels were significantly increased in the Dyt1 knock-in mice. Our results support a role of the cerebellum in the pathogenesis of DYT1 dystonia. Manipulating the Purkinje cell firing and cerebellar output may show great promise for treating DYT1 dystonia.

TorsinB overexpression prevents abnormal twisting in DYT1 dystonia mouse models.

Genetic redundancy can be exploited to identify therapeutic targets for inherited disorders. We explored this possibility in DYT1 dystonia, a neurodevelopmental movement disorder caused by a loss-of-function (LOF) mutation in the TOR1A gene encoding torsinA. Prior work demonstrates that torsinA and its paralog torsinB have conserved functions at the nuclear envelope. This work established that low neuronal levels of torsinB dictate the neuronal selective phenotype of nuclear membrane budding. Here, we examined whether torsinB expression levels impact the onset or severity of abnormal movements or neuropathological features in DYT1 mouse models. We demonstrate that torsinB levels bidirectionally regulate these phenotypes. Reducing torsinB levels causes a dose-dependent worsening whereas torsinB overexpression rescues torsinA LOF-mediated abnormal movements and neurodegeneration. These findings identify torsinB as a potent modifier of torsinA LOF phenotypes and suggest that augmentation of torsinB expression may retard or prevent symptom development in DYT1 dystonia.

Trihexyphenidyl rescues the deficit in dopamine neurotransmission in a mouse model of DYT1 dystonia

Trihexyphenidyl, a nonselective muscarinic receptor antagonist, is the small molecule drug of choice for the treatment of DYT1 dystonia, but it is poorly tolerated due to significant side effects. A better understanding of the mechanism of action of trihexyphenidyl is needed for the development of improved treatments. Because DTY1 dystonia is associated with both abnormal cholinergic neurotransmission and abnormal dopamine regulation, we tested the hypothesis that trihexyphenidyl normalizes striatal dopamine release in a mouse model of DYT1 dystonia using ex vivo fast scan cyclic voltammetry and in vivo microdialysis. Trihexyphenidyl increased striatal dopamine release and efflux as assessed by ex vivo voltammetry and in vivo microdialysis respectively. In contrast, ʟ-DOPA, which is not usually effective for the treatment of DYT1 dystonia, did not increase dopamine release in either Dyt1 or control mice. Trihexyphenidyl was less effective at enhancing dopamine release in Dyt1 mice relative to controls ex vivo (mean increase WT: 65% vs Dyt1: 35%). Trihexyphenidyl required nicotinic receptors but not glutamate receptors to increase dopamine release. Dyt1 mice were more sensitive to the dopamine release decreasing effects of nicotinic acetylcholine receptor antagonism (IC50: WT = 29.46 nM, Dyt1 = 12.26 nM) and less sensitive to acetylcholinesterase inhibitors suggesting that nicotinic acetylcholine receptor neurotransmission is altered in Dyt1 mice, that nicotinic receptors indirectly mediate the differential effects of trihexyphenidyl in Dyt1 mice, and that nicotinic receptors may be suitable therapeutic targets for DYT1 dystonia.

RGS9‐2 rescues dopamine D2 receptor levels and signaling in DYT1 dystonia mouse models

Dopamine D2 receptor signaling is central for striatal function and movement, while abnormal activity is associated with neurological disorders including the severe early‐onset DYT1 dystonia. Nevertheless, the mechanisms that regulate D2 receptor signaling in health and disease remain poorly understood. Here, we identify a reduced D2 receptor binding, paralleled by an abrupt reduction in receptor protein level, in the striatum of juvenile Dyt1 mice. This occurs through increased lysosomal degradation, controlled by competition between β‐arrestin 2 and D2 receptor binding proteins. Accordingly, we found lower levels of striatal RGS9‐2 and spinophilin. Further, we show that genetic depletion of RGS9‐2 mimics the D2 receptor loss of DYT1 dystonia striatum, whereas RGS9‐2 overexpression rescues both receptor levels and electrophysiological responses in Dyt1 striatal neurons. This work uncovers the molecular mechanism underlying D2 receptor downregulation in Dyt1 mice and in turn explains why dopaminergic drugs lack efficacy in DYT1 patients despite significant evidence for striatal D2 receptor dysfunction. Our data also open up novel avenues for disease‐modifying therapeutics to this incurable neurological disorder.

Forebrain knock-out of torsinA reduces striatal free-water and impairs whole-brain functional connectivity in a symptomatic mouse model of DYT1 dystonia

Multiple lines of evidence implicate striatal dysfunction in the pathogenesis of dystonia, including in DYT1, a common inherited form of the disease. The impact of striatal dysfunction on connected motor circuits and their interaction with other brain regions is poorly understood. Conditional knock-out (cKO) of the DYT1 protein torsinA from forebrain cholinergic and GABAergic neurons creates a symptomatic model that recapitulates many characteristics of DYT1 dystonia, including the developmental onset of overt twisting movements that are responsive to antimuscarinic drugs. We performed diffusion MRI and resting-state functional MRI on cKO mice of either sex to define abnormalities of diffusivity and functional connectivity in cortical, subcortical, and cerebellar networks. The striatum was the only region to exhibit an abnormality of diffusivity, indicating a selective microstructural deficit in cKO mice. The striatum of cKO mice exhibited widespread increases in functional connectivity with somatosensory cortex, thalamus, vermis, cerebellar cortex and nuclei, and brainstem. The current study provides the first in vivo support that direct pathological insult to forebrain torsinA in a symptomatic mouse model of DYT1 dystonia can engage genetically normal hindbrain regions into an aberrant connectivity network. These findings have important implications for the assignment of a causative region in CNS disease.

In vivo imaging reveals impaired connectivity across cortical and subcortical networks in a mouse model of DYT1 dystonia

Developing in vivo functional and structural neuroimaging assays in Dyt1 ΔGAG heterozygous knock-in (Dyt1 KI) mice provide insight into the pathophysiology underlying DYT1 dystonia. In the current study, we examined in vivo functional connectivity of large-scale cortical and subcortical networks in Dyt1 KI mice and wild-type (WT) controls using resting-state functional magnetic resonance imaging (MRI) and an independent component analysis. In addition, using diffusion MRI we examined how structural integrity across the basal ganglia and cerebellum directly relates to impairments in functional connectivity. Compared to WT mice, Dyt1 KI mice revealed increased functional connectivity across the striatum, thalamus, and somatosensory cortex; and reduced functional connectivity in the motor and cerebellar cortices. Further, Dyt1 KI mice demonstrated elevated free-water (FW) in the striatum and cerebellum compared to WT mice, and increased FW was correlated with impairments in functional connectivity across basal ganglia, cerebellum, and sensorimotor cortex. The current study provides the first in vivo MRI-based evidence in support of the hypothesis that the deletion of a 3-base pair (ΔGAG) sequence in the Dyt1 gene encoding torsinA has network level effects on in vivo functional connectivity and microstructural integrity across the sensorimotor cortex, basal ganglia, and cerebellum.

Electromyographic evidence in support of a knock-in mouse model of DYT1 Dystonia.

DYT1 dystonia is an autosomal-dominant movement disorder characterized by abnormal, often repetitive, movements and postures. Its hallmark feature is sustained or intermittent contractions of muscles involving co-contractions of antagonist muscle pairs. The symptoms are relieved with the anticholinergic drug trihexyphenidyl. The primary mutation is a trinucleotide deletion (ΔGAG) in DYT1/TOR1A, which codes for torsinA. Previous studies showed that (1) heterozygous Dyt1 ΔGAG knock-in mice, which have an analogous mutation in the endogenous gene, exhibit motor deficits and altered corticostriatal synaptic plasticity in the brain and (2) these deficits can be rescued by trihexyphenidyl. However, brain imaging studies suggest that the Dyt1 knock-in mouse models nonmanifesting mutation carriers of DYT1 dystonia. The aim of this work was to examine the hallmark features of DYT1 dystonia in the Dyt1 knock-in mice by analyzing muscular activities. Wireless telemetry devices with biopotential channels were implanted to the bicep and the rectus femori muscles in Dyt1 knock-in mice, and muscular activities were recorded before and after trihexyphenidyl administration. (1) Consistent with DYT1 dystonia patients, Dyt1 knock-in mice showed sustained contractions and co-contractions of the antagonistic bicep femoris and rectus femoris. (2) The abnormal muscle contractions were normalized by trihexyphenidyl. The results suggest that the motor deficits in Dyt1 knock-in mice are likely produced by abnormal muscle contractions, and Dyt1 knock-in mice can potentially be used as a manifesting disease model to study pathophysiology and develop novel therapeutics. © 2016 International Parkinson and Movement Disorder Society.

Dopamine receptor and Gα(olf) expression in DYT1 dystonia mouse models during postnatal development.

BackgroundDYT1 dystonia is a heritable, early-onset generalized movement disorder caused by a GAG deletion (ΔGAG) in the DYT1 gene. Neuroimaging studies and studies using mouse models suggest that DYT1 dystonia is associated with dopamine imbalance. However, whether dopamine imbalance is key to DYT1 or other forms of dystonia continues to be debated.Methodology/Principal FindingsWe used Dyt1 knock out (Dyt1 KO), Dyt1 ΔGAG knock-in (Dyt1 KI), and transgenic mice carrying one copy of the human DYT1 wild type allele (DYT1 hWT) or human ΔGAG mutant allele (DYT1 hMT). D1R, D2R, and Gα(olf) protein expression was analyzed by western blot in the frontal cortex, caudate-putamen and ventral midbrain in young adult (postnatal day 60; P60) male mice from all four lines; and in the frontal cortex and caudate putamen in juvenile (postnatal day 14; P14) male mice from the Dyt1 KI and KO lines. Dopamine receptor and Gα(olf) protein expression were significantly decreased in multiple brain regions of Dyt1 KI and Dyt1 KO mice and not significantly altered in the DYT1 hMT or DYT1 hWT mice at P60. The only significant change at P14 was a decrease in D1R expression in the caudate-putamen of the Dyt1 KO mice.Conclusion/SignificanceWe found significant decreases in key proteins in the dopaminergic system in multiple brain regions of Dyt1 KO and Dyt1 KI mouse lines at P60. Deletion of one copy of the Dyt1 gene (KO mice) produced the most pronounced effects. These data offer evidence that impaired dopamine receptor signaling may be an early and significant contributor to DYT1 dystonia pathophysiology.

Direct measurement of the Hypercholinergic state in a mouse model of DYT1 Dystonia

Direct measurement of the Hypercholinergic state in a mouse model of DYT1 Dystonia

Regional specificity of synaptic plasticity deficits in a knock-in mouse model of DYT1 dystonia

DYT1 dystonia is a movement disorder caused by a deletion in the C-terminal of the protein torsinA. It is unclear how torsinA mutation might disrupt cellular processes encoding motor activity, and whether this impairment occurs in specific brain regions. Here, we report a selective impairment of corticostriatal synaptic plasticity in knock-in mice heterozygous for Δ-torsinA (Tor1a+/Δgag mice) as compared to controls (Tor1a+/+ mice). In striatal spiny neurons from Tor1a+/Δgag mice, high-frequency stimulation failed to induce long-term depression (LTD), whereas long-term potentiation (LTP) exhibited increased amplitude. Of interest, blockade of D2 dopamine receptors (D2Rs) increased LTP in Tor1a+/+ mice to a level comparable to that measured in Tor1a+/Δgag mice and normalized the levels of potentiation across mouse groups. A low-frequency stimulation (LFS) protocol was unable to depotentiate corticostriatal synapses in Tor1a+/Δgag mice. Muscarinic M1 acetylcholine receptor (mAChR) blockade rescued plasticity deficits. Additionally, we found an abnormal responsiveness of cholinergic interneurons to D2R activation, consisting in an excitatory response rather than the expected inhibition, further confirming an imbalance between dopaminergic and cholinergic signaling in the striatum. Conversely, synaptic activity and plasticity in the CA1 hippocampal region were unaltered in Tor1a+/Δgag mice. Importantly, the M1 mAChR-dependent enhancement of hippocampal LTP was unaffected in both genotypes. Similarly, both basic properties of dopaminergic nigral neurons and their responses to D2R activation were normal.These results provide evidence for a regional specificity of the electrophysiological abnormalities observed and demonstrate the reproducibility of such alterations in distinct models of DYT1 dystonia.

Subtle microstructural changes of the cerebellum in a knock-in mouse model of DYT1 dystonia

The dystonias are a group of disorders characterized by involuntary twisting and repetitive movements. DYT1 dystonia is an inherited form of dystonia caused by a mutation in the TOR1A gene, which encodes torsinA. TorsinA is expressed in many regions of the nervous system, and the regions responsible for causing dystonic movements remain uncertain. Most prior studies have focused on the basal ganglia, although there is emerging evidence for abnormalities in the cerebellum too. In the current studies, we examined the cerebellum for structural abnormalities in a knock-in mouse model of DYT1 dystonia. The gross appearance of the cerebellum appeared normal in the mutant mice, but stereological measures revealed the cerebellum to be 5% larger in mutant compared to control mice. There were no changes in the numbers of Purkinje cells, granule cells, or neurons of the deep cerebellar nuclei. However, Golgi histochemical studies revealed Purkinje cells to have thinner dendrites, and fewer and less complex dendritic spines. There also was a higher frequency of heterotopic Purkinje cells displaced into the molecular layer. These results reveal subtle structural changes of the cerebellum that are similar to those reported for the basal ganglia in the DYT1 knock-in mouse model.

Dystonia-associated protein torsinA is not detectable at the nerve terminals of central neurons

Presynaptic functions of the mammalian central neurons are regulated by a network of protein interactions. Synaptic vesicle recycling in and neurotransmitter release from the presynaptic nerve terminals are altered when a glutamate-deleting mutation is present in the torsinA protein (ΔE-torsinA). This mutation is linked with a hereditary form of the movement disorder dystonia known as DYT1 dystonia. Although torsinA expression is prevalent throughout the central nervous system, its subcellular localization – in particular with respect to presynaptic nerve terminals – remains unclear. This information would be useful in narrowing down possible models for how wild-type torsinA affects presynaptic function, as well as the nature of the presynaptic dysfunction that arises in the context of ΔE-torsinA mutation. Here we report on an analysis of the presynaptic localization of torsinA in cultured neurons obtained from a knock-in mouse model of DYT1 dystonia. Primary cultures of neurons were established from heterozygous and homozygous ΔE-torsinA knock-in mice, as well as from their wild-type littermates. Neurons were obtained from the striatum, cerebral cortex and hippocampus of these mice, and were subjected to immunocytochemistry. This analysis revealed the expression of both proteins in the somata and dendrites. However, neither the nerve terminals nor axonal shafts were immunoreactive. These results were confirmed by fluorogram-based quantitation. Our findings indicate that neither the wild-type nor the ΔE-torsinA mutant protein is present at substantial levels in the presynaptic structures of cultured neurons. Thus, the effects of torsinA, in wild-type and mutant forms, appear to influence presynaptic function indirectly, without residing in presynaptic structures.

Abnormal cytoplasmic calcium dynamics in central neurons of a dystonia mouse model

Increased activities of cytoplasmic calcium and the excitatory neurotransmitter glutamate have been independently implicated in dystonia pathophysiology. However, cellular-level evidence linking these two features is not available. Here we show that glutamate-dependent changes in neuronal calcium dynamics occur in a knock-in mouse model of DYT1 dystonia, the most common hereditary form of this disorder. Fluorescence-based analysis of the dynamics of cytoplasmic calcium concentration ([Ca2+]c) in cultured hippocampal neurons shows that electrical stimulation depolarizes the neurons and increases the dendritic [Ca2+]c, which then decays slowly to the pre-stimulus level. Whereas the peak amplitude of [Ca2+]c was not affected, the decay period was prolonged in neurons of heterozygous mice whose genotype reflects the human condition. We found that this effect was blocked by the antagonists of ionotropic glutamate receptors, and confirmed that glutamate receptors are present in these neurons. As the [Ca2+]c is readout and regulator of neuronal excitability, its abnormality represents an important cellular phenotype of dystonia

Functional analysis of dopaminergic systems in a DYT1 knock-in mouse model of dystonia

The dystonias are a group of disorders characterized by involuntary twisting movements and abnormal posturing. The most common of the inherited dystonias is DYT1 dystonia, which is due to deletion of a single GAG codon (ΔE) in the TOR1A gene that encodes torsinA. Since some forms of dystonia have been linked with dysfunction of brain dopamine pathways, the integrity of these pathways was explored in a knock-in mouse model of DYT1 dystonia. In DYT1(ΔE) knock-in mice, neurochemical measures revealed only small changes in the content of dopamine or its metabolites in tissue homogenates from caudoputamen or midbrain, but microdialysis studies revealed robust decreases in baseline and amphetamine-stimulated extracellular dopamine in the caudoputamen. Quantitative stereological methods revealed no evidence for striatal or midbrain atrophy, but substantia nigra neurons immunopositive for tyrosine hydroxylase were slightly reduced in numbers and enlarged in size. Behavioral studies revealed subtle abnormalities in gross motor activity and motor coordination without overt dystonia. Neuropharmacological challenges of dopamine systems revealed normal behavioral responses to amphetamine and a minor increase in sensitivity to haloperidol. These results demonstrate that this DYT1(ΔE) knock-in mouse model of dystonia harbors neurochemical and structural changes of the dopamine pathways, as well as motor abnormalities.

Genetic Background Modulates the Phenotype of a Mouse Model of DYT1 Dystonia

DYT1 dystonia is a debilitating neurological disease characterized by involuntary twisting movements. The disease is caused by an in-frame deletion (GAG, “ΔE”) mutation in the TOR1A gene that encodes the torsinA protein. Intriguingly, only 30% of mutation carriers exhibit motor symptoms despite the fact that functional brain imaging studies show abnormal brain metabolism in all carriers. Because genetic modifiers may be a determinant of this reduced penetrance, we examined the genetic contribution of three different inbred strains of mice on the DYT1 mutation in animals that are homozygous (Tor1aΔE/ΔE) or heterozygous (Tor1aΔE/+; disease state) for the disease-causing ΔE mutation. We find that the DBA/2J, C57BL/6J, and CD1-ICR contribution of genes significantly alter lifespan in Tor1aΔE/ΔE mice, which die during the first few days of life on the 129S6/SvEvTac (129) background. The C57BL/6J (B6) strain significantly decreases life expectancy of Tor1aΔE/ΔE animals but, like 129S6/SvEvTac Tor1aΔE/+ mice, congenic C57BL/6J Tor1aΔE/+ mice do not exhibit any motor abnormalities. In contrast, the DBA/2J (D2) strain significantly increases life expectancy. This effect was not present in congenic DBA/2J Tor1aΔE/ΔE mice, indicating that the extended lifespan of F2 129/D2 mice was due to a combination of homozygous and heterozygous allelic effects. Our observations suggest that genetic modifiers may alter the penetrance of the ΔE mutation, and that mapping these modifiers may provide fresh insight into the torsinA molecular pathway.

Synaptic vesicle recycling is enhanced by torsinA that harbors the DYT1 dystonia mutation

Early-onset generalized dystonia, DYT1, is caused by a mutation in the gene encoding the evolutionarily conserved AAA+ ATPase torsinA. Synaptic abnormalities have been implicated in DYT1 dystonia, but the details of the synaptic pathophysiology are only partially understood. Here, we demonstrate a novel role for torsinA in synaptic vesicle recycling, using cultured hippocampal neurons from a knock-in mouse model of DYT1 dystonia (ΔE-torsinA) and live-cell imaging with styryl FM dyes. Neurons from heterozygous ΔE-torsinA mice released a larger fraction of the total recycling pool (TRP) during a single round of electrical stimulation than did wild-type neurons. Moreover, when the neurons were subjected to prior high activity, the time course of release was shortened. In neurons from homozygous mice, these enhanced exocytosis phenotypes were similar, but in addition the size of the TRP was reduced. Notably, when release was triggered by applying a calcium ionophore rather than electrical stimuli, neither a single nor two ΔE-torsinA alleles affected the time course of release. Thus, the site of action of ΔE-torsinA is at or upstream of the rise in calcium concentration in nerve terminals. Our results suggest that torsinA regulates synaptic vesicle recycling in central neurons. They also indicate that this regulation is influenced by neuronal activity, further supporting the idea that synaptic abnormalities contribute to the pathophysiology of DYT1 dystonia.

An anticholinergic reverses motor control and corticostriatal LTD deficits in Dyt1 ΔGAG knock-in mice

DYT1 early-onset generalized torsion dystonia is an inherited movement disorder associated with mutations in DYT1 that codes for torsinA protein. The most common mutation seen in this gene is a trinucleotide deletion of GAG. We previously reported a motor control deficit on a beam-walking task in our Dyt1 ΔGAG knock-in heterozygous mice. In this report we show the reversal of this motor deficit with the anticholinergic trihexyphenidyl (THP), a drug commonly used to treat movement problems in dystonia patients. THP also restored the reduced corticostriatal long-term depression (LTD) observed in these mice. Corticostriatal LTD has long been known to be dependent on D2 receptor activation. In this mouse model, striatal D2 receptors were expressed at lower quantities in comparison to wild-type mice. Furthermore, the mice were also partially resistant to FPL64176, an agonist of L-type calcium channels that have been previously reported to cause severe dystonic-like symptoms in wild-type mice. Our findings collectively suggest that altered communication between cholinergic interneurons and medium spiny neurons is responsible for the LTD deficit and that this synaptic plasticity modification may be involved in the striatal motor control abnormalities in our mouse model of DYT1 dystonia.