A loss-of-function variant in canine GLRA1 associates with a neurological disorder resembling human hyperekplexia.

Hereditary hyperekplexia is a rare neuronal disorder characterized by an exaggerated startle response to sudden tactile or acoustic stimuli. In this study, we present a Miniature Australian Shepherd family showing clinical signs, which have genetic and phenotypic similarities with human hereditary hyperekplexia: episodes of muscle stiffness that could occasionally be triggered by acoustic stimuli. Whole genome sequence data analysis of two affected dogs revealed a 36-bp deletion spanning the exon-intron boundary in the glycine receptor alpha 1 (GLRA1) gene. Further validation in pedigree samples and an additional cohort of 127 Miniature Australian Shepherds, 45 Miniature American Shepherds and 74 Australian Shepherds demonstrated complete segregation of the variant with the disease, according to an autosomal recessive inheritance pattern. The protein encoded by GLRA1 is a subunit of the glycine receptor, which mediates postsynaptic inhibition in the brain stem and spinal cord. The canine GLRA1 deletion is located in the signal peptide and is predicted to cause exon skipping and subsequent premature stop codon resulting in a significant defect in glycine signaling. Variants in GLRA1 are known to cause hereditary hyperekplexia in humans; however, this is the first study to associate a variant in canine GLRA1 with the disorder, establishing a spontaneous large animal disease model for the human condition.

Hereditary Hyperekplexia in Saudi Arabia.

BACKGROUND: Hyperekplexia is a rare disorder characterized by exaggerated startle responses to unexpected sensory stimuli, recurrent apneas, and stiffness. Only few studies have been published on this disorder in populations with high rates of consanguinity. METHODS: We retrospectively reviewed Saudi patients with genetically confirmed hereditary hyperekplexia using a standard questionnaire that was sent to nine major referral hospitals in Saudi Arabia. RESULTS: A total of 22 Saudi patients (11 males, 11 females) from 20 unrelated families who had hereditary hyperekplexia were included. Based on molecular studies, they were classified into different subtypes: SLC6A5 variant (12 patients, 54.5%), GLRB variant (seven patients, 31.8%), and GLRA1 variant (three patients, 13.7%). All patients were homozygous for the respective causal variant. The combined carrier frequency of hereditary hyperekplexia for the encountered founder mutations in the Saudi population is 10.9 per 10,000, which translates to a minimum disease burden of 13 patients per 1,000,000. CONCLUSION: Our study provides comprehensive epidemiologic information, prevalence figures, and clinical characteristics of a large cohort of patients with hereditary hyperekplexia.

Hereditary Hyperekplexia: A New Family and a Systematic Review of GLRA1 Gene-Related Phenotypes.

Hereditary hyperekplexia (HPX) is a genetic neurodevelopmental disorder recently defined by the triad of (1) neonatal hypertonia, (2) excessive startle reflexes, and (3) generalized stiffness following the startle. Defects in GLRA1 are the most common cause of HPX, inherited both in an autosomal dominant and autosomal recessive manner. GLRA1 mutations can also cause milder phenotypes in the startle syndromes spectrum, but the prevalence is uncertain and no clear genotype-phenotype correlation has emerged yet. Moreover, the prevalence of neurodevelopmental outcomes has not been clearly defined. Here we report a new family of patients with a typical HPX phenotype, linked to a novel GLRA1 mutation, inherited with a recessive pattern. We then perform a systematic review of the literature of GLRA1-related HPX, describing the main epidemiological features of 210 patients. We found that GLRA1-related phenotypes do not necessarily fulfill the current criteria for HPX, including also milder and later-onset phenotypes. Among clinical features of the disease, neurodevelopmental issues were reported in a third of the sample; interestingly, we found that these problems, particularly when severe, were more common in homozygous than in heterozygous patients. Additional clinical and preclinical studies are needed to define predictors of adverse neurodevelopmental outcomes and underlying mechanisms.

Advances in hyperekplexia and other startle syndromes.

Startle, a basic alerting reaction common to all mammals, is described as a sudden involuntary movement of the body evoked by all kinds of sudden and unexpected stimulus. Startle syndromes are heterogeneous groups of disorders with abnormal and exaggerated responses to startling events, including hyperekplexia, stimulus-induced disorders, and neuropsychiatric startle syndromes. Hyperekplexia can be attributed to a genetic, idiopathic, or symptomatic cause. Excluding secondary factors, hereditary hyperekplexia, a rare neurogenetic disorder with highly genetic heterogeneity, is characterized by neonatal hypertonia, exaggerated startle response provoked by the sudden external stimuli, and followed by a short period of general stiffness. It mainly arises from defects of inhibitory glycinergic neurotransmission. GLRA1 is the major pathogenic gene of hereditary hyperekplexia, along with many other genes involved in the function of glycinergic inhibitory synapses. While about 40% of patients remain negative genetic findings. Clonazepam, which can specifically upgrade the GABARA1 chloride channels, is the main and most effective administration for hereditary hyperekplexia patients. In this review, with the aim at enhancing the recognition and prompting potential treatment for hyperekplexia, we focused on discussing the advances in hereditary hyperekplexia genetics and the expound progress in pathogenic mechanisms of the glycinergic-synapse-related pathway and then followed by a brief overview of other common startle syndromes.

Teaching Video NeuroImage: Hereditary Hyperekplexia Mimicking Tonic Seizures in an Infant.

Hereditary hyperekplexia is a rare neurologic disorder characterized by an exaggerated startle response with profound muscle stiffness.1,2 Given the nature of the spells, this condition is often misdiagnosed as epilepsy. Mutations in glycine receptors and transporters are the primary cause of this syndrome.1 We present an example of stimulus-induced hyperekplexia captured on video EEG in a 7-week-old girl with compound heterozygous variants in the presynaptic glycine transporter gene SLC6A5.

Hyperekplexia and other startle syndromes.

Abnormal startle syndromes are classified into hyperekplexia, stimulus-induced, and neuropsychiatric startle syndromes. Hyperekplexia is attributed to a genetic, idiopathic, or symptomatic cause. Hereditary hyperekplexia is a treatable neurogenetic disorder. In patients with a hyperactive startle response, the first step is to characterize the extent and associations of 'response.' Secondary or symptomatic causes are particularly important in children, as they provide useful clinical clues to an underlying neurodevelopmental or neurodegenerative disorders. Particular attention should be given to any neonate or infant with generalized or episodic stiffness, drug-refractory seizures, recurrent apnea, stimulus-sensitive behavioral states, or sudden infant death syndrome. Eliciting a non-habituating head-retraction reflex to repeated nose tapping should be a part of routine examination of all new-borns. Vigevano maneuver should be taught to all families and health-care workers as an emergency rescue measure. The onset of excessive startle after infancy should direct investigations for an acquired cause such as brainstem injury, antibodies against glycine receptors, and neurodegeneration. Finally, one should not forget to evaluate unexplained cases of abnormal gait and frequent falls in adults for underlying undiagnosed startle syndromes. Oral clonazepam is an effective therapy besides behavioral and safety interventions for hereditary cases. The outcomes in genetic cases are good overall.

A Novel Glycine Receptor Variant with Startle Disease Affects Syndapin I and Glycinergic Inhibition.

Glycine receptors (GlyRs) are the major mediators of fast synaptic inhibition in the adult human spinal cord and brainstem. Hereditary mutations to GlyRs can lead to the rare, but potentially fatal, neuromotor disorder hyperekplexia. Most mutations located in the large intracellular domain (TM3-4 loop) of the GlyRα1 impair surface expression levels of the receptors. The novel GLRA1 mutation P366L, located in the TM3-4 loop, showed normal surface expression but reduced chloride currents, and accelerated whole-cell desensitization observed in whole-cell recordings. At the single-channel level, we observed reduced unitary conductance accompanied by spontaneous opening events in the absence of extracellular glycine. Using peptide microarrays and tandem MS-based analysis methods, we show that the proline-rich stretch surrounding P366 mediates binding to syndapin I, an F-BAR domain protein involved in membrane remodeling. The disruption of the noncanonical Src homology 3 recognition motif by P366L reduces syndapin I binding. These data suggest that the GlyRα1 subunit interacts with intracellular binding partners and may therefore play a role in receptor trafficking or synaptic anchoring, a function thus far only ascribed to the GlyRß subunit. Hence, the P366L GlyRα1 variant exhibits a unique set of properties that cumulatively affect GlyR functionality and thus might explain the neuropathological mechanism underlying hyperekplexia in the mutant carriers. P366L is the first dominant GLRA1 mutation identified within the GlyRα1 TM3-4 loop that affects GlyR physiology without altering protein expression at the whole-cell and surface levels.SIGNIFICANCE STATEMENT We show that the intracellular domain of the inhibitory glycine receptor α1 subunit contributes to trafficking and synaptic anchoring. A proline-rich stretch in this receptor domain forms a noncanonical recognition motif important for the interaction with syndapin I (PACSIN1). The disruption of this motif, as present in a human patient with hyperekplexia led to impaired syndapin I binding. Functional analysis revealed that the altered proline-rich stretch determines several functional physiological parameters of the ion channel (e.g., faster whole-cell desensitization) reduced unitary conductance and spontaneous opening events. Thus, the proline-rich stretch from the glycine receptor α1 subunit represents a multifunctional intracellular protein motif.

C.292G>A, a novel glycine receptor alpha 1 subunit gene (GLRA1) mutation found in a Chinese patient with hyperekplexia: A case report.

INTRODUCTION: Hyperekplexia is a rare hereditary neurological disorder; only 5 glycine receptor alpha 1 subunit gene (GLRA1) mutations have been reported in 5 Chinese patients. We report a Chinese infant with hyperekplexia and a novel mutation at c.292G > A. PATIENT CONCERNS: A Chinese infant with hyperekplexia and a novel mutation at c.292G > A. DIAGNOSIS: All exons of GLRA1 were sequenced in her parents and her, which revealed a mutation at c.1030C > T and another novel mutation at c.292G > A. Her diagnosis was confirmed as hereditary hyperekplexia with GlRA1 hybrid gene mutations based on the sequencing results. INTERVENTIONS: She was treated with clonazepam. OUTCOMES: Her muscle hypertonia recovered rapidly and the excessive startle reflex to unexpected stimuli was significantly reduced. CONCLUSION: Genetic DNA sequencing is a crucial method for diagnosing hyperekplexia-related gene mutation.

Primary DQ effect in the association between HLA and neurological syndromes with anti-GAD65 antibodies.

The primary cause of neurological syndromes with antibodies against glutamic acid decarboxylase 65 (GAD65-Ab) is unknown, but genetic predisposition may exist as it is suggested by the co-occurrence in patients and their relatives of other organ-specific autoimmune diseases, notably type 1 diabetes mellitus (T1DM), and by the reports of a few familial cases. We analyzed the human leukocyte antigen (HLA) in 32 unrelated patients and compared them to an ethnically matched sample of 137 healthy controls. Four-digit resolution HLA alleles were imputed from available Genome Wide Association data, and full HLA next-generation sequencing-based typing was also performed. HLA DQA1*05:01-DQB1*02:01-DRB1*03:01 was the most frequent class II haplotype in patients (13/32, 41%). DQB1*02:01 was the only allele found to be significantly more common in patients than in controls (20/137, 15%, corrected p = 0.03, OR 3.96, 95% CI [1.54-10.09]). There was also a trend towards more frequent DQA1*05:01 among patients compared to controls (22/137, 16%; corrected p = 0.05, OR 3.54, 95% CI [1.40-8.91]) and towards a protective effect of DQB1*03:01 (2/32, 6% in patients vs. 42/137, 31% in control group; corrected p = 0.05, OR 0.15, 95% CI [0.02-0.65]). There was no significant demographic or clinical difference between DQ2 and non-DQ2 carriers (p > 0.05). Taken together, these findings suggest a primary DQ effect on GAD65-Ab neurological diseases, partially shared with other systemic organ-specific autoimmune diseases such as T1DM. However, it is likely that other non-HLA loci are involved in the genetic predisposition of GAD65-Ab neurological syndromes.

[The Frozen Man and the Chinese Alphabet]. / L'Homme Gelé (渐冻人) et le déficit en cristalline αB.

The Confucian philosophy teaches us that the search for truth does not always follow a straight line. The clinical observation presented here illustrates this perfectly and is about a child afflicted by a rare neuromuscular disorder (in Chinese, the word 'myopathy' is translated to meaning 'frozen man') in whom was suspected a deficit in αB crystallin. The authors take the opportunity to put the spotlight on China, this great country which did not wait for Alain Peyrefitte to wake up or, more precisely, to rewake. In the light of past and recent missions in the former Middle Kingdom, an update is made on the medico-scientific but also societal issues of this country on the verge of becoming, perhaps, a giant in the field of neuromuscular diseases.

TITLE: L'Homme Gelé (渐冻人) et le déficit en cristalline αB. ABSTRACT: La philosophie confucéenne nous enseigne que la recherche de la vérité n'emprunte pas toujours un chemin rectiligne. L'observation clinique présentée ici l'illustre parfaitement. Il y est question d'un enfant souffrant d'une maladie neuromusculaire rare (en chinois, le mot myopathie se traduit par 渐冻人 soit « homme gelé ¼) chez qui fut suspecté un déficit en cristalline αB. Les auteurs profitent de l'occasion pour mettre le projecteur sur la Chine, ce grand pays qui n'a pas attendu Alain Peyrefitte pour s'éveiller ou, plus exactement, se réveiller. à la lumière de missions passées et récentes dans l'ex-Empire du Milieu, le point est fait sur les enjeux médico-scientifiques mais aussi sociétaux de ce pays en passe de devenir, peut-être, un géant dans le domaine des maladies neuromusculaires.

A glycine transporter SLC6A5 frameshift mutation causes startle disease in Spanish greyhounds.

Startle disease, or hyperekplexia, is a glycinergic disorder characterized by hypertonia and apnea that is triggered by noise and/or touch. Mutations in five genes have been associated with startle disease in humans, dogs, cattle, and mice. We identified a novel recessive startle disease in a family of Spanish greyhounds. Whole genome resequencing of an affected dog revealed a homozygous two base pair deletion in the ninth exon of SLC6A5, encoding the presynaptic glycine transporter. The deletion is predicted to cause a frameshift, p.S460FfsX47, leading to a premature stop codon that truncates over a third of the protein. Family members were genotyped for the deletion, and findings were consistent with an autosomal recessive inheritance pattern. The pathogenic variant was absent from 34 unrelated greyhounds, 659 domestic dogs of pure and mixed breeds, and 54 wild canids, suggesting it occurred recently and may be private to the family. The findings of this study can be used to inform future breeding decisions and prevent dissemination of the deleterious allele in greyhounds.

Reduced display of conformational epitopes in the N-terminal truncated GAD65 isoform: relevance for people with stiff person syndrome or DQ8/8-positive Type 1 diabetes mellitus.

AIMS: To investigate whether the N-terminal truncated glutamic acid decarboxylase 65 (GAD65) isoform is as well recognized by people with stiff person syndrome as it is by people with Type 1 diabetes, and whether conformational GAD65 antibody epitopes are displayed properly by the isoform. METHODS: GAD65 antibody-positive healthy individuals (n=13), people with stiff-person syndrome (n=15) and children with new-onset Type 1 diabetes (n=654) were analysed to determine binding to full-length GAD65 and the N-terminal truncated GAD65 isoform in each of these settings. GAD65 autoantibody epitope specificity was correlated with binding ratios of full-length GAD65/N-terminal truncated GAD65. RESULTS: The N-terminal truncated GAD65 isoform was significantly less recognized in GAD65Ab-positive people with stiff-person syndrome (P=0.002) and in healthy individuals (P=0.0001) than in people with Type 1 diabetes. Moreover, at least two specific conformational GAD65Ab epitopes were not, or were only partially, presented by the N-terminal truncated GAD65 isoform compared to full-length GAD65. Finally, an N-terminal conformational GAD65Ab epitope was significantly less recognized in DQ8/8 positive individuals with Type 1 diabetes (P=0.02). CONCLUSIONS: In people with stiff person syndrome preferred binding to the full-length GAD65 isoform over the N-terminal truncated molecule was observed. This binding characteristic is probably attributable to reduced presentation of two conformational epitopes by the N-terminal truncated molecule. These findings support the notion of disease-specific GAD65Ab epitope specificities and emphasize the need to evaluate the applicability of novel assays for different medical conditions.

[A pedigree of hereditary hyperekplexia].

A 31-year old women presented with excessive startle reflex and frequent falls. Her startle reflex is induced by slight stimuli which are not problematic in most people. Soon after her startle reflex is evoked, generalized muscle stiffness occurs. She becomes rigid and falls down without loss of consciousness. Because she cannot protect herself when she is startled and falls, she has repeatedly bruised her head and face. The pedigree includes her father and two sisters with similar symptoms. Gene analysis revealed GLRA1 mutation, and she was diagnosed with hereditary hyperekplexia (HPX). Symptoms improved with clonazepam 1 mg/day. HPX patients live with severe anxiety about frequent falls and sometimes suffer serious injury, such as cerebral concussion or bone fracture. Although HPX might sometimes be underestimated, accurate diagnosis is very important for effective treatment.

Hyperekplexia: A forgotten diagnosis clinched by next-generation sequencing.

Hyperekplexia is a rare early neonatal onset, potentially treatable, neurological disorder, characterized by a triad of immediate neonatal-onset stiffness, an exaggerated startle reflex in response to tactile or auditory stimuli followed by short periodical generalized stiffness. It is a monogenic genetically heterogeneous condition which can be potentially life threatening due to apneic episodes and is usually misdiagnosed as seizures. Here, we report two female siblings with hyperekplexia who were being treated by multiple antiepileptic medications for seizure-like episodes. Hyperekplexia was diagnosed by next-generation sequencing, which has emerged as a powerful diagnostic tool over the last few years.

Disruption of a Structurally Important Extracellular Element in the Glycine Receptor Leads to Decreased Synaptic Integration and Signaling Resulting in Severe Startle Disease.

Functional impairments or trafficking defects of inhibitory glycine receptors (GlyRs) have been linked to human hyperekplexia/startle disease and autism spectrum disorders. We found that a lack of synaptic integration of GlyRs, together with disrupted receptor function, is responsible for a lethal startle phenotype in a novel spontaneous mouse mutant shaky, caused by a missense mutation, Q177K, located in the extracellular ß8-ß9 loop of the GlyR α1 subunit. Recently, structural data provided evidence that the flexibility of the ß8-ß9 loop is crucial for conformational transitions during opening and closing of the ion channel and represents a novel allosteric binding site in Cys-loop receptors. We identified the underlying neuropathological mechanisms in male and female shaky mice through a combination of protein biochemistry, immunocytochemistry, and both in vivo and in vitro electrophysiology. Increased expression of the mutant GlyR α1Q177K subunit in vivo was not sufficient to compensate for a decrease in synaptic integration of α1Q177Kß GlyRs. The remaining synaptic heteromeric α1Q177Kß GlyRs had decreased current amplitudes with significantly faster decay times. This functional disruption reveals an important role for the GlyR α1 subunit ß8-ß9 loop in initiating rearrangements within the extracellular-transmembrane GlyR interface and that this structural element is vital for inhibitory GlyR function, signaling, and synaptic clustering.SIGNIFICANCE STATEMENT GlyR dysfunction underlies neuromotor deficits in startle disease and autism spectrum disorders. We describe an extracellular GlyR α1 subunit mutation (Q177K) in a novel mouse startle disease mutant shaky Structural data suggest that during signal transduction, large transitions of the ß8-ß9 loop occur in response to neurotransmitter binding. Disruption of the ß8-ß9 loop by the Q177K mutation results in a disruption of hydrogen bonds between Q177 and the ligand-binding residue R65. Functionally, the Q177K change resulted in decreased current amplitudes, altered desensitization decay time constants, and reduced GlyR clustering and synaptic strength. The GlyR ß8-ß9 loop is therefore an essential regulator of conformational rearrangements during ion channel opening and closing.

The astrocytic transporter SLC7A10 (Asc-1) mediates glycinergic inhibition of spinal cord motor neurons.

SLC7A10 (Asc-1) is a sodium-independent amino acid transporter known to facilitate transport of a number of amino acids including glycine, L-serine, L-alanine, and L-cysteine, as well as their D-enantiomers. It has been described as a neuronal transporter with a primary role related to modulation of excitatory glutamatergic neurotransmission. We find that SLC7A10 is substantially enriched in a subset of astrocytes of the caudal brain and spinal cord in a distribution corresponding with high densities of glycinergic inhibitory synapses. Accordingly, we find that spinal cord glycine levels are significantly reduced in Slc7a10-null mice and spontaneous glycinergic postsynaptic currents in motor neurons show substantially diminished amplitudes, demonstrating an essential role for SLC7A10 in glycinergic inhibitory function in the central nervous system. These observations establish the etiology of sustained myoclonus (sudden involuntary muscle movements) and early postnatal lethality characteristic of Slc7a10-null mice, and implicate SLC7A10 as a candidate gene and auto-antibody target in human hyperekplexia and stiff person syndrome, respectively.

Hyperekplexia, microcephaly and simplified gyral pattern caused by novel ASNS mutations, case report.

BACKGROUND: Asparagine synthetase deficiency (OMIM# 615574) is a very rare newly described neurometabolic disorder characterized by congenital microcephaly and severe global developmental delay, associated with intractable seizures or hyperekplexia. Brain MRI typically shows cerebral atrophy with simplified gyral pattern and delayed myelination. Only 12 cases have been described to date. The disease is caused by homozygous or compound heterozygous mutations in the ASNS gene on chromosome 7q21. CASE PRESENTATION: Family 1 is a multiplex consanguineous family with five affected members, while Family 2 is simplex. One affected from each family was available for detailed phenotyping. Both patients (Patients 1 and 2) presented at birth with microcephaly and severe hyperekplexia, and were found to have gross brain malformation characterized by simplified gyral pattern, and hypoplastic cerebellum and pons. EEG showed no epileptiform discharge in Patient 2 but multifocal discharges in patient 1. Patient 2 is currently four years old with severe neurodevelopmental delay, quadriplegia and cortical blindness. Whole exome sequencing (WES) revealed a novel homozygous mutation in ASNS (NM_001178076.1) in each patient (c.970C > T:p.(Arg324*) and c.944A > G:p.(Tyr315Cys)). CONCLUSION: Our results expand the mutational spectrum of the recently described asparagine synthetase deficiency and show a remarkable clinical homogeneity among affected individuals, which should facilitate its recognition and molecular confirmation for pertinent and timely genetic counseling.