A case report of severe tuberous sclerosis complex detected in utero and linked to a novel duplication in the TSC2 gene.

BACKGROUND: Disease severity is tremendously variable in tuberous sclerosis complex (TSC). In contrast with the detailed guidelines available for TSC diagnosis and management, clinical practice lacks adequate tools to evaluate the prognosis, especially in the case of in utero diagnosis. In addition, the correlation between genotypes and phenotypes remains a challenge, in part due to the large number of mutations linked to TSC. In this report, we describe a case of severe TSC diagnosed in utero and associated with a specific mutation in the gene tuberous sclerosis complex 2 (TSC2). CASE PRESENTATION: A mother was referred for a thorough investigation following the observation by ultrasound of cardiac abnormalities in her fetus. The mother was healthy and reported frequent, intense and long-lasting hiccups/spasms in the fetus. The fetus of gestational age 33 weeks and 4 days was found to have multiple cardiac tumors with cardiac ultrasound. Brain magnetic resonance imaging (MRI) performed in utero revealed the presence of sub-ependymal nodules and of abnormal signals disseminated in the white matter, in the cerebral cortex and in the cerebellum. Following diagnosis of definite TSC, pregnancy interruption was chosen by the parents. Genetic testing of the fetus exposed a duplication in exon 41 of TSC2 (c.5169dupA), which was absent in the parents. The autopsy ascertained the high severity of brain damage characterized by an extensive disorganisation of white and grey matter in most cerebral lobes. CONCLUSIONS: This case presentation is the first to depict the association between a de novo TSC2 c.5169dupA and multi-organ manifestation together with indications of a particularly high disease severity. This report can help physicians to perform early clinical diagnosis of TSC and to evaluate the prognosis.

Bidirectional relationships between sleep and amyloid-beta in the hippocampus.

Alzheimer's disease (AD) is a debilitating neurodegenerative disease characterized by progressive hippocampal-dependent explicit memory deficits that begin at the onset of the illness. An early hallmark of AD is the accumulation of amyloid-beta (Aß) proteins in brain structures involved in encoding and consolidation of memory, like the hippocampus and prefrontal cortex. Aß neurotoxicity is known to induce synaptic dysfunctions and neuronal death leading to cognitive decline. Another recurrent event observed in AD is sleep disturbances. Decreased sleep duration, sleep fragmentation, and circadian alterations are often observed in early AD. The origin of these disturbances, and especially the specific contribution of the hippocampal Aß pathology, remains to be determined. It is required to identify mechanisms impacting wakefulness and sleep architecture and microarchitecture given the role of sleep in memory encoding and consolidation. Sleep perturbations in AD are thus likely contributing to memory decline in the course of the disease. The central aim of this review is to address the bidirectional relationship between sleep and hippocampal Aß by discussing the literature featuring data on wakefulness and sleep variables (i.e., duration, electroencephalographic activity, daily distribution) in AD mouse models and on the effect of enforced sleep loss on Aß pathology in the hippocampus. The current state of knowledge on this topic emphasizes a clear need for more efforts to assess the precise impact of hippocampal Aß on wakefulness and sleep quality as well as the mechanisms mediating their reciprocal relationship.

Characterization of Sleep, Emotional, and Cognitive Functions in a New Rat Model of Concomitant Spinal Cord and Traumatic Brain Injuries.

Traumatic injuries to the spinal cord or the brain have serious medical consequences and lead to long-term disability. The epidemiology, medical complications, and prognosis of isolated spinal cord injury (SCI) and traumatic brain injury (TBI) have been well described. However, there are limited data on patients suffering from concurrent SCI and TBI, even if a large proportion of SCI patients have concomitant TBI. The complications associated with this "dual-diagnosis" such as cognitive or behavioral dysfunction are well known in the rehabilitation setting, but evidence-based and standardized approaches for diagnosis and treatment are lacking. Our goal was to develop and characterize a pre-clinical animal model of concurrent SCI and TBI to help identifying "dual-diagnosis" tools. Female rats received a unilateral contusive SCI at the thoracic level alone (SCI group) or combined with a TBI centered on the contralateral sensorimotor cortex (SCI-TBI group). We first validated that the SCI extent was comparable between SCI-TBI and SCI groups, and that hindlimb function was impaired. We characterized various neurological outcomes, including locomotion, sleep architecture, brain activity during sleep, depressive- and anxiety-like behaviors, and working memory. We report that SCI-TBI and SCI groups show similar impairments in global locomotor function. While wake/sleep amount and distribution and anxiety- and depression-like symptoms were not affected in SCI-TBI and SCI groups in comparison to the control group (laminectomy and craniotomy only), working memory was impaired only in SCI-TBI rats. This pre-clinical model of concomitant SCI and TBI, including more severe variations of it, shows a translational value for the identification of biomarkers to refine the "dual-diagnosis" of neurotrauma in humans.

Probing pathways by which rhynchophylline modifies sleep using spatial transcriptomics.

BACKGROUND: Rhynchophylline (RHY) is an alkaloid component of Uncaria, which are plants extensively used in traditional Asian medicines. Uncaria treatments increase sleep time and quality in humans, and RHY induces sleep in rats. However, like many traditional natural treatments, the mechanisms of action of RHY and Uncaria remain evasive. Moreover, it is unknown whether RHY modifies key brain oscillations during sleep. We thus aimed at defining the effects of RHY on sleep architecture and oscillations throughout a 24-h cycle, as well as identifying the underlying molecular mechanisms. Mice received systemic RHY injections at two times of the day (beginning and end of the light period), and vigilance states were studied by electrocorticographic recordings. RESULTS: RHY enhanced slow wave sleep (SWS) after both injections, suppressed paradoxical sleep (PS) in the light but enhanced PS in the dark period. Furthermore, RHY modified brain oscillations during both wakefulness and SWS (including delta activity dynamics) in a time-dependent manner. Interestingly, most effects were larger in females. A brain spatial transcriptomic analysis showed that RHY modifies the expression of genes linked to cell movement, apoptosis/necrosis, and transcription/translation in a brain region-independent manner, and changes those linked to sleep regulation (e.g., Hcrt, Pmch) in a brain region-specific manner (e.g., in the hypothalamus). CONCLUSIONS: The findings provide support to the sleep-inducing effect of RHY, expose the relevance to shape wake/sleep oscillations, and highlight its effects on the transcriptome with a high spatial resolution. The exposed molecular mechanisms underlying the effect of a natural compound should benefit sleep- and brain-related medicine.

Transcriptional regulation of the mouse EphA4, Ephrin-B2 and Ephrin-A3 genes by the circadian clock machinery.

Circadian rhythms originate from molecular feedback loops. In mammals, the transcription factors CLOCK and BMAL1 act on regulatory elements (i.e. E-boxes) to shape biological functions in a rhythmic manner. The EPHA4 receptor and its ligands Ephrins (EFN) are cell adhesion molecules regulating neurotransmission and neuronal morphology. Previous studies showed the presence of E-boxes in the genes of EphA4 and specific Ephrins, and that EphA4 knockout mice have an altered circadian rhythm of locomotor activity. We thus hypothesized that the core clock machinery regulates the gene expression of EphA4, EfnB2 and EfnA3. CLOCK and BMAL1 (or NPAS2 and BMAL2) were found to have transcriptional activity on distal and proximal regions of EphA4, EfnB2 and EfnA3 putative promoters. A constitutively active form of glycogen synthase kinase 3ß (GSK3ß; a negative regulator of CLOCK and BMAL1) blocked the transcriptional induction. Mutating the E-boxes of EphA4 distal promoter sequence reduced transcriptional induction. EPHA4 and EFNB2 protein levels did not show circadian variations in the mouse suprachiasmatic nucleus or prefrontal cortex. The findings uncover that core circadian transcription factors can regulate the gene expression of elements of the Eph/Ephrin system, which might contribute to circadian rhythmicity in biological processes in the brain or peripheral tissues.

Electrocorticographic Recording of Cerebral Cortex Areas Manipulated Using an Adeno-Associated Virus Targeting Cofilin in Mice.

The use of electrocorticographic (ECoG) recordings in rodents is relevant to sleep research and to the study of a wide range of neurological conditions. Adeno-associated viruses (AAVs) are increasingly used to improve understanding of brain circuits and their functions. The AAV-mediated manipulation of specific cell populations and/or of precise molecular components has been tremendously useful to identify new sleep regulatory circuits/molecules and key proteins contributing to the adverse effects of sleep loss. For instance, inhibiting activity of the filamentous actin-severing protein cofilin using AAV prevents sleep deprivation-induced memory impairment. Here, a protocol is described that combines the manipulation of cofilin function in a cerebral cortex area with the recording of ECoG activity to examine whether cortical cofilin modulates the wakefulness and sleep ECoG signals. AAV injection is performed during the same surgical procedure as the implantation of ECoG and electromyographic (EMG) electrodes in adult male and female mice. Mice are anesthetized, and their heads are shaved. After skin cleaning and incision, stereotaxic coordinates of the motor cortex are determined, and the skull is pierced at this location. A cannula prefilled with an AAV expressing cofilinS3D, an inactive form of cofilin, is slowly positioned in the cortical tissue. After AAV infusion, gold-covered screws (ECoG electrodes) are screwed through the skull and cemented to the skull with gold wires inserted in the neck muscles (EMG electrodes). The animals are allowed three weeks to recover and to ensure sufficient expression of cofilinS3D. The infected area and cell type are verified using immunohistochemistry, and the ECoG is analyzed using visual identification of vigilance states and spectral analysis. In summary, this combined methodological approach allows the investigation of the precise contribution of molecular components regulating neuronal morphology and connectivity to the regulation of synchronized cerebral cortex activity during wakefulness and sleep.

Neuroligin-1 is altered in the hippocampus of Alzheimer's disease patients and mouse models, and modulates the toxicity of amyloid-beta oligomers.

Synapse loss occurs early and correlates with cognitive decline in Alzheimer's disease (AD). Synaptotoxicity is driven, at least in part, by amyloid-beta oligomers (Aßo), but the exact synaptic components targeted by Aßo remain to be identified. We here tested the hypotheses that the post-synaptic protein Neuroligin-1 (NLGN1) is affected early in the process of neurodegeneration in the hippocampus, and specifically by Aßo, and that it can modulate Aßo toxicity. We found that hippocampal NLGN1 was decreased in patients with AD in comparison to patients with mild cognitive impairment and control subjects. Female 3xTg-AD mice also showed a decreased NLGN1 level in the hippocampus at an early age (i.e., 4 months). We observed that chronic hippocampal Aßo injections initially increased the expression of one specific Nlgn1 transcript, which was followed by a clear decrease. Lastly, the absence of NLGN1 decreased neuronal counts in the dentate gyrus, which was not the case in wild-type animals, and worsens impairment in spatial learning following chronic hippocampal Aßo injections. Our findings support that NLGN1 is impacted early during neurodegenerative processes, and that Aßo contributes to this effect. Moreover, our results suggest that the presence of NLGN1 favors the cognitive prognosis during Aßo-driven neurodegeneration.

Regulation of the Neuroligin-1 Gene by Clock Transcription Factors.

NEUROLIGIN-1 (NLGN1) is a postsynaptic adhesion molecule involved in the regulation of glutamatergic transmission. It has been associated with several features of sleep and psychiatric disorders. Our previous work suggested that transcription of the Nlgn1 gene could be regulated by the transcription factors CLOCK and BMAL1 because they bind to the Nlgn1 gene promoter in vivo. However, whether CLOCK/BMAL1 can directly activate Nlgn1 transcription is not yet known. We thus aimed to verify whether CLOCK/BMAL1, as well as their homologs NPAS2 and BMAL2, can activate transcription via the Nlgn1 promoter by using luciferase assays in COS-7 cells. We also investigated how Nlgn1 expression was affected in Clock mutant mice. Our results show transcriptional activation in vitro mediated by CLOCK/BMAL1 and by combinations with their homologs NPAS2 and BMAL2. Moreover, CLOCK/BMAL1 activation via the Nlgn1 gene fragment was repressed by GSK3ß. In vivo, Nlgn1 mRNA expression was significantly modified in the forebrain of Clock mutant mice in a transcript variant-dependent manner. However, no significant change in NLGN1 protein level was observed in Clock mutant mice. These findings will increase knowledge about the transcriptional regulation of Nlgn1 and the relationship between circadian rhythms, mental health, and sleep.

The Tyrosine Phosphatase STEP Is Involved in Age-Related Memory Decline.

Cognitive disabilities that occur with age represent a growing and expensive health problem. Age-associated memory deficits are observed across many species, but the underlying molecular mechanisms remain to be fully identified. Here, we report elevations in the levels and activity of the striatal-enriched phosphatase (STEP) in the hippocampus of aged memory-impaired mice and rats, in aged rhesus monkeys, and in people diagnosed with amnestic mild cognitive impairment (aMCI). The accumulation of STEP with aging is related to dysfunction of the ubiquitin-proteasome system that normally leads to the degradation of STEP. Higher level of active STEP is linked to enhanced dephosphorylation of its substrates GluN2B and ERK1/2, CREB inactivation, and a decrease in total levels of GluN2B and brain-derived neurotrophic factor (BDNF). These molecular events are reversed in aged STEP knockout and heterozygous mice, which perform similarly to young control mice in the Morris water maze (MWM) and Y-maze tasks. In addition, administration of the STEP inhibitor TC-2153 to old rats significantly improved performance in a delayed alternation T-maze memory task. In contrast, viral-mediated STEP overexpression in the hippocampus is sufficient to induce memory impairment in the MWM and Y-maze tests, and these cognitive deficits are reversed by STEP inhibition. In old LOU/C/Jall rats, a model of healthy aging with preserved memory capacities, levels of STEP and GluN2B are stable, and phosphorylation of GluN2B and ERK1/2 is unaltered. Altogether, these data suggest that elevated levels of STEP that appear with advancing age in several species contribute to the cognitive declines associated with aging.