Biological Networks and Complexity in Early-Onset Motor Neuron Diseases

Motor neuron diseases (MNDs) are neuromuscular disorders where the spinal motor neurons–either the cell bodies themselves or their axons–are the primary cells affected. To date, there are 120 different genes that are lost or mutated in pediatric-onset MNDs. Most of these childhood-onset disorders, aside from spinal muscular atrophy (SMA), lack viable therapeutic options. Previous research on MNDs has focused on understanding the pathobiology of a single, specific gene mutation and targeting therapies to that pathobiology. This reductionist approach has yielded therapeutic options for a specific disorder, in this case SMA. Unfortunately, therapies specific for SMA have not been effective against other pediatric-onset MNDs. Pursuing the same approach for the other defined MNDs would require development of at least 120 independent treatments raising feasibility issues. We propose an alternative to this this type of reductionist approach by conceptualizing MNDs in a complex adaptive systems framework that will allow identification of common molecular and cellular pathways which form biological networks that are adversely affected in early-onset MNDs and thus MNDs with similar phenotypes despite diverse genotypes. This systems biology approach highlights the complexity and self-organization of the motor system as well as the ways in which it can be affected by these genetic disorders. Using this integrated approach to understand early-onset MNDs, we would be better poised to expand the therapeutic repertoire for multiple MNDs

ACTH Prevents Deficits in Fear Extinction Associated with Early Life Seizures

Objective: Early life seizures (ELS) are often associated with cognitive and psychiatric comorbidities that are detrimental to quality of life. In a rat model of ELS, we explored long-term cognitive outcomes in adult rats. Using ACTH, an endogeneous HPA-axis hormone given to children with severe epilepsy, we sought to prevent cognitive deficits. Through comparisons with dexamethasone, we sought to dissociate the corticosteroid effects of ACTH from other potential mechanisms of action. Results: Although rats with a history of ELS were able to acquire a conditioned fear learning paradigm and controls, these rats had significant deficits in their ability to extinguish fearful memories. ACTH treatment did not alter any seizure parameters but nevertheless was able to significantly improve this fear extinction, while dexamethasone treatment during the same period did not. This ACTH effect was specific for fear extinction deficits and not for spatial learning deficits in a water maze. Additionally, ACTH did not alter seizure latency or duration suggesting that cognitive and seizure outcomes may be dissociable. Expression levels of melanocortin receptors, which bind ACTH, were found to be significantly lower in animals that had experienced ELS than in control animals, potentially implicating central melanocortin receptor dysregulation in the effects of ELS, and suggesting a mechanism of action for ACTH. Interpretation: Taken together, these data suggest that early treatment with ACTH can have significant long-term consequences for cognition in animals with a history of ELS independently of seizure cessation and may act in part through a CNS melanocortin receptor pathway

Differential regulation of gene expression pathways with dexamethasone and ACTH after early life seizures.

Early-life seizures (ELS) are associated with persistent cognitive deficits such as ADHD and memory impairment. These co-morbidities have a dramatic negative impact on the quality of life of patients. Therapies that improve cognitive outcomes have enormous potential to improve patients\u27 quality of life. Our previous work in a rat flurothyl-induction model showed that administration of adrenocorticotropic hormone (ACTH) at time of seizure induction led to improved learning and memory in the animals despite no effect on seizure latency or duration. Administration of dexamethasone (Dex), a corticosteroid, did not have the same positive effect on learning and memory and has even been shown to exacerbate injury in a rat model of temporal lobe epilepsy. We hypothesized that ACTH exerted positive effects on cognitive outcomes through beneficial changes to gene expression and proposed that administration of ACTH at seizure induction would return gene-expression in the brain towards the normal pattern of expression in the Control animals whereas Dex would not. Twenty-six Sprague-Dawley rats were randomized into vehicle- Control, and ACTH-, Dex-, and vehicle- ELS. Rat pups were subjected to 60 flurothyl seizures from P5 to P14. After seizure induction, brains were removed and the hippocampus and PFC were dissected, RNA was extracted and sequenced, and differential expression analysis was performed using generalized estimating equations. Differential expression analysis showed that ACTH pushes gene expression in the brain back to a more normal state of expression through enrichment of pathways involved in supporting homeostatic balance and down-regulating pathways that might contribute to excitotoxic cell-damage post-ELS

Short Duration Waveforms Recorded Extracellularly from Freely Moving Rats are Representative of Axonal Activity

While extracellular somatic action potentials from freely moving rats have been well characterized, axonal activity has not. We report direct extracellular tetrode recordings of putative axons whose principal feature is a short duration waveform (SDW) with an average peak-trough length less than 179 μs. While SDW recordings using tetrodes have previously been treated as questionable or classified as cells, we hypothesize that they are representative of axonal activity. These waveforms have significantly shorter duration than somatic action potentials, are triphasic and are therefore similar to classic descriptions of microelectrode recordings in white matter and of in vitro action potential propagation along axons. We describe SDWs recorded from pure white-matter tracts including the alveus and corpus callosum. Recordings of several SDWs in the alveus exhibit grid-like firing patterns suggesting these axons carry spatial information from entorhinal cortical neurons. Finally, we locally injected the GABAA agonist Muscimol into layer CA1 of the hippocampus while simultaneously recording somatic activity and SDWs on the same tetrodes. The persistent activity of SDWs during Muscimol inactivation of somatic action potentials indicates that SDWs are representative of action potential propagation along axons projecting from more distal somata. This characterization is important as it illustrates the dangers of exclusively using spike duration as the sole determinant of unit type, particularly in the case of interneurons whose peak-trough times overlap with SDWs. It may also allow future studies to explore how axonal projections from disparate brain regions integrate spatial information in the hippocampus, and provide a basis for studying the effects of pharmaceutical agents on signal transmission in axons, and ultimately to aid in defining the potential role of axons in cognition

Oscillation Phase Locking and Late ERP Components of Intracranial Hippocampal Recordings Correlate to Patient Performance in a Working Memory Task

In working memory tasks, stimulus presentation induces a resetting of intracranial temporal lobe oscillations in multiple frequency bands. To further understand the functional relevance of this phenomenon, we investigated whether working memory performance depends on the phase precision of ongoing oscillations in the hippocampus. We recorded intra-hippocampal local field potentials in individuals performing a working memory task. Two types of trials were administered. For high memory trials presentation of a list of four letters ( List ) was followed by a single letter memory probe ( Test ). Low memory load trials, consisting of four identical letters (AAAA) followed by a probe with the same letter (A), were interspersed. Significant phase locking of ongoing oscillations across trials, estimated by the Pairwise Phase Consistency Index (PPCI) was observed in delta (0.5-4 Hz), theta (5-7 Hz), and alpha (8-12 Hz) bands during stimulus presentation and recall but was increased in low memory load trials. Across patients however, higher delta PPCIs during recall in the left hippocampus were associated with faster reaction times. Because phase locking could also be interpreted as a consequence of a stimulus evoked potential, we performed event related potential analysis (ERP) and examined the relationship of ERP components with performance. We found that both amplitude and latency of late ERP components correlated with both reaction time and accuracy. We propose that, in the Sternberg task, phase locking of oscillations, or alternatively its ERP correlate, synchronizes networks within the hippocampus and connected structures that are involved in working memory

Next-generation sequencing reveals broad down-regulation of microRNAs in secondary progressive multiple sclerosis CD4+ T cells

Background Immunoactivation is less evident in secondary progressive MS (SPMS) compared to relapsing-remitting disease. MicroRNA (miRNA) expression is integral to the regulation of gene expression; determining their impact on immune-related cell functions, especially CD4+ T cells, during disease progression will advance our understanding of MS pathophysiology. This study aimed to compare miRNA profiles of CD4+ T cells from SPMS patients to healthy controls (HC) using whole miRNA transcriptome next-generation sequencing (NGS). Total RNA was extracted from CD4+ T cells and miRNA expression patterns analyzed using Illumina-based small-RNA NGS in 12 SPMS and 12 HC samples. Results were validated in a further cohort of 12 SPMS and 10 HC by reverse transcription quantitative polymerase chain reaction (RT-qPCR). Results The ten most dysregulated miRNAs identified by NGS were selected for qPCR confirmation; five (miR-21-5p, miR-26b-5p, miR-29b-3p, miR-142-3p, and miR-155-5p) were confirmed to be down-regulated in SPMS (p < 0.05). SOCS6 is targeted by eight of these ten miRNAs. Consistent with this, SOCS6 expression is up-regulated in SPMS CD4+ T cells (p < 0.05). This is of particular interest as SOCS6 has previously been shown to act as a negative regulator of T cell activation. Conclusions Ninety-seven percent of miRNA candidates identified by NGS were down-regulated in SPMS. The down-regulation of miRNAs and increased expression of SOCS6 in SPMS CD4+ T cells may contribute to reduced immune system activity in progressive MS

Enrichment and Training Improve Cognition in Rats with Cortical Malformations

Children with malformations of cortical development (MCD) frequently have associated cognitive impairments which reduce quality of life. We hypothesized that cognitive deficits associated with MCD can be improved with environmental manipulation or additional training. The E17 methylazoxymethanol acetate (MAM) exposure model bears many anatomical hallmarks seen in human MCDs as well as similar behavioral and cognitive deficits. We divided control and MAM exposed Sprague-Dawley rats into enriched and non-enriched groups and tested performance in the Morris water maze. Another group similarly divided underwent sociability testing and also underwent Magnetic Resonance Imaging (MRI) scans pre and post enrichment. A third group of control and MAM rats without enrichment were trained until they reached criterion on the place avoidance task. MAM rats had impaired performance on spatial tasks and enrichment improved performance of both control and MAM animals. Although MAM rats did not have a deficit in sociability they showed similar improvement with enrichment as controls. MRI revealed a whole brain volume decrease with MAM exposure, and an increase in both MAM and control enriched volumes in comparison to non-enriched animals. In the place avoidance task, MAM rats required approximately 3 times as long to reach criterion as control animals, but with additional training were able to reach control performance. Environmental manipulation and additional training can improve cognition in a rodent MCD model. We therefore suggest that patients with MCD may benefit from appropriate alterations in educational strategies, social interaction and environment. These factors should be considered in therapeutic strategies

Drug-resistant focal epilepsy in children is associated with increased modal controllability of the whole brain and epileptogenic regions

Network control theory provides a framework by which neurophysiological dynamics of the brain can be modelled as a function of the structural connectome constructed from diffusion MRI. Average controllability describes the ability of a region to drive the brain to easy-to-reach neurophysiological states whilst modal controllability describes the ability of a region to drive the brain to difficult-to-reach states. In this study, we identify increases in mean average and modal controllability in children with drug-resistant epilepsy compared to healthy controls. Using simulations, we purport that these changes may be a result of increased thalamocortical connectivity. At the node level, we demonstrate decreased modal controllability in the thalamus and posterior cingulate regions. In those undergoing resective surgery, we also demonstrate increased modal controllability of the resected parcels, a finding specific to patients who were rendered seizure free following surgery. Changes in controllability are a manifestation of brain network dysfunction in epilepsy and may be a useful construct to understand the pathophysiology of this archetypical network disease. Understanding the mechanisms underlying these controllability changes may also facilitate the design of network-focussed interventions that seek to normalise network structure and function

Symptoms of anxiety and depression in school-aged children with active epilepsy: A population-based study

Children (5-15 years) with active epilepsy were screened using the parent-report (n=69) and self-report (n=48) versions of the Spence Children's Anxiety Scale (SCAS) and the self-report version of the Children's Depression Inventory (CDI) (n=48) in a population-based sample. A total of 32.2% of children (self-report) and 15.2% of children (parent-report) scored ≥1 SD above the mean on the SCAS total score. The subscales where most difficulty were reported on parent-report were Physical Injury and Separation Anxiety. There was less variation on self-report. On the CDI, 20.9% of young people scored ≥1 SD above the mean. Children reported significantly more symptoms of anxiety on the SCAS total score and three of the subscales (p&lt;.05). There was a significant effect on the SCAS total score of respondents by seizure type interaction, suggesting higher scores on SCAS for children with generalized seizures on self- but not parent-report. Higher CDI scores were significantly associated with generalized seizures (p&gt;.05).Symptoms of anxiety were more common based on self-report compared with parent-report. Children with generalized seizures reported more symptoms of depression and anxiety

Child and parental sleep in young children with epilepsy: A population-based case-control study

Objective: To determine the prevalence of parent‐reported sleep problems in young children with epilepsy and their parents, and to compare findings with those in a non–epilepsy‐related neurodisability (neurodevelopmental/neurological difficulties) group. Method: Parents of young children (1–7 years) with epilepsy (n = 48 [91% ascertainment]) completed the Child Sleep Habits Questionnaire (CSHQ). Parents (mothers and fathers) also completed the Pittsburgh Sleep Quality Index (PSQI) and the Iowa Fatigue Scale (IFS) in relation to their own functioning. The responses of parents of children with epilepsy were compared with parents of developmental‐, age‐, and gender‐matched children with nonepilepsy‐related neurodisability (n = 48). Results: There was not a significant difference in the proportion of children with epilepsy and the children with neurodisability scoring in the at‐risk range on the CSHQ (81% vs. 71% respectively) (p = 0.232). 62% of mothers and 44% of fathers of children with epilepsy had ‘poor quality sleep’ on the PSQI; there was not a significant difference between mothers of children with epilepsy and those of children with neurodisability (p = 0.526) or IFS (p = 0.245) total scores. However, mothers of children with epilepsy had significantly more difficulties on the productivity subscale of the IFS (p = 0.004). There were no significant differences between fathers’ scores on either measure. In the epilepsy group, child behavioral problems (p = 0.001) were independently associated with child sleep difficulties and maternal mental health problems were associated with parental sleep difficulties (p = 0.04) and fatigue (p = 0.018). Significance: Young children with epilepsy and their parents have a high rate of sleep difficulties. There is a need to develop effective interventions for this population, taking into consideration of the role of child behavioral problems and parental mental health difficulties