Possible implication of undescribed SMN1-SMN2 genotype in chronic EMG-pattern of SMA with transitory acute denervation.

Spinal muscular atrophy (SMA) refers to a group of genetic neuromuscular disorders affecting lower motor neurons causative of numerous phenotypes. To date, according to the age of onset, maximum muscular activity achieved, and life expectation four types of SMA are recognized, all caused by mutations in the SMN1 gene with SMN2 copy number influencing disease severity. Herein, we describe the case of a 31-year-old young male with normal psychomotor development who has experienced fatigue, cramps, and muscle fasciculations in the lower limbs for a period of 2 months. Based on electrophysiological and clinical findings we performed SMA genetic, clinical exome and RNA expression of candidate genes which led us to suggest SMN1-SMN2 genes [(2+0) and (0+0)] combination as possibly being implicated in the phenotype.

Sox11 is required to maintain proper levels of Hedgehog signaling during vertebrate ocular morphogenesis.

Ocular coloboma is a sight-threatening malformation caused by failure of the choroid fissure to close during morphogenesis of the eye, and is frequently associated with additional anomalies, including microphthalmia and cataracts. Although Hedgehog signaling is known to play a critical role in choroid fissure closure, genetic regulation of this pathway remains poorly understood. Here, we show that the transcription factor Sox11 is required to maintain specific levels of Hedgehog signaling during ocular development. Sox11-deficient zebrafish embryos displayed delayed and abnormal lens formation, coloboma, and a specific reduction in rod photoreceptors, all of which could be rescued by treatment with the Hedgehog pathway inhibitor cyclopamine. We further demonstrate that the elevated Hedgehog signaling in Sox11-deficient zebrafish was caused by a large increase in shha transcription; indeed, suppressing Shha expression rescued the ocular phenotypes of sox11 morphants. Conversely, over-expression of sox11 induced cyclopia, a phenotype consistent with reduced levels of Sonic hedgehog. We screened DNA samples from 79 patients with microphthalmia, anophthalmia, or coloboma (MAC) and identified two novel heterozygous SOX11 variants in individuals with coloboma. In contrast to wild type human SOX11 mRNA, mRNA containing either variant failed to rescue the lens and coloboma phenotypes of Sox11-deficient zebrafish, and both exhibited significantly reduced transactivation ability in a luciferase reporter assay. Moreover, decreased gene dosage from a segmental deletion encompassing the SOX11 locus resulted in microphthalmia and related ocular phenotypes. Therefore, our study reveals a novel role for Sox11 in controlling Hedgehog signaling, and suggests that SOX11 variants contribute to pediatric eye disorders.

Dysregulation of group-I metabotropic glutamate (mGlu) receptor mediated signalling in disorders associated with Intellectual Disability and Autism.

Activation of group-I metabotropic glutamate receptors, mGlu1 and mGlu5, triggers a variety of signalling pathways in neurons and glial cells, which are differently implicated in synaptic plasticity. The earliest and much of key studies discovered abnormal mGlu5 receptor function in Fragile X syndrome (FXS) mouse models which then motivated more recent work that finds mGlu5 receptor dysfunction in related disorders such as intellectual disability (ID), obsessive-compulsive disorder (OCD) and autism. Therefore, mGlu1/5 receptor dysfunction may represent a common aetiology of these complex diseases. Furthermore, many studies have focused on dysregulation of mGlu5 signalling to synaptic protein synthesis. However, emerging evidence finds abnormal mGlu5 receptor interactions with its scaffolding proteins in FXS which results in mGlu5 receptor dysfunction and phenotypes independent of signalling to protein synthesis. Finally, both an increased and reduced mGlu5 functioning seem to be associated with ID and autism spectrum disorders, with important consequences for potential treatment of these developmental disorders.

The 9-bp deletion in region V of mtDNA: a risk factor of hearing loss and encephalomyopathy in Caucasian populations?

A deletion of one of the two copies of the 9-bp tandem repeat sequence (CCCCCTCTA), in the small non-coding/untranslated segment located between the cytochrome oxidase II and lysine tRNA genes of mitochondrial DNA (mtDNA), has previously been used as a polymorphic anthropological marker (MIC9D) for people of Africa and Asia, but it has been rarely reported in Europe. 32 Sicilian patients with syndromic hearing loss, negative for mutations in GJB2 and GJB6 genes, were tested for mtDNA known point mutations associated with syndromic or non-syndromic hearing loss by RFLP and/or direct sequencing. We identified the presence of the MIC9D in homoplasmy in lymphocytes and muscle of three subjects with sensorineural hearing loss and encephalomyopathy, two of these also presented moderate mental retardation. This deletion was absent in 300 Caucasian controls. Although further studies are warranted, our results suggest that the MIC9D polymorphism could have a susceptibility role in Caucasus, such as Sicily population.

Activation of 5-HT7 serotonin receptors reverses metabotropic glutamate receptor-mediated synaptic plasticity in wild-type and Fmr1 knockout mice, a model of Fragile X syndrome.

BACKGROUND: Fragile X syndrome (FXS) is a genetic cause of intellectual disability and autism. Fmr1 knockout (Fmr1 KO) mice, an animal model of FXS, exhibit spatial memory impairment and synapse malfunctioning in the hippocampus, with abnormal enhancement of long-term depression mediated by metabotropic glutamate receptors (mGluR-LTD). The neurotransmitter serotonin (5-HT) modulates hippocampal-dependent learning through serotonin 1A (5-HT1A) and serotonin 7 (5-HT7) receptors; the underlying mechanisms are unknown. METHODS: We used electrophysiology to test the effects of 5-HT on mGluR-LTD in wild-type and Fmr1 KO mice and immunocytochemistry and biotinylation assay to study related changes of 2-amino-3-(5-methyl-3-oxo-1,2-oxazol-4-yl)propanoic acid (AMPA) glutamate receptor surface expression. RESULTS: Application of 5-HT or 8-OH-DPAT (a mixed 5-HT1A/5-HT7 agonist) reversed mGluR-LTD in hippocampal slices. Reversal of mGluR-LTD by 8-OH-DPAT persisted in the presence of the 5-HT1A receptor antagonist WAY-100635, was abolished by SB-269970 (5-HT7 receptor antagonist), and was mimicked by LP-211, a novel selective 5-HT7 receptor agonist. Consistently, 8-OH-DPAT decreased mGluR-mediated reduction of AMPA glutamate receptor 2 (GluR2) subunit surface expression in hippocampal slices and cultured hippocampal neurons, an effect mimicked by LP-211 and blocked by SB-269970. In Fmr1 KO mice, mGluR-LTD was abnormally enhanced; similarly to wild-type, 8-OH-DPAT reversed mGluR-LTD and decreased mGluR-induced reduction of surface AMPA receptors, an effect antagonized by SB-269970. CONCLUSIONS: Serotonin 7 receptor activation reverses metabotropic glutamate receptor-induced AMPA receptor internalization and LTD both in wild-type and in Fmr1 KO mice, correcting excessive mGluR-LTD. Therefore, selective activation of 5-HT7 receptors may represent a novel strategy in the therapy of FXS.

Complex segmental duplications mediate a recurrent dup(X)(p11.22-p11.23) associated with mental retardation, speech delay, and EEG anomalies in males and females.

Submicroscopic copy-number variations make a considerable contribution to the genetic etiology of human disease. We have analyzed subjects with idiopathic mental retardation (MR) by using whole-genome oligonucleotide-based array comparative genomic hybridization (aCGH) and identified familial and de novo recurrent Xp11.22-p11.23 duplications in males and females with MR, speech delay, and a peculiar electroencephalographic (EEG) pattern in childhood. The size of the duplications ranges from 0.8-9.2 Mb. Most affected females show preferential activation of the duplicated X chromosome. Carriers of the smallest duplication show X-linked recessive inheritance. All other affected individuals present dominant expression and comparable clinical phenotypes irrespective of sex, duplication size, and X-inactivation pattern. The majority of the rearrangements are mediated by recombination between flanking complex segmental duplications. The identification of common clinical features, including the typical EEG pattern, predisposing genomic structure, and peculiar X-inactivation pattern, suggests that duplication of Xp11.22-p11.23 constitutes a previously undescribed syndrome.

Audiogenic seizure susceptibility is reduced in fragile X knockout mice after introduction of FMR1 transgenes.

The Fmr1 knockout (KO) mouse is characterized by an increased audiogenic seizure (AGS) susceptibility and is considered a good animal model for epilepsy and seizures in the human fragile-X (FRAX) syndrome. Here, we tested the hypothesis that the reintroduction of the FMR1 gene is able to revert the AGS susceptibility characterizing Fmr1 KO mice. To this aim, two groups of Fmr1 KO transgenic mice, which have additional copies of the human FMR1 gene (YAC) or FMR1 cDNA (G6) were used. AGS susceptibility of these mice was examined and compared to that of Fmr1 KO, wild type, and wild-type animals in whom the FMR1gene was also introduced (over-expressed). Mice were tested at different ages because AGS susceptibility is age dependent. The intensity of response was scored and the results were analyzed by means of 2-way analysis of variance to evaluate the effects of age and genetic condition. We found that AGS susceptibility rescue is complete in the G6 mice and partial in YAC mice. Our data indicate that the introduction of the human FMR1 gene in Fmr1 KO mice is able to revert the Fmr1 KO epileptic phenotype.

6q terminal deletion syndrome associated with a distinctive EEG and clinical pattern: a report of five cases.

PURPOSE: Mental retardation, facial dysmorphisms, and neurologic and brain abnormalities are features of 6q terminal deletions. Epilepsy is frequently associated with this chromosome abnormality, but electroclinical findings are not well delineated. We report five unrelated patients with 6q terminal deletions and a peculiar clinical, EEG, and neuroradiologic picture of epilepsy, mental retardation, and colpocephaly. METHODS: These three male and two female patients underwent general and neurologic examinations, repeated awake and sleep EEGs, and brain magnetic resonance imaging (MRI). A cytogenetic study and fluorescent in situ hybridization (FISH) with chromosome-specific subtelomeric probes were carried out in all cases. RESULTS: All subjects had seizures characterized by vomiting, cyanosis, and head and eye version, with and without loss of consciousness. In four cases, EEGs showed posterior spike-and-wave complexes, which were activated by sleep. No patient had status epilepticus or prolonged seizures. Brain MRI revealed colpocephaly and dysgenesis of the corpus callosum and brainstem in four patients; three of them also had hypertrophic massa intermedia. FISH analysis revealed a 6q terminal deletion in all patients, which ranged between 9 Mb (cases 2 and 3) and 16 Mb (case 4). CONCLUSIONS: We suggest that epilepsy associated with 6q terminal deletions is a new entity. Patients with dysmorphic features associated with focal occipital epilepsy, colpocephaly, and dysgenesis of the corpus callosum, thalami, and brainstem should be considered candidates for testing for 6q subtelomere deletions.

Sleep breathing and periodic leg movement pattern in Angelman Syndrome: a polysomnographic study.

OBJECTIVE: The aim of this study was to evaluate the sleep breathing patterns and to detect the eventual presence of periodic leg movements (PLMs) in patients affected by Angelman syndrome (AS). METHODS: Ten children with AS were recruited to participate in the study; the clinical diagnosis was confirmed by the genetic analysis (maternal 15q deletion, uniparental paternal disomy, or mutation of the UBE3A gene). All patients but two had presented epileptic seizures. Two age-matched groups of patients with mental retardation (MR) associated (MRE+) or not (MRE-) to epilepsy were used as control groups. All subjects underwent one polysomnographic recording, after one adaptation night. Sleep stages were scored according to standard criteria slightly modified in order to take into account the specific EEG patterns of AS, also the apnea/hypopnea index (AHI) was quantified; PLMs were identified and the PLM index (PLMI) was computed. The statistical analysis was carried out by means of the one-way ANOVA, followed by the Fisher LSD post-hoc test, when appropriate, and by means of the linear correlation coefficient between AHI and PLMI. RESULTS: Sleep macrostructure showed only few significant differences between children with AS and the other two groups of subjects: AS patients showed higher percentage of wakefulness after sleep onset and sleep onset latency; moreover, the percentage of REM sleep was reduced in AS and in MRE+ subjects. A tendency for AS subjects to present a higher PLMI than the other two groups was also found. AHI >5 was found in 30% of AS subjects, in 30.8% of MRE+, and only in 20% of MRE- patients (chi(2) = 2.359, NS); 70% of AS patients, 38.5% of MRE+, and 46.7% of MRE- subjects had PLMI >5 (chi(2) = 3.088, NS). CONCLUSIONS: These results confirm our previous questionnaire-based findings of a high prevalence of sleep breathing disorder and important PLMs in AS and allow us to hypothesize that epilepsy, rather than mental retardation, might exacerbate these sleep disorders. SIGNIFICANCE: Sleep breathing disorder and PLMs might contribute to the cognitive impairment and to the worsening of life quality of subjects with AS and with MR (mostly those with epilepsy). Therefore, our findings suggest the need to explore these sleep disorders in children affected by MR and to set up a correct treatment.

Different EEG frequency band synchronization during nocturnal frontal lobe seizures.

OBJECTIVE: In this article we describe the course of synchronization between different EEG channels during nocturnal seizures in one patient with nocturnal frontal lobe epilepsy (NFLE). METHODS: The functional interactions between the different EEG channels during the nocturnal seizures were analyzed by means of the so-called synchronization likelihood (SL). SL is a measure of the dynamical interdependencies between a time series (EEG channel) and one or more other time series. In contrast to coherence, SL measures linear as well as non-linear interdependencies and it can do so as a function of time, making it suitable for non-stationary time series. RESULTS: The main result of our single-patient study is the demonstration of a significant hyper-synchronization during NFLE seizures in the 8-12 Hz band which seems to be stopped by an increase in synchronization in the 0.5-4 Hz band, towards the end of each ictal episode. CONCLUSIONS: We suggest that a self-inhibiting complex mechanism might be responsible for the termination of ictal episodes which might take place at the level of the cortical layers and might involve mainly pyramidal neurons. SIGNIFICANCE: This study shows that advanced EEG analysis methods can help the current understanding of ictal manifestations of NFLE.

The mismatch negativity and the P3a components of the auditory event-related potentials in autistic low-functioning subjects.

OBJECTIVE: In order to understand better the psychophysiological basis of auditory processing abnormalities in autism, we decided to study two automatic components of the auditory event-related potentials (ERPs): the mismatch negativity (MMN)--a component of the ERP which is recorded when, during repetitive auditory stimulation, rare changes are introduced--and the novelty-related P3a which is recorded as a response to unexpected novel events occurring in a sequence of repetitive stimuli. METHODS: Ten male subjects, mean age 12.3 years (SD 4.95), affected by autism and mental retardation were admitted to this study. All patients were also mentally retarded. Ten normal male subjects, mean age 12.2 years (SD 3.94), were used as controls. Auditory evoked potentials were recorded from 19 scalp electrodes (10-20 system), and stimuli were presented in sequences consisting of 2000 tones (70 dB, ISI=800 ms). Three types of stimuli were presented: (1) standard stimuli (1000 Hz tones, 80% of total stimuli), (2) deviant stimuli (1300 Hz tones, 10% of total stimuli), and (3) novel stimuli (complex and non-monotonal, 10% of total stimuli). To quantify the MMN, the evoked response to the standard tones was subtracted from the corresponding deviant stimulus response and its amplitude and latency at peak were measured over Fz, Cz and Pz; similarly, the P3a component of the ERP was obtained by subtracting the response to the standard tone from that to the novel stimuli and its amplitude and latency at peak were measured over Fz, Cz and Pz. Also, the amplitude and latency at peak for the N1 component of the auditory evoked potential obtained with the standard stimuli were measured over Fz, Cz and Pz. The correlation between age and MMN and P3a amplitude was also analyzed. RESULTS: N1 showed significantly shorter latencies in the autistic groups. MMN elicited by deviant stimuli, but not that elicited by novel stimuli, was found to be significantly larger in autistic children than in normal controls. P3a showed higher amplitude in autistic subjects than in normal controls during childhood; the opposite was observed during young adulthood. DISCUSSION: Our findings indicate that significant changes in ERPs can also be seen in non-cooperative individuals with autism and mental retardation, which might be different from the changes already reported for high-functioning autistic subjects and deserve further insight. These changes show developmental modifications that should be taken into consideration when analyzing data from autistic subjects.

Nonlinear EEG analysis during sleep in premature and full-term newborns.

OBJECTIVE: Recently it has been shown that the adult sleep EEG is mostly determined by high-dimensional, linear dynamics with the exception of the A phase of the cyclic alternating pattern which displays more synchronized nonlinear dynamics. It is not known how these two different types of brain dynamics develop in early life; for this reason the aim of this study was that to extend the nonlinear analysis to the EEG during sleep recorded in premature and full-term newborns. METHODS: EEG epochs were chosen from a total of 24 polygraphic recordings from 14 babies (9 males and 5 females) aged between 33 weeks 3 days and 4 months conceptional age. All subjects were neurologically normal and showed normal psychomotor development at follow-up. A total of 243 artifact-free epochs was chosen during active sleep (AS, 74 total epochs), quiet sleep (QS, 76 total epochs) and indeterminate sleep (IS, 93 total epochs). The dynamic properties of the EEG were assessed by means of the nonlinear cross prediction test which uses 3 different 'model' time series in order to predict nonlinearly the original data set (Pred, Ama, and Tir). Pred is a measure of the predictability of the time series, and Ama and Tir are measures of asymmetry, indicating nonlinear structure. RESULTS: Our results show that the structure of sleep EEG in newborns is significantly different from that of adults, it cannot be distinguished from that of high-dimensional noise in the majority of epochs, and shows a tendency to become nonlinear in nature, mostly during QS, in a small percentage of the epochs analyzed. CONCLUSIONS: These findings can be interpreted as the effect of immature synaptic interconnections between neurons in the newborn brain.

Peripheral markers of the gamma-aminobutyric acid (GABA)ergic system in Angelman's syndrome.

It has recently been demonstrated that patients with Angelman's syndrome who exhibited a deletion on cytogenetic tests show more severe clinical pictures with drug-resistant epilepsy than patients with Angelman's syndrome not carrying the deletion. To verify if this difference in clinical severity can be attributed to genes for the three gamma-aminobutyric acid (GABA)A receptor subunits (GABRB3, GABRA5, GABRG3) located in the deleted region, a possible modification of peripheral markers of the GABAergic system was investigated in 12 subjects with Angelman's syndrome and 20 age-matched subjects (8 with idiopathic epilepsy and 12 not affected by neurologic diseases). The results confirmed a more severe clinical picture, and epilepsy syndrome in particular, in Angelman's syndrome patients with deletions versus patients without deletions. In contrast, biochemical study (based on dosage of plasma levels of GABA and diazepam binding inhibitor, an endogenous ligand of GABAA and peripheral benzodiazepine receptors, showed contradictory results: patients with Angelman's syndrome showed significantly higher levels of GABA and diazepam binding inhibitor than patients without neurologic impairment but significantly lower levels than epileptic controls.

Heart rate variability during sleep in children with partial epilepsy.

Alterations in autonomic control of cardiac activity in epileptic patients have been reported by several studies in the past, and both ictal and interictal modifications of heart rate regulation have been described. Alterations of autonomic control of cardiac activity can play an important role in sudden unexplained death in patients with epilepsy (SUDEP). However, the presence of specific changes in heart rate variability (HRV) during sleep, not correlated with seizures, has not been assessed in children with epilepsy; for this reason, we evaluated features of cardiac autonomic function during sleep without ictal epileptiform electroencephalogram (EEG) activity in a group of children with partial epilepsy. Eleven patients (five males and six females; mean age 11.5 years, SD: 3.65 years) affected by partial epilepsy were admitted to this study; 11 normal subjects (five males and six females; mean age 12.9 years, SD: 2.72 years) served as a control group. All subjects slept in the laboratory for two consecutive nights. The data were analyzed during the second night. Sleep was polygraphically recorded [including one electrocardiography (ECG) channel] and signals were digitally stored. A series of 5-min ECG epochs were chosen from each sleep stage, during periods without evident ictal epileptiform activity in the EEG. Electrocardiography signals were analyzed for automatic detection of R-waves and, subsequently, a series of time- and frequency-domain measures were calculated. Epileptic subjects tended to show an overall lower HRV in both time- and frequency-domain parameters, principally during rapid eye movement (REM) sleep and, to a lesser extent, during sleep stage 2. Among the different bands, this decrease was most evident for the high-frequency band (HF) absolute power. For this reason, the ratio between the low-frequency band (LF) and HF was always higher in epileptic patients than in normal controls and the difference was statistically significant during sleep stages 3 and/or 4 and REM sleep. Our results indicate that during sleep, a particular condition of basal modification in autonomic characteristics occurs (mostly during REM sleep) in partial epilepsy patients. This finding might represent an important factor contributing to the complex mechanism of SUDEP which takes place most often during sleep and supports the need of studying HRV specifically during this state in subjects with seizures.

Non-linear EEG measures during sleep: effects of the different sleep stages and cyclic alternating pattern.

The objective of this work was to study the non-linear aspects of sleep EEG, taking into account the different sleep stages and the peculiar organization of its phasic events in ordered sequences (CAP) by applying a series of new non-linear measures (non-linear cross prediction or NLCP), which appear more reliable for the detection and characterization of non-linear structures in experimental data than the commonly used correlation dimension. Eight healthy subjects aged 18-20 years participated in this study. Polysomnography was performed in all subjects; signals were sampled at 128 Hz and stored on hard disk. The C3 or C4 derivation was used for all the subsequent computational steps, which were performed on EEG epochs (4096 data points) selected from sleep stage 2 (S2) and slow-wave sleep (SWS), in both CAP and non-CAP (NCAP) conditions. Also, epochs from sleep stage 1 (S1), REM and wakefulness preceding sleep were recorded. The dynamic properties of the EEG were assessed by means of the non-linear cross-prediction test, which uses three different 'model' time series in order to predict non-linearly the original data set (Pred, Ama, and Tir). Pred is a measure of the predictability of the time series, and Ama and Tir are measures of asymmetry, indicating non-linear structure. The non-linear measures applied in this study indicate that sleep EEG tends to show non-linear structure only during CAP periods, both during S2 and SWS. Moreover, during CAP periods, non-linearity can only be detected during the phase A1 subtypes (and partially A2) of CAP. The A3 phases show characteristics of non-stationarity and bear some resemblance to wakefulness. Based on the results of this study, sleep might be considered as a dynamically evolving sequence of different states of the EEG, which we could track by detecting non-linearity, mostly in association with CAP. Our results clearly show that detectable non-linearity in the EEG is closely related to the occurrence of the phase A of CAP.