Periventricular nodular heterotopia (PNH) is a common type of heterotopia usually characterized by epilepsy. Previous studies have identified alterations in structural and functional connectivity related to this disorder, but its local functional neural basis has received less attention. The purpose of this study was to combine univariate analysis and a Gaussian process classifier (GPC) to assess local activity and further explore neuropathological mechanisms in PNH-related epilepsy.

We aimed to delineate the distribution of periventricular nodular heterotopia (PNH) in patients with 22q11.2 microdeletion syndrome (22q11.2DS) and place this in the context of other genetic forms of PNH.

Pathogenic FLNA variants can be identified in patients with seizures accompanied by periventricular nodular heterotopia (PVNH). It is unusual to find FLNA aberrations in epileptic patients without PVNH on brain imaging. We report a boy with cryptogenic West syndrome followed by refractory seizures and psychomotor delay. We performed whole-exome sequencing and identified a de novo missense variant in FLNA. It is noteworthy that this patient showed no PVNH. As no other pathogenic variants were found in epilepsy-related genes, this FLNA variant likely caused West syndrome but with no PVNH.

We investigated systematic differences in corpus callosum morphology in periventricular nodular heterotopia (PVNH). Differences in corpus callosum mid-sagittal area and subregional area changes were measured using an automated software-based method. Heterotopic gray matter deposits were automatically labeled and compared with corpus callosum changes. The spatial pattern of corpus callosum changes were interpreted in the context of the characteristic anterior-posterior development of the corpus callosum in healthy individuals. Individuals with periventricular nodular heterotopia were imaged at the Melbourne Brain Center or as part of the multi-site Epilepsy Phenome Genome project. Whole brain T1 weighted MRI was acquired in cases (n=48) and controls (n=663). The corpus callosum was segmented on the mid-sagittal plane using the software "yuki". Heterotopic gray matter and intracranial brain volume was measured using Freesurfer. Differences in corpus callosum area and subregional areas were assessed, as well as the relationship between corpus callosum area and heterotopic GM volume. The anterior-posterior distribution of corpus callosum changes and heterotopic GM nodules were quantified using a novel metric and compared with each other. Corpus callosum area was reduced by 14% in PVNH (p=1.59×10(-9)). The magnitude of the effect was least in the genu (7% reduction) and greatest in the isthmus and splenium (26% reduction). Individuals with higher heterotopic GM volume had a smaller corpus callosum. Heterotopic GM volume was highest in posterior brain regions, however there was no linear relationship between the anterior-posterior position of corpus callosum changes and PVNH nodules. Reduced corpus callosum area is strongly associated with PVNH, and is probably associated with abnormal brain development in this neurological disorder. The primarily posterior corpus callosum changes may inform our understanding of the etiology of PVNH. Our results suggest that interhemispheric pathways are affected in PVNH.

Periventricular nodular heterotopia (PNH) is a neural migration disorder which often presents clinically with seizures. However, the underlying functional neural basis of PNH is still unclear. We aimed to explore the underlying pathological mechanism of PNH by combining both whole brain functional connectivity (FC) and seed-based FC analyses. We utilized resting-state fMRI to measure functional connectivity strength (FCS) in 38 patients with PNH-related epilepsy and 38 control subjects. The regions with FCS alterations were selected as seeds in the following FC analyses. Pearson correlation analyses were performed to explore associations between these functional neural correlates and clinical features. In comparison with controls, PNH patients showed lower FCS in bilateral insula (P < 0.05, family wise error (FWE) correction), higher FC in the default mode network and lower FC in the fronto-limbic-cerebellar circuits (P < 0.05, FWE correction). Pearson correlation analyses revealed that FCS in bilateral insula was negatively correlated with the epilepsy duration (P < 0.05); medial prefronto-insular connectivity was negatively correlated with Hamilton Anxiety Scale (P < 0.05) and cerebellar-insular connectivity was also negatively correlated with Hamilton Depression Scale (P < 0.05). Using the resting-state FCS analytical approach, we identified significant insular hypoactivation in PNH patients, which suggests that the insula might represent the cortical hub of the whole-brain networks in this condition. Additionally, disruption of resting state FC in large-scale neural networks pointed to a connectivity-based neuropathological process in PNH.

Periventricular nodular heterotopia (PVNH) is a malformation of cortical development commonly associated with epilepsy. We exome sequenced 202 individuals with sporadic PVNH to identify novel genetic risk loci. We first performed a trio-based analysis and identified 219 de novo variants. Although no novel genes were implicated in this initial analysis, PVNH cases were found overall to have a significant excess of nonsynonymous de novo variants in intolerant genes (p = 3.27x10-7), suggesting a role for rare new alleles in genes yet to be associated with the condition. Using a gene-level collapsing analysis comparing cases and controls, we identified a genome-wide significant signal driven by four ultra-rare loss-of-function heterozygous variants in MAP1B, including one de novo variant. In at least one instance, the MAP1B variant was inherited from a parent with previously undiagnosed PVNH. The PVNH was frontally predominant and associated with perisylvian polymicrogyria. These results implicate MAP1B in PVNH. More broadly, our findings suggest that detrimental mutations likely arising in immediately preceding generations with incomplete penetrance may also be responsible for some apparently sporadic diseases.

Periventricular nodular heterotopia (PNH) is a rare malformation of cortical development often associated with drug resistant focal onset epilepsy. The link between nodules and neocortex have been demonstrated with depth electrodes investigations showing that seizures may arise from both structures. In the last years fMRI resting-state (fMRI-RS) have received a surge in interest due to its capability to track non-invasively physiological and pathological relevant differences in brain network organization. We performed a cerebro-cerebellar voxel-wise and region-of-interest resting state fMRI (RS-fMRI) functional connectivity analysis in a seizure-free epilepsy patient with a PNH in the right temporal horn. Our finding confirms a spontaneous synchronization between PNH and its surrounding cortex, specifically in the inferior temporal, fusiform and occipital gyrus. We also found a significant connectivity with bilateral cerebellum, more intense and widespread on the PNH cerebellar contralateral lobule. RS-fMRI confirmed its potential as a promising tool for non-invasive mapping of cortical and subcortical brain functional organization.

Periventricular nodular heterotopia (PVNH) is caused by abnormal neuronal migration, resulting in the neurons accumulate as nodules along the surface of the lateral ventricles. PVNH often cause epilepsy, psychomotor development or cognition problem. Mutations in FLNA (Filamin A) is the most common underlying genetic etiology. Our purpose is to delineate the clinical and imaging spectrum that differentiates FLNA-positive and FLNA-negative PVNH patients.

Periventricular nodular heterotopias (PVNH) are areas of neurons abnormally located in the white matter that might be involved in physiological cortical functions. Autoscopic hallucinations are changes in self-consciousness determined by a mismatch in integration of multiple sensory inputs. Our goal is to highlight the brain network involved in generation of autoscopic hallucination elicited by electrical stimulation of a PVNH in a drug resistant epilepsy patient. Our patient was explored using stereo-electroencephalography with electrodes covering the right posterior temporal PVNH and the adjacent cortex. Direct electrical high frequency stimulation of the PVNH elicited autoscopic hallucinations mainly involving the face and upper trunk. We then used multiple modalities to determine brain connectivity: single pulse electrical stimulation of the PVNH and stimulation-evoked potentials were used to highlight resting state effective connectivity. High-frequency stimulation using alternating polarity pulses enabled us to identify the network involved, time-locked to the clinical effect and to map symptom-related effective connectivity. Functional connectivity using a non-linear regression method was used to determine dependencies between different cortical regions following the stimulation. Finally, structural connectivity was highlighted using deterministic fiber tracking. Multi-modal connectivity analysis identified a network involving the PVNH, occipital and temporal neocortex, fusiform gyrus and parietal cortex.

To investigate the quantitative diffusion properties of the corpus callosum (CC) in a large group of patients with periventricular nodular heterotopia (PNH) related epilepsy and to further investigate the effect of Filamin A (FLNA) mutation on these properties.

Periventricular nodular heterotopia (PNH) is a common structural malformation of cortical development. Mutations in the filamin A gene are frequent in familial cases with X-linked PNH. However, many cases with sporadic PNH remain genetically unexplained. Although medically refractory epilepsy often brings attention to the underlying PNH, patients are often not candidates for surgical resection. This limits access to neuronal tissue harboring causal mutations. We evaluated a patient with PNH and medically refractory focal epilepsy who underwent a presurgical evaluation with stereotactically placed electroencephalographic (SEEG) depth electrodes. Following SEEG explantation, we collected trace tissue adherent to the electrodes and extracted the DNA. Whole-exome sequencing performed in a Clinical Laboratory Improvement Amendments-approved genetic diagnostic laboratory uncovered a de novo heterozygous pathogenic variant in novel candidate PNH gene MEN1 (multiple endocrine neoplasia type 1; c.1546dupC, p.R516PfsX15). The variant was absent in an earlier exome profiling of the venous blood-derived DNA. The MEN1 gene encodes the ubiquitously expressed, nuclear scaffold protein menin, a known tumor suppressor gene with an established role in the regulation of transcription, proliferation, differentiation, and genomic integrity. Our study contributes a novel candidate gene in PNH generation and a novel practical approach that integrates electrophysiological and genetic explorations of epilepsy.

Defects in neuronal migration cause brain malformations, which are associated with intellectual disability (ID) and epilepsy. Using exome sequencing, we identified compound heterozygous variants (p.Arg71His and p. Leu729ThrfsTer6) in TMTC3, encoding transmembrane and tetratricopeptide repeat containing 3, in four siblings with nocturnal seizures and ID. Three of the four siblings have periventricular nodular heterotopia (PVNH), a common brain malformation caused by failure of neurons to migrate from the ventricular zone to the cortex. Expression analysis using patient-derived cells confirmed reduced TMTC3 transcript levels and loss of the TMTC3 protein compared to parental and control cells. As TMTC3 function is currently unexplored in the brain, we gathered support for a neurobiological role for TMTC3 by generating flies with post-mitotic neuron-specific knockdown of the highly conserved Drosophila melanogaster TMTC3 ortholog, CG4050/tmtc3. Neuron-specific knockdown of tmtc3 in flies resulted in increased susceptibility to induced seizures. Importantly, this phenotype was rescued by neuron-specific expression of human TMTC3, suggesting a role for TMTC3 in seizure biology. In addition, we observed co-localization of TMTC3 in the rat brain with vesicular GABA transporter (VGAT), a presynaptic marker for inhibitory synapses. TMTC3 is localized at VGAT positive pre-synaptic terminals and boutons in the rat hypothalamus and piriform cortex, suggesting a role for TMTC3 in the regulation of GABAergic inhibitory synapses. TMTC3 did not co-localize with Vglut2, a presynaptic marker for excitatory neurons. Our data identified TMTC3 as a synaptic protein that is involved in PVNH with ID and epilepsy, in addition to its previously described association with cobblestone lissencephaly.

Neurodevelopmental disorders associated with periventricular nodular heterotopia (PVNH) are characterized by phenotypic and genetic heterogeneity. NEDD4L mutation can lead to PVNH7. However, at present, only eight NEDD4L pathogenic variants have been identified across 15 cases of PVNH7 worldwide. Given this dearth of evidence, the precise correlations between genetic pathogenesis and phenotypes remain to be determined.

Neurodevelopmental disorders with periventricular nodular heterotopia (PNH) are etiologically heterogeneous, and their genetic causes remain in many cases unknown. Here we show that missense mutations in NEDD4L mapping to the HECT domain of the encoded E3 ubiquitin ligase lead to PNH associated with toe syndactyly, cleft palate and neurodevelopmental delay. Cellular and expression data showed sensitivity of PNH-associated mutants to proteasome degradation. Moreover, an in utero electroporation approach showed that PNH-related mutants and excess wild-type NEDD4L affect neurogenesis, neuronal positioning and terminal translocation. Further investigations, including rapamycin-based experiments, found differential deregulation of pathways involved. Excess wild-type NEDD4L leads to disruption of Dab1 and mTORC1 pathways, while PNH-related mutations are associated with deregulation of mTORC1 and AKT activities. Altogether, these data provide insights into the critical role of NEDD4L in the regulation of mTOR pathways and their contributions in cortical development.

Epilepsy associated with periventricular nodular heterotopia (PNH) is characterized by complex relationships between the heterotopic and the normotopic cortex during the interictal state and at seizure onset. High-frequency oscillations (HFO) have been proposed as a marker of epileptogenicity that might reflect disease activity. The effects of thermocoagulations on epileptogenicity in this context remain unknown. We aimed to investigate the interictal HFO- and spike profiles of different cortical structures before and after two consecutive SEEG-guided thermocoagulations, in correlation with seizure outcome, in a patient with PNH-related drug-resistant epilepsy.

Pathogenic variants of the X-linked FLNA gene encoding filamin A protein have been associated with a wide spectrum of symptoms, including the recently described pulmonary phenotype with childhood-onset panlobular emphysema. We describe three female patients from two families with novel heterozygous FLNA variants c.5837_2del and c.508C > T. Analysis of immunofluorescence of peripheral blood smears and platelet function was performed for all patients. FLNA-negative platelets were observed, suggesting that these variants result in the loss of a functional protein product. All three patients also had periventricular nodular heterotopia and panlobular emphysema. However, they had considerably milder symptoms and later age of onset than in the previously reported cases. Therefore, patients with pathogenic FLNA variants should be studied actively for lung involvement even in the absence of pronounced respiratory symptoms. Conversely, any patient with unexplained panlobular emphysema should be analyzed for pathogenic FLNA variants. We also suggest that immunofluorescence analysis is a useful tool for investigating the pathogenicity of novel FLNA variants.

Kabuki syndrome is a heterogeneous condition characterized by distinctive facial features, intellectual disability, growth retardation, skeletal abnormalities and a range of organ malformations. Although at least two major causative genes have been identified, these do not explain all cases. Here we describe a patient with a complex Kabuki-like syndrome that included nodular heterotopia, in whom testing for several single-gene disorders had proved negative. Exome sequencing uncovered a de novo c.931_932insTT variant in HNRNPK (heterogeneous nuclear ribonucleoprotein K). Although this variant was identified in March 2012, its clinical relevance could only be confirmed following the August 2015 publication of two cases with HNRNPK mutations and an overlapping phenotype that included intellectual disability, distinctive facial dysmorphism and skeletal/connective tissue abnormalities. Whilst we had attempted (unsuccessfully) to identify additional cases through existing collaborators, the two published cases were 'matched' using GeneMatcher, a web-based tool for connecting researchers and clinicians working on identical genes. Our report therefore exemplifies the importance of such online tools in clinical genetics research and the benefits of periodically reviewing cases with variants of unproven significance. Our study also suggests that loss of function variants in HNRNPK should be considered as a molecular basis for patients with Kabuki-like syndrome.

Progressive lung involvement in Filamin A (FLNA)-related cerebral periventricular nodular heterotopia (PVNH) has been reported in a limited number of cases.

We are currently investigating various treatments which could determine, in the rat brain, structural abnormalities mimicking those reported in human brain dysgeneses. We can induce the formation of neuronal heterotopia in the progeny of rats by means of a double injection of the cytotoxic agent methylazoxymethanol acetate (MAM) on embryonic day 15. We have now investigated the anatomical connections of these heterotopia by means of anterograde and retrograde tract tracing techniques. The induced heterotopia along the border of the lateral ventricles shared common anatomical features with the periventricular nodules in human periventricular or subcortical nodular heterotopia (PNH). The tract tracing data demonstrated the existence of reciprocal connections between the neuronal heterotopia and the ipsilateral and contralateral cortical areas, and the presence of abnormal cortico-hippocampal and cortico-cortical connections. On the basis of the connectivity patterns, it may be speculated that some cells in the heterotopia could be neurons originally committed to the cortex, that were interrupted in their migration by the MAM treatment. Given the common morphological features seen in human PNH and MAM-induced brain heterotopia, the anatomical and developmental analysis of MAM-treated rats may shed light on the mechanisms by which human brain dysgeneses develop in human patients.

The mammalian neocortex has undergone remarkable changes through evolution. A consequence of such rapid evolutionary events could be a trade-off that has rendered the brain susceptible to certain neurodevelopmental and neuropsychiatric conditions. We analyzed the exomes of 65 patients with the structural brain malformation periventricular nodular heterotopia (PH). De novo coding variants were observed in excess in genes defining a transcriptomic signature of basal radial glia, a cell type linked to brain evolution. In addition, we located two variants in human isoforms of two genes that have no ortholog in mice. Modulating the levels of one of these isoforms for the gene PLEKHG6 demonstrated its role in regulating neuroprogenitor differentiation and neuronal migration via RhoA, with phenotypic recapitulation of PH in human cerebral organoids. This suggests that this PLEKHG6 isoform is an example of a primate-specific genomic element supporting brain development.