Overt Dystonia Research Articles

Conditional Knockout of Striatal Gnal Produces Dystonia-like Motor Phenotypes.

Loss-of-function mutations in GNAL have been linked to an adult-onset, isolated dystonia that is largely indistinguishable from idiopathic dystonia. GNAL encodes Gα olf , a heterotrimeric G-protein α subunit with a defined molecular function to increase the production of the second messenger cAMP. Gα olf is abundant in the striatum, and is the only stimulatory G-protein in many cell types of the striatum. Due to the defined molecular signaling pathway and expression pattern of Gα olf , the clear genetic link to dystonia makes GNAL an exciting target to understand the pathological mechanisms of not only this genetic dystonia, but also the larger idiopathic disease. To better understand GNAL -linked dystonia, we generated a novel genetic mouse model that allows us to conditionally knock out Gnal in a site and time-specific manner. In the current study we used genetic or AAV based approaches to express Cre to knockout striatal Gnal in our novel Gnal fl/fl model. We then performed motor behavioral testing and ex vivo whole-cell patch clamp electrophysiology of striatal spiny projection neurons to interrogate how loss of Gnal leads to dystonia. Mice with conditional striatal knockout of Gnal show hindlimb clasping, other dystonia-like postures, less motor coordination, slowness, and torticollis as compared to age-matched controls. Furthermore, striatal spiny projection neurons show increased excitability in Gnal knockout animals. These exciting data are the first to report uninduced, overt dystonia in a mouse model of GNAL- linked dystonia, and directly correlate these with changes in spiny projection neuron electrophysiological properties. Our results show that adult loss of Gnal in the striatum leads to the development of dystonia, through homeostatic, paradoxical increases in spiny projection neuron excitability, and suggest that therapeutic strategies aimed at decreasing this hyperexcitable phenotype may provide symptomatic relief for patients with disease. One Sentence Summary: When Gnal is knocked out in the striatum of mice we observe overt behavioral symptoms and hyperexcitability in striatal spiny projection neurons.

Comparing Essential Tremor with and without Soft Dystonic Signs and Tremor Combined with Dystonia: The TITAN Study.

Tremor disorders remain as clinical diagnoses and the rate of misdiagnosis between the commonest non-parkinsonian tremors is relatively high. To compare the clinical features of Essential Tremor without other features (pure ET), ET plus soft dystonic signs (ET + DS), and tremor combined with dystonia (TwD). We compared the clinical features of patients with pure ET, ET + DS, and TwD enrolled in The ITAlian tremor Network (TITAN). Linear regression models were performed to determine factors associated with health status and quality of life. Three-hundred-eighty-three patients were included. Sex distribution was significantly different between the groups with males being more represented in pure ET and females in TwD. The initial site of tremor was different between the groups with about 40% of TwD having head tremor and ET + DS unilateral upper limb tremor at onset. This pattern mirrored the distribution of overt dystonia and soft dystonic signs at examination. Sensory trick, task-specificity, and position-dependence were more common, but not exclusive, to TwD. Pure ET patients showed the lowest degree of alcohol responsiveness and ET + DS the highest. Midline tremor was more commonly encountered and more severe in TwD than in the other groups. Regression analyses demonstrated that tremor severity, sex, age, and to a lesser degree the variable "group", independently predicted health status and quality of life, suggesting the existence of other determinants beyond tremor. Pure ET and TwD manifest with a phenotypic overlap, which calls for the identification of diagnostic biomarkers. ET + DS shared features with both syndromes, suggesting intra-group heterogeneity.

In vivo optogenetic inhibition of striatal parvalbumin-reactive interneurons induced genotype-specific changes in neuronal activity without dystonic signs in male DYT1 knock-in mice.

The pathophysiology of early-onset torsion dystonia (TOR1A/DYT1) remains unclear. Like 70% of human mutation carriers, rodent models with ΔGAG mutation such as DYT1 knock-in (KI) mice do not show overt dystonia but have subtle sensorimotor deficits and pattern of abnormal synaptic plasticity within the striatal microcircuits. There is evidence that dysfunction of striatal parvalbumin-reactive (Parv+) fast-spiking interneurons (FSIs) can be involved in dystonic signs. To elucidate the relevance of these GABAergic interneurons in the pathophysiology of DYT1 dystonia, we used in vivo optogenetics to specifically inhibit Parv+ and to detect changes in motor behavior and neuronal activity. Optogenetic fibers were bilaterally implanted into the dorsal striatum of male DYT1 KI mice and wild-type (WT) littermates expressing halorhodopsin (eNpHR3.0) in Parv+ interneurons. While stimulations with yellow light pulses for up to 60 min at different pulse durations and interval lengths did not induce abnormal movements, such as dystonic signs, immunohistochemical examinations revealed genotype-dependent differences. In contrast to WT mice, stimulated DYT1 KI showed decreased striatal neuronal activity, that is, less c-Fos reactive neurons, and increased activation of cholinergic interneurons after optogenetic inhibition of Parv+ interneurons. These findings suggest an involvement of Parv+ interneurons in an impaired striatal network in DYT1 KI mice, but at least short-term inhibition of these GABAergic interneurons is not sufficient to trigger a dystonic phenotype, similar to previously shown optogenetic activation of cholinergic interneurons.

Motor deficit and lack of overt dystonia in Dlx conditional Dyt1 knockout mice

DYT1 or DYT-TOR1A dystonia is early-onset generalized dystonia caused by a trinucleotide deletion of GAG in the TOR1A or DYT1 gene leads to the loss of a glutamic acid residue in the resulting torsinA protein. A mouse model with overt dystonia is of unique importance to better understand the DYT1 pathophysiology and evaluate preclinical drug efficacy. DYT1 dystonia is likely a network disorder involving multiple brain regions, particularly the basal ganglia. Tor1a conditional knockout in the striatum or cerebral cortex leads to motor deficits, suggesting the importance of corticostriatal connection in the pathogenesis of dystonia. Indeed, corticostriatal long-term depression impairment has been demonstrated in multiple targeted DYT1 mouse models. Pappas and colleagues developed a conditional knockout line (Dlx-CKO) that inactivated Tor1a in the forebrain and surprisingly displayed overt dystonia. We set out to validate whether conditional knockout affecting both cortex and striatum would lead to overt dystonia and whether machine learning-based video behavioral analysis could be used to facilitate high throughput preclinical drug screening. We generated Dlx-CKO mice and found no overt dystonia or motor deficits at 4 months. At 8 months, retesting revealed motor deficits in rotarod, beam walking, grip strength, and hyperactivity in the open field; however, no overt dystonia was visually discernible or through the machine learning-based video analysis. Consistent with other targeted DYT1 mouse models, we observed age-dependent deficits in the beam walking test, which is likely a better motor behavioral test for preclinical drug testing but more labor-intensive when overt dystonia is absent.

A man with an arched back

A 19-year-old Chinese man presented with a four-month history of progressive lower back pain and stiffness. He had progressive truncal arching, but no limb, cranial or vocal abnormalities. This affected his ability to walk and fall asleep. His symptoms worsened with anxiety, but improved during deep sleep. Four years prior, he had been diagnosed with depression. He was taking fluoxetine intermittently, but there was no prior neuroleptic exposure. A trial of levodopa/benserazide up to 100 mg twice daily for over three months had no effect. His brother was unaffected, but his father reported feeling a cramping sensation in his dominant hand when writing under stressful conditions – the latter had a bilateral postural hand tremor without overt dystonia on examination.

Motor Deficit and Lack of Overt Dystonia in Dlx Conditional Dyt1 Knockout Mice

Motor Deficit and Lack of Overt Dystonia in Dlx Conditional Dyt1 Knockout Mice

Optogenetic augmentation of the hypercholinergic endophenotype in DYT1 knock-in mice induced erratic hyperactive movements but not dystonia.

BackgroundThe most prevalent inherited form of generalized dystonia is caused by a mutation in torsinA (DYT1, ∆GAG) with incomplete penetrance. Rodent models with mutated torsinA do not develop dystonic symptoms, but previous ex vivo studies indicated abnormal excitation of cholinergic interneurons (ChI) and increased striatal acetylcholine.MethodsWe used in vivo optogenetics to exacerbate this endophenotype in order to determine its capacity to trigger dystonic symptoms in freely behaving mice. Tor1a+/Δgag DYT1 mice and wildtype littermates expressing channelrhodopsin2 under the Chat promotor were implanted bilaterally with optical LED cannulae and stimulated with blue light pulses of varied durations.FindingsSix months old DYT1 KI mice but not wildtype controls responded with hyperactivity to blue light specifically at 25 ms pulse duration, 10 Hz frequency. Neuronal activity (c-Fos) in cholinergic interneurons was increased immediately after light stimulation and persisted only in DYT1 KI over 15 min. Substance P was increased specifically in striosome compartments in naïve DYT1 KI mice compared to wildtype. Under optogenetic stimulation substance P increased in wildtype to match levels in Dyt1 KI, and acetylcholinesterase was elevated in the striatum of stimulated DYT1 KI. No signs of dystonic movements were observed under stimulation of up to one hour in both genotypes and age groups, and the sensorimotor deficit previously observed in 6 months old DYT1 KI mice persisted under stimulation.InterpretationOverall this supports an endophenotype of dysregulated cholinergic activity in DYT1 dystonia, but depolarizing cholinergic interneurons was not sufficient to induce overt dystonia in DYT1 KI mice.

P 77 Optogenetic stimulations of striatal cholinergic interneurons in an animal model of dystonia

Dystonias are movement disorders, defined by sustained or intermittent muscle contractions causing twisting movements and postures. The most prevalent inherited form of dystonia is caused by a mutation in the gene for torsin A (DYT1, ΔGAG) with incomplete penetrance. It has been hypothesized that an increased activity of cholinergic interneurons in the striatum, resulting in abnormal synaptic plasticity, plays an important role in the disease pathophysiology. However, this hypothesis is merely based on ex vivo electrophysiological recordings in brain slices of animal models which do not show a dystonic phenotype. Here we aim to substantiate the role of this alteration for motor dysfunctions in vivo by using optogenetic activation of striatal cholinergic interneurons in freely behaving mice. For this purpose we crossed Dyt1 ΔGAG heterozygous knock-in mice (DYT1) with mice expressing a light-dependent cation channel (Channelrhodopsin2, ChR2), essential for optogenetic examinations, specifically in cholinergic interneurons (Chat promotor). ChR2 is opened by blue light (wavelength 470 nm), leading to depolarization and acetylcholine release. The light (LED) is conveyed by optical cannulae, which were chronically implanted by stereotaxic surgery into the striatum of DYT1/ChR2 mice and wildtype (WT)/ChR2 littermates. First, we ensured that ChR2 is expressed in cholinergic interneurons of ChR2 + mice by immunohistochemistry and confocal microscopy and that these neurons respond to blue light exposure in brain slices. Subsequently, effects of different stimulation parameters (e.g. pulse width, Hz) on motor activity were examined in WT/ChR2 ( n = 9) and DYT1/ChR2 mice ( n = 6). Bilateral and unilateral photostimulations with blue LED light were compared to yellow LED light which does not open ChR2 (negative control). As expected, yellow light had no influence on behavior. Effects of bilateral stimulations with blue light were more robust than unilateral stimulations. Selected stimulation parameters induced significant increases of general activity (increased distance moved) specifically in DYT1/ChR2 mice compared to before or after stimulation, while there were no effects in WT/ChR2. This substantiates a specific role of striatal cholinergic neurons in motor activity in DYT1 dystonia. We have recently shown that DYT1 KI mice, which do not develop overt dystonia, present with sensorimotor deficits in specific behavioral paradigms. Optogenetic activation of cholinergic interneurons ameliorated this deficit temporarily. Preliminary data of an investigation of neuronal activity in the striatum after stimulation indicated an increased number of ChR2 cells positive for c-Fos in Dyt1/ChR2 ( n = 4) in comparison to WT/ChR2 ( n = 5) while there was no difference in total number of c-Fos positive cells. Total number of c-Fos positive cells did also not differ between naive WT/ChR2 and Dyt1/ChR2 mice, supporting that increased c-Fos expression in cholinergic interneurons is the expected effect of optogenetic stimulation on neuronal activity. We now perform comprehensive studies with short- and long-term stimulations in the Dyt1 mice to clarify the importance of overactive striatal cholinergic interneurons for the manifestation and abnormal striatal plasticity in DYT1 dystonia. For this purpose optogenetics will be combined with pharmacological, electrophysiological, neurochemical and molecular examinations.

Tremor entities and their classification: an update.

This review focuses on important new findings in the field of tremor and illustrates the consequences for the current definition and classification of tremor. Since 1998 when the consensus criteria for tremor were proposed, new variants of tremors and new diagnostic methods were discovered that have changed particularly the concepts of essential tremor and dystonic tremor. Accumulating evidence exists that essential tremor is not a single entity rather different conditions that share the common symptom action tremor without other major abnormalities. Tremor is a common feature in patients with adult-onset focal dystonia and may involve several different body parts and forms of tremor. Recent advances, in particular, in the field of genetics, suggest that dystonic tremor may even be present without overt dystonia. Monosymptomatic asymmetric rest and postural tremor has been further delineated, and apart from tremor-dominant Parkinson's disease, there are several rare conditions including rest and action tremor with normal dopamine transporter imaging (scans without evidence of dopaminergic deficit) and essential tremor with tremor at rest. Increasing knowledge in the last decades changed the view on tremors and highlights several caveats in the current tremor classification. Given the ambiguous assignment between tremor phenomenology and tremor etiology, a more cautious definition of tremors on the basis of clinical assessment data is needed.

Adult-Onset Primary Dystonic Tics: A Different Entity?

Based on their phenomenology, tics can be classified as clonic, dystonic, and tonic. Although tic syndromes are considered to be childhood disorders, there are reports on (clonic) tics developing late in life. Literature on dystonic tics is sparse, and it is unclear whether adult-onset dystonic tics are part of the same disorder spectrum that includes Tourette's syndrome or represent a discrete entity. We describe here 11 patients with adult-onset primary dystonic tics. Ten patients (90.1%) were males. Mean age at onset was 42.2±14.9years. More than 60% had both motor clonic and dystonic tics. Dystonic tics most frequently involved the cranial-cervical region and the shoulders and, less frequently, the limbs. Psychiatric comorbidities were present in 5 patients. Family history for any movement disorder or for psychiatric disorders was present in 2 cases. One patient showed a sensory geste, which allowed him to partially control his tics, whereas another developed overt dystonia 3years after his first assessment. The hyperkinesias exhibited by these patients were likely consistent with tics. However, other clinical features would allow us to argue that adult-onset dystonic tics may represent a discrete entity, which is intermediate between tics and dystonia.

Rest and other types of tremor in adult-onset primary dystonia

IntroductionKnowledge regarding tremor prevalence and phenomenology in patients with adult-onset primary dystonia is limited. Dystonic tremor is presumably under-reported, and we aimed to assess the prevalence and the clinical correlates...

P 182. Normal lateral inhibition mechanism during sensory-motor plasticity in dystonia

Objective To explore if sensory information is processed and integrated during sensory-motor plasticity phenomena by using lateral inhibition mechanisms in normal humans and in patients with dystonia. Background Several evidence suggest that lateral inhibition is a system within sensory-motor cortex operating during the acquisition of new motor tasks in order to select the appropriate muscle sequence to be stored within the final motor engram. This mechanism is thought to be lost in dystonia and this should explain the development of redundant motor memories which could culminate in overflow phenomena and overt dystonia. Methods We have used transcranial magnetic stimulation to explore lateral inhibition during sensory-motor plasticity in 12 dystonic patients (7 focal hand dystonia, 5 cranial dystonia) and in 8 healthy subjects. In particular we looked at motor evoked potential (MEP) facilitation, in the abductor pollicis brevis (APB) and abductor digiti minimi (ADM), obtained after 5 Hz repetitive paired associative stimulation after median (PAS M), ulnar nerve stimulation (PAS U) and median + ulnar nerve (PAS MU) stimulation. In this way we evaluated the ratio MU/M + Ux100 (lateral inhibition index). Moreover, we evaluated the lateral inhibition index (Li index). This parameter is easily obtained by using the following formula: ratio MU/(M + U) x 100. MU is the MEP facilitation measured after PAS with simultaneous stimulation of median and ulnar and M + U is the amount of MEP facilitation, after PAS, induced from stimulation of the individual nerves. Results Our data confirmed that patients with dystonia had two main abnormalities: first the amount of facilitation was larger than normal subjects; second and more important the spatial specificity was lost. A three-factorial ANOVA demonstrated a significant time × group × conditioning interaction ( F = 7.9; p = 0.005). Lateral inhibition index was similar (about 50%) in healthy subjects and dystonic patients. Conclusions These data suggest that lateral inhibition is normal in dystonia during sensory-motor plasticity. Another mechanism could contribute to the formation of motor memories with redundant information, which could culminate in overt 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.

Cholinergic dysregulation produced by selective inactivation of the dystonia-associated protein torsinA

DYT1 dystonia, a common and severe primary dystonia, is caused by a 3-bp deletion in TOR1A which encodes torsinA, a protein found in the endoplasmic reticulum. Several cellular functions are altered by the mutant protein, but at a systems level the link between these and the symptoms of the disease is unclear. The most effective known therapy for DYT1 dystonia is the use of anticholinergic drugs. Previous studies have revealed that in mice, transgenic expression of human mutant torsinA under a non-selective promoter leads to abnormal function of striatal cholinergic neurons. To investigate what pathological role torsinA plays in cholinergic neurons, we created a mouse model in which the Dyt1 gene, the mouse homolog of TOR1A, is selectively deleted in cholinergic neurons (ChKO animals). These animals do not have overt dystonia, but do have subtle motor abnormalities. There is no change in the number or size of striatal cholinergic cells or striatal acetylcholine content, uptake, synthesis, or release in ChKO mice. There are, however, striking functional abnormalities of striatal cholinergic cells, with paradoxical excitation in response to D2 receptor activation and loss of muscarinic M2/M4 receptor inhibitory function. These effects are specific for cholinergic interneurons, as recordings from nigral dopaminergic neurons revealed normal responses. Amphetamine stimulated dopamine release was also unaltered. These results demonstrate a cell-autonomous effect of Dyt1 deletion on striatal cholinergic function. Therapies directed at modifying the function of cholinergic neurons may prove useful in the treatment of the human disorder.

Dopa-responsive dystonia simulating cerebral palsy

Five patients presented in infancy or early childhood with various combinations of pyramidal and extrapyramidal signs with normal cognitive function. Their perinatal courses were unremarkable. In each patient, initial impressions listed by several examiners included spastic diplegia or cerebral palsy. Later in each course, either extrapyramidal features or progression suggested dopa-responsive dystonia. In 4 of the 5 children, cerebrospinal fluid was obtained and disclosed reduced levels of biopterin, neopterin, and homovanillic acid in all 4. Levodopa therapy resulted in prompt improvement with normal function returning within 6 months. The disappearance of the “spasticity,” extensor plantar responses, and extrapyramidal signs, following levodopa therapy, confirmed the diagnosis of dopa-responsive dystonia in these patients. Three had apparently sporadic disease; the other 2 were siblings with an affected paternal grandmother. Three had onset in infancy with delayed sitting and walking before the appearance of overt dystonia; infantile onset is infrequent in dopa-responsive dystonia. The other 2 had normal milestones, but developed gait disorders with prominent imbalance in early childhood. The diagnosis of dopa-responsive dystonia should be considered in children with unexplained or atypical “cerebral palsy.”