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Identifying the genetic determinants of pain is a scientific imperative given the magnitude of the global health burden that pain causes. Here, we report a genetic screen for nociception, performed under the auspices of the International Mouse Phenotyping Consortium. A biased set of 110 single-gene knockout mouse strains was screened for 1 or more nociception and hypersensitivity assays, including chemical nociception (formalin) and mechanical and thermal nociception (von Frey filaments and Hargreaves tests, respectively), with or without an inflammatory agent (complete Freund's adjuvant). We identified 13 single-gene knockout strains with altered nocifensive behavior in 1 or more assays. All these novel mouse models are openly available to the scientific community to study gene function. Two of the 13 genes (Gria1 and Htr3a) have been previously reported with nociception-related phenotypes in genetically engineered mouse strains and represent useful benchmarking standards. One of the 13 genes (Cnrip1) is known from human studies to play a role in pain modulation and the knockout mouse reported herein can be used to explore this function further. The remaining 10 genes (Abhd13, Alg6, BC048562, Cgnl1, Cp, Mmp16, Oxa1l, Tecpr2, Trim14, and Trim2) reveal novel pathways involved in nociception and may provide new knowledge to better understand genetic mechanisms of inflammatory pain and to serve as models for therapeutic target validation and drug development.

Truncating mutations of the maternally imprinted, paternally expressed MAGEL2 gene are the predicted genetic cause of several rare neurodevelopmental disorders including Schaaf-Yang (SYS), Chitayat-Hall and Opitz Trigonocephaly C syndromes. MAGEL2 is also deleted or inactivated in Prader-Willi syndrome (PWS). Previous studies in mice have utilized Magel2 gene deletion models to examine the consequences of its absence. In this study, we report the generation, molecular validation, and phenotypic characterization of a novel rat model with a truncating Magel2 mutation generating a mutant peptide sequence more closely modeling variants associated with SYS-causing mutations. Within the hypothalamus, a brain region wherein mouse and human MAGEL2 is paternally-expressed, we demonstrate at the level of transcript and peptide detection that Magel2 in the rat exhibits a paternal, parent-of-origin effect. In the evaluation of behavioral features across several domains, juvenile Magel2 mutant rats display select alterations in anxiety-like behavior and sociability measures. Moreover, the analysis of peripheral organ systems detected alterations in body composition, cardiac structure and function, and breathing irregularities in Magel2 mutant rats. Several of these findings are concordant with reported mouse phenotypes, signifying the conservation of MAGEL2 function across rodent species for specific behavioral outcome measures. We conclude that our comprehensive analysis demonstrating impairments across multiple domains demonstrates the tractability of this model system for the study of truncating MAGEL2 mutations.

Progress in addressing the origins of intellectual and developmental disabilities accelerated with the establishment 50 years ago of the Eunice Kennedy Shriver National Institute of Child Health and Human Development of the National Institutes of Health and associated Intellectual and Developmental Disabilities Research Centers. Investigators at these Centers have made seminal contributions to understanding human brain and behavioral development and defining mechanisms and treatments of disorders of the developing brain. ANN NEUROL 2019;86:332-343.

Purpose To evaluate the phenotypes of individuals with pathogenic and likely pathogenic variants in the MECP2 gene. Methods We surveyed exome sequencing data from a large clinical care cohort for deleterious variation in the MECP2 gene. We reviewed de-identified clinical information for these individuals to interrogate for neurodevelopmental and neuropsychiatric phenotypes. Results We identified pathogenic and likely pathogenic variants in MECP2 in individuals with typical and atypical Rett syndrome, and neuropsychiatric phenotypes, and estimate a prevalence of MECP2- associated disorders of 1 in 2,645 individuals. We observed a 7.45x increased relative risk of neuropsychiatric phenotypes, especially major depression, in adult individuals with deleterious variants in MECP2 without a diagnosis of Rett syndrome. Male individuals with missense pathogenic variants in MECP2 appear to have more severe neuropsychiatric phenotypes. Conclusions We identified and report individuals with heterozygous pathogenic variants in MECP2 and their phenotypes in a large clinical cohort. The observed prevalence of MECP2-associated disorders in our cohort is higher than estimated in the literature. Individuals with pathogenic variants in MECP2 can survive into adulthood but are at increased risk of developing neuropsychiatric disorders, mainly major depression. Pathogenic variation in MECP2 is a likely important contributor to neuropsychiatric disorders in the general population.

Sustained attention is supported by circuits in the frontoparietal attention network. In human and primate studies, the right posterior parietal cortex (PPC) shows dominance for sustained attention, and phase-amplitude coupling (PAC) throughout the frontoparietal network correlates with performance on attention tasks. Here we evaluate oscillatory dynamics of bilateral PPC in mice during the 5-Choice Serial Reaction Time Task (5-CSRTT). Right PPC theta-gamma PAC (TG-PAC) and gamma power were independently elevated to a greater degree than the left PPC during the period prior to a correct response and were significantly correlated with accuracy in both simple and difficult tasks. Greater task difficulty was also associated with greater hemispheric asymmetry in TG-PAC, favoring the right PPC. These findings highlight the engagement of PPC with sustained attention in mice, reflected by increases in TG-PAC and gamma power, with maximal expression in the right hemisphere.

Motivation High-throughput phenomic projects generate complex data from small treatment and large control groups that increase the power of the analyses but introduce variation over time. A method is needed to utlize a set of temporally local controls that maximizes analytic power while minimizing noise from unspecified environmental factors. Results Here we introduce ‘soft windowing’, a methodological approach that selects a window of time that includes the most appropriate controls for analysis. Using phenotype data from the International Mouse Phenotyping Consortium (IMPC), adaptive windows were applied such that control data collected proximally to mutants were assigned the maximal weight, while data collected earlier or later had less weight. We applied this method to IMPC data and compared the results with those obtained from a standard non-windowed approach. Validation was performed using a resampling approach in which we demonstrate a 10% reduction of false positives from 2.5 million analyses. We applied the method to our production analysis pipeline that establishes genotype–phenotype associations by comparing mutant versus control data. We report an increase of 30% in significant P-values, as well as linkage to 106 versus 99 disease models via phenotype overlap with the soft-windowed and non-windowed approaches, respectively, from a set of 2082 mutant mouse lines. Our method is generalizable and can benefit large-scale human phenomic projects such as the UK Biobank and the All of Us resources. Availability and implementation The method is freely available in the R package SmoothWin, available on CRAN http://CRAN.R-project.org/package=SmoothWin. Supplementary information Supplementary data are available at Bioinformatics online.

Behavioral neuroscience research incorporates the identical high level of meticulous methodologies and exacting attention to detail as all other scientific disciplines. To achieve maximal rigor and reproducibility of findings, well-trained investigators employ a variety of established best practices. Here we explicate some of the requirements for rigorous experimental design and accurate data analysis in conducting mouse and rat behavioral tests. Novel object recognition is used as an example of a cognitive assay which has been conducted successfully with a range of methods, all based on common principles of appropriate procedures, controls, and statistics. Directors of Rodent Core facilities within Intellectual and Developmental Disabilities Research Centers contribute key aspects of their own novel object recognition protocols, offering insights into essential similarities and less-critical differences. Literature cited in this review article will lead the interested reader to source papers that provide step-by-step protocols which illustrate optimized methods for many standard rodent behavioral assays. Adhering to best practices in behavioral neuroscience will enhance the value of animal models for the multiple goals of understanding biological mechanisms, evaluating consequences of genetic mutations, and discovering efficacious therapeutics.

BACKGROUND Ongoing research is necessary to better understand the causes of autism spectrum disorder (ASD), the developmental outcomes for individuals diagnosed with ASD, and the efficacy of the interventions. However, it is often difficult to recruit sufficient numbers of participants for studies, and despite the prevalence of ASD (currently estimated to affect 1 in 54 children), little research has focused on how to efficiently recruit participants with ASD. OBJECTIVE The aim of this study was to determine the efficacy of two different paid advertisements—social media and radio advertising—in recruiting participants for a study enrolling people with ASD and their family members by examining the number of participants enrolled, the cost per participant, and the geographic reach of each type of advertising. METHODS We examined participant enrollment in a study following nonoverlapping paid advertisements on a popular FM radio station (aired in three cities across two states) and Facebook (six advertisements that ran in five cities across two states). The total paid investment in the radio campaign was $12,030 and that in the Facebook campaign was $2950. Following the advertising campaigns, 1391 participants in the study who were affiliated with the Houston, Texas, site received email invitations to participate in a brief survey about the ways in which they learned about the study (eg, social media, medical provider, website) and which of these were most influential in their decisions to participate; 374 (26.8%) of the participants completed this survey. RESULTS Social media advertising outperformed radio in all three parameters examined by enrolling more participants (338 vs 149), with a lower average cost per participant ($8.73 vs $80.74) and a wider geographic reach, based on a comparison of the number of zip codes within and outside of Texas for questionnaire respondents who rated social media as the most influential method of contact (n=367, χ21=5.85, P=.02). Of the 374 survey participants, 139 (37.2%) reported that they had seen the study on social media prior to enrollment, while only 9 (2.4%) said they heard about it via radio. CONCLUSIONS Our findings suggest that advertising on social media can efficiently reach a large pool of potential participants with ASD, increasing the likelihood of meeting study enrollment goals. Researchers should consider allocating at least some portion of recruitment dollars to social media platforms as a means of quickly and inexpensively reaching out to their target populations, including for studies with in-person procedures.

Background Ongoing research is necessary to better understand the causes of autism spectrum disorder (ASD), the developmental outcomes for individuals diagnosed with ASD, and the efficacy of the interventions. However, it is often difficult to recruit sufficient numbers of participants for studies, and despite the prevalence of ASD (currently estimated to affect 1 in 54 children), little research has focused on how to efficiently recruit participants with ASD. Objective The aim of this study was to determine the efficacy of two different paid advertisements—social media and radio advertising—in recruiting participants for a study enrolling people with ASD and their family members by examining the number of participants enrolled, the cost per participant, and the geographic reach of each type of advertising. Methods We examined participant enrollment in a study following nonoverlapping paid advertisements on a popular FM radio station (aired in three cities across two states) and Facebook (six advertisements that ran in five cities across two states). The total paid investment in the radio campaign was $12,030 and that in the Facebook campaign was $2950. Following the advertising campaigns, 1391 participants in the study who were affiliated with the Houston, Texas, site received email invitations to participate in a brief survey about the ways in which they learned about the study (eg, social media, medical provider, website) and which of these were most influential in their decisions to participate; 374 (26.8%) of the participants completed this survey. Results Social media advertising outperformed radio in all three parameters examined by enrolling more participants (338 vs 149), with a lower average cost per participant ($8.73 vs $80.74) and a wider geographic reach, based on a comparison of the number of zip codes within and outside of Texas for questionnaire respondents who rated social media as the most influential method of contact (n=367, χ21=5.85, P=.02). Of the 374 survey participants, 139 (37.2%) reported that they had seen the study on social media prior to enrollment, while only 9 (2.4%) said they heard about it via radio. Conclusions Our findings suggest that advertising on social media can efficiently reach a large pool of potential participants with ASD, increasing the likelihood of meeting study enrollment goals. Researchers should consider allocating at least some portion of recruitment dollars to social media platforms as a means of quickly and inexpensively reaching out to their target populations, including for studies with in-person procedures.

De novo and inherited rare genetic disorders (RGDs) are a major cause of human morbidity, frequently involving neuropsychiatric symptoms. Recent advances in genomic technologies and data sharing have revolutionized the identification and diagnosis of RGDs, presenting an opportunity to elucidate the mechanisms underlying neuropsychiatric disorders by investigating the pathophysiology of high-penetrance genetic risk factors. Here we seek out the best path forward for achieving these goals. We think future research will require consistent approaches across multiple RGDs and developmental stages, involving both the characterization of shared neuropsychiatric dimensions in humans and the identification of neurobiological commonalities in model systems. A coordinated and concerted effort across patients, families, researchers, clinicians and institutions, including rapid and broad sharing of data, is now needed to translate these discoveries into urgently needed therapies. Rare genetic diseases frequently involve a neuropsychiatric component for which a defined framework of investigation will expedite our understanding for these diseases as a whole.

Deep brain stimulation (DBS) has improved the prospects for many individuals with diseases affecting motor control, and recently it has shown promise for improving cognitive function as well. Several studies in individuals with Alzheimer disease and in amnesic rats have demonstrated that DBS targeted to the fimbria-fornix, the region that appears to regulate hippocampal activity, can mitigate defects in hippocampus-dependent memory. Despite these promising results, DBS has not been tested for its ability to improve cognition in any childhood intellectual disability disorder. Such disorders are a pressing concern: they affect as much as 3% of the population and involve hundreds of different genes. We proposed that stimulating the neural circuits that underlie learning and memory might provide a more promising route to treating these otherwise intractable disorders than seeking to adjust levels of one molecule at a time. We therefore studied the effects of forniceal DBS in a well-characterized mouse model of Rett syndrome (RTT), which is a leading cause of intellectual disability in females. Caused by mutations that impair the function of MeCP2 (ref. 6), RTT appears by the second year of life in humans, causing profound impairment in cognitive, motor and social skills, along with an array of neurological features. RTT mice, which reproduce the broad phenotype of this disorder, also show clear deficits in hippocampus-dependent learning and memory and hippocampal synaptic plasticity. Here we show that forniceal DBS in RTT mice rescues contextual fear memory as well as spatial learning and memory. In parallel, forniceal DBS restores in vivo hippocampal long-term potentiation and hippocampal neurogenesis. These results indicate that forniceal DBS might mitigate cognitive dysfunction in RTT.

15q13.3 microdeletion syndrome is characterized by a wide spectrum of neurodevelopmental disorders, including developmental delay, intellectual disability, epilepsy, language impairment, abnormal behaviors, neuropsychiatric disorders, and hypotonia. This syndrome is caused by a deletion on chromosome 15q, which typically encompasses six genes. Here, through studies on OTU deubiquitinase 7A (Otud7a) knockout mice, we identify OTUD7A as a critical gene responsible for many of the cardinal phenotypes associated with 15q13.3 microdeletion syndrome. Otud7a-null mice show reduced body weight, developmental delay, abnormal electroencephalography patterns and seizures, reduced ultrasonic vocalizations, decreased grip strength, impaired motor learning/motor coordination, and reduced acoustic startle. We show that OTUD7A localizes to dendritic spines and that Otud7a-null mice have decreased dendritic spine density compared to their wild-type littermates. Furthermore, frequency of miniature excitatory postsynaptic currents (mEPSCs) is reduced in the frontal cortex of Otud7a-null mice, suggesting a role of Otud7a in regulation of dendritic spine density and glutamatergic synaptic transmission. Taken together, our results suggest decreased OTUD7A dosage as a major contributor to the neurodevelopmental phenotypes associated with 15q13.3 microdeletion syndrome, through the misregulation of dendritic spine density and activity.

Neurobehavioral disorders comprised of neurodegenerative, neurodevelopmental, and psychiatric disorders together represent leading causes of morbidity and mortality. Despite significant academic research and industry efforts to elucidate the disease mechanisms operative in these disorders and to develop mechanism-based therapies, our understanding remains incomplete and our access to tractable therapeutic interventions severely limited. The magnitude of these short-comings can be measured by the growing list of disappointing clinical trials based on initially promising compounds identified in genetic animal models. This review and commentary will explore why this may be so, focusing on the central role that genetic models of neurobehavioral disorders have come to occupy in current efforts to identify disease mechanisms and therapies. In particular, we will highlight the unique pitfalls and challenges that have hampered success in these models as compared to genetic models of non-neurological diseases as well as to symptom-based models of the early 20th century that led to the discovery of all major classes of psychoactive pharmaceutical compounds still used today. Using examples from specific genetic rodent models of human neurobehavioral disorders, we will highlight issues of reproducibility, construct validity, and translational relevance in the hopes that these examples will be instructive toward greater success in future endeavors. Lastly, we will champion a two-pronged approach toward identifying novel therapies for neurobehavioral disorders that makes greater use of the historically more successful symptom-based approaches in addition to more mechanism-based approaches.

Intellectual and developmental disabilities (IDDs) are a common group of disorders that frequently share overlapping symptoms, including cognitive deficits, altered attention, seizures, impaired social interactions, and anxiety. The causes of these disorders are varied ranging from early prenatal/postnatal insults to genetic variants that either cause or are associated with an increased likelihood of an IDD. As many of the symptoms observed in individuals with IDDs are a manifestation of altered nervous system function resulting in altered behaviors, it should not be surprising that the field is very dependent upon in vivo model systems. This special issue of Neurobiology of Learning and Memory is focused on the methods and approaches that are being used to model and understand these disorders in mammals. While surveys by the Pew Foundation continue to find a high degree of confidence/trust in scientists by the public, several recent studies have documented issues with reproducibility in scientific publications. This special issue includes both primary research articles and review articles in which careful attention has been made to transparently report methods and use rigorous approaches to ensure reproducibility. Although there have been and will continue to be remarkable advances for treatment of subset of IDDs, it is clear that this field is still in its early stages. There is no doubt that the strategies being used to model IDDs will continue to evolve. We hope this special issue will support this evolution so that we can maintain the trust of the public and elected officials, and continue developing evidence-based approaches to new therapeutics.

Autism spectrum disorder (ASD) is a constellation of neurodevelopmental presentations with high heritability and both phenotypic and genetic heterogeneity. To date, mutations in hundreds of genes have been associated to varying degrees with increased ASD risk. A better understanding of the functions of these genes and whether they fit together in functional groups or impact similar neuronal circuits is needed to develop rational treatment strategies. We will review current areas of emphasis in ASD research, starting from human genetics and exploring how mouse models of human mutations have helped identify specific molecular pathways (protein synthesis and degradation, chromatin remodeling, intracellular signaling), which are linked to alterations in circuit function and cognitive/social behavior. We will conclude by discussing how we can leverage the findings on molecular and cellular alterations found in ASD to develop therapies for neurodevelopmental disorders.

Rett Syndrome (RTT) is a neurodevelopmental disorder characterized by loss of acquired skills during development, autonomic dysfunction, and an increased risk for premature lethality. Clinical experience identified a subset of individuals with RTT that present with urological dysfunction including individuals with frequent urinary tract infections, kidney stones, and urine retention requiring frequent catheterization for bladder voiding. To determine if urologic dysfunction is a feature of RTT, we queried the Rett Syndrome Natural History Study, a repository of clinical data from over 1000 individuals with RTT and found multiple instances of urological dysfunction. We then evaluated urological function in a mouse model of RTT and found an abnormal pattern of micturition. Both male and female mice possessing Mecp2 mutations show a decrease in urine output per micturition event. Furthermore, we identified signs of kidney failure secondary to urethral obstruction. Although genetic strain background significantly affects both survival and penetrance of the urethral obstruction phenotype, survival and penetrance of urethral obstruction do not directly correlate. We have identified an additional phenotype caused by loss of MeCP2, urological dysfunction. Furthermore, we urge caution in the interpretation of survival data as an endpoint in preclinical studies, especially where causes of mortality are poorly characterized.

Mouse models of the transcriptional modulator Methyl-CpG-Binding Protein 2 (MeCP2) have advanced our understanding of Rett syndrome (RTT). RTT is a 'prototypical' neurodevelopmental disorder with many clinical features overlapping with other intellectual and developmental disabilities (IDD). Therapeutic interventions for RTT may therefore have broader applications. However, the reliance on the laboratory mouse to identify viable therapies for the human condition may present challenges in translating findings from the bench to the clinic. In addition, the need to identify outcome measures in well-chosen animal models is critical for preclinical trials. Here, we report that a novel Mecp2 rat model displays high face validity for modelling psychomotor regression of a learned skill, a deficit that has not been shown in Mecp2 mice. Juvenile play, a behavioural feature that is uniquely present in rats and not mice, is also impaired in female Mecp2 rats. Finally, we demonstrate that evaluating the molecular consequences of the loss of MeCP2 in both mouse and rat may result in higher predictive validity with respect to transcriptional changes in the human RTT brain. These data underscore the similarities and differences caused by the loss of MeCP2 among divergent rodent species which may have important implications for the treatment of individuals with disease-causing MECP2 mutations. Taken together, these findings demonstrate that the Mecp2 rat model is a complementary tool with unique features for the study of RTT and highlight the potential benefit of cross-species analyses in identifying potential disease-relevant preclinical outcome measures.

Genomic duplications spanning Xq28 are associated with a spectrum of phenotypes, including anxiety and autism. The minimal region shared among affected individuals includes MECP2 and IRAK1, although it is unclear which gene when overexpressed causes anxiety and social behavior deficits. We report that doubling MECP2 levels causes heightened anxiety and autism-like features in mice and alters the expression of genes that influence anxiety and social behavior, such as Crh and Oprm1. To test the hypothesis that alterations in these two genes contribute to heightened anxiety and social behavior deficits, we analyzed MECP2 duplication mice (MECP2-TG1) that have reduced Crh and Oprm1 expression. In MECP2-TG1 animals, reducing the levels of Crh or its receptor, Crhr1, suppressed anxiety-like behavior; in contrast, reducing Oprm1 expression improved abnormal social behavior. These data indicate that increased MeCP2 levels affect molecular pathways underlying anxiety and social behavior and provide new insight into potential therapies for MECP2-related disorders.

Introduction 1Rett syndrome (RTT, MIM 312750), originally described by the Austrian pediatrician Andreas Rett, is an X-linked neurodevelopmental disorder that primarily affects girls at a frequency of 1:10,000 live female births ([Rett, 1966][1]; [Hagberg, 1985][2]; [Laurvick et al., 2006][3

Mutations in the X-linked MECP2 gene, which encodes the transcriptional regulator methyl-CpG-binding protein 2 (MeCP2), cause Rett syndrome and several neurodevelopmental disorders including cognitive disorders, autism, juvenile-onset schizophrenia and encephalopathy with early lethality. Rett syndrome is characterized by apparently normal early development followed by regression, motor abnormalities, seizures and features of autism, especially stereotyped behaviours. The mechanisms mediating these features are poorly understood. Here we show that mice lacking Mecp2 from GABA (γ-aminobutyric acid)-releasing neurons recapitulate numerous Rett syndrome and autistic features, including repetitive behaviours. Loss of MeCP2 from a subset of forebrain GABAergic neurons also recapitulates many features of Rett syndrome. MeCP2-deficient GABAergic neurons show reduced inhibitory quantal size, consistent with a presynaptic reduction in glutamic acid decarboxylase 1 (Gad1) and glutamic acid decarboxylase 2 (Gad2) levels, and GABA immunoreactivity. These data demonstrate that MeCP2 is critical for normal function of GABA-releasing neurons and that subtle dysfunction of GABAergic neurons contributes to numerous neuropsychiatric phenotypes.

Rett Syndrome, an X-linked dominant neurodevelopmental disorder characterized by regression of language and hand use, is primarily caused by mutations in methyl-CpG-binding protein 2 (MECP2). Loss of function mutations in MECP2 are also found in other neurodevelopmental disorders such as autism, Angelman-like syndrome and non-specific mental retardation. Furthermore, duplication of the MECP2 genomic region results in mental retardation with speech and social problems. The common features of human neurodevelopmental disorders caused by the loss or increase of MeCP2 function suggest that even modest alterations of MeCP2 protein levels result in neurodevelopmental problems. To determine whether a small reduction in MeCP2 level has phenotypic consequences, we characterized a conditional mouse allele of Mecp2 that expresses 50% of the wild-type level of MeCP2. Upon careful behavioral analysis, mice that harbor this allele display a spectrum of abnormalities such as learning and motor deficits, decreased anxiety, altered social behavior and nest building, decreased pain recognition and disrupted breathing patterns. These results indicate that precise control of MeCP2 is critical for normal behavior and predict that human neurodevelopmental disorders will result from a subtle reduction in MeCP2 expression.

Autism is a common neurodevelopmental disorder of complex genetic etiology. Rett syndrome, an X-linked dominant disorder caused by MECP2 mutations, and Angelman syndrome, an imprinted disorder caused by maternal 15q11-q13 or UBE3A deficiency, have phenotypic and genetic overlap with autism. MECP2 encodes methyl-CpG-binding protein 2 that acts as a transcriptional repressor for methylated gene constructs but is surprisingly not required for maintaining imprinted gene expression. Here, we test the hypothesis that MECP2 deficiency may affect the level of expression of UBE3A and neighboring autism candidate gene GABRB3 without necessarily affecting imprinted expression. Multiple quantitative methods were used including automated quantitation of immunofluorescence and in situ hybridization by laser scanning cytometry on tissue microarrays, immunoblot and TaqMan PCR. The results demonstrated significant defects in UBE3A/E6AP expression in two different Mecp2 deficient mouse strains and human Rett, Angelman and autism brains compared with controls. Although no difference was observed in the allelic expression of several imprinted transcripts in Mecp2-null brain, Ube3a sense expression was significantly reduced, consistent with the decrease in protein. A non-imprinted gene from 15q11-q13, GABRB3, encoding the beta3 subunit of the GABAA receptor, also showed significantly reduced expression in multiple Rett, Angelman and autism brain samples, and Mecp2 deficient mice by quantitative immunoblot. These results suggest an overlapping pathway of gene dysregulation within 15q11-q13 in Rett, Angelman and autism and implicate MeCP2 in the regulation of UBE3A and GABRB3 expressions in the postnatal mammalian brain.

Rett syndrome (RTT) is an X-linked neurodevelopmental disorder caused by mutations in MECP2, encoding methyl-CpG-binding protein 2 (MeCP2). The onset of symptoms in RTT is delayed until 6-18 months and 4-6 months in the Mecp2(-/+) mouse model, corresponding to a dynamic and gradual accumulation of MeCP2 expression in individual neurons of the postnatal brain. Because of X chromosome inactivation (XCI), cells within RTT females are mosaic for expression of the heterozygous MECP2 mutation. Using the targeted Mecp2 mouse model, we investigated the effect of Mecp2 mutation on XCI and developmental MeCP2 expression in wild-type (wt)-expressing neurons by quantitative laser scanning cytometry. Mecp2(-/+) female mice exhibited uniform regional distribution of Mecp2 mutant-expressing cells in brain, but unbalanced XCI in the population, favoring expression of the Mecp2 wt allele. Interestingly, MeCP2 expression in Mecp2 wt-expressing cells from Mecp2(-/+) mice was significantly lower than those from Mecp2(+/+) age-matched controls. The negative effect of Mecp2 mutation on wt Mecp2 expression correlated with the percentage of Mecp2 mutant-expressing cells in the cortex. Similar results were observed in two RTT females with identical MECP2 mutations but different XCI ratios. These results demonstrate that Mecp2-mutant neurons affect the development of surrounding neurons in a non-cell-autonomous manner and suggest that environmental influences affect the level of MeCP2 expression in wt neurons. These results help in explaining the role of XCI in the pathogenesis of RTT and have important implications in designing therapies for female RTT patients.

In September of 2011, the National Institute of Neurological Disorders and Stroke (NINDS), the Eunice Kennedy Shriver National Institute of Child Health and Human Development (NICHD), the International Rett Syndrome Foundation (IRSF) and the Rett Syndrome Research Trust (RSRT) convened a workshop involving a broad cross-section of basic scientists, clinicians and representatives from the National Institutes of Health (NIH), the US Food and Drug Administration (FDA), the pharmaceutical industry and private foundations to assess the state of the art in animal studies of Rett syndrome (RTT). The aim of the workshop was to identify crucial knowledge gaps and to suggest scientific priorities and best practices for the use of animal models in preclinical evaluation of potential new RTT therapeutics. This review summarizes outcomes from the workshop and extensive follow-up discussions among participants, and includes: (1) a comprehensive summary of the physiological and behavioral phenotypes of RTT mouse models to date, and areas in which further phenotypic analyses are required to enhance the utility of these models for translational studies; (2) discussion of the impact of genetic differences among mouse models, and methodological differences among laboratories, on the expression and analysis, respectively, of phenotypic traits; and (3) definitions of the standards that the community of RTT researchers can implement for rigorous preclinical study design and transparent reporting to ensure that decisions to initiate costly clinical trials are grounded in reliable preclinical data.

Rett syndrome (RTT) is an X-linked neurological disorder caused by mutations in the gene encoding the transcriptional modulator methyl-CpG-binding protein 2 (MeCP2). Typical RTT primarily affects girls and is characterized by a brief period of apparently normal development followed by the loss of purposeful hand skills and language, the onset of anxiety, hand stereotypies, autistic features, seizures and autonomic dysfunction. Mecp2 mouse models have extensively been studied to demonstrate the functional link between MeCP2 dysfunction and RTT pathogenesis. However, the majority of studies have focused primarily on the molecular and behavioral consequences of the complete absence of MeCP2 in male mice. Studies of female Mecp2(+/-) mice have been limited because of potential phenotypic variability due to X chromosome inactivation effects. To determine whether reproducible and reliable phenotypes can be detected Mecp2(+/-) mice, we analyzed Mecp2(+/-) mice of two different F1 hybrid isogenic backgrounds and at young and old ages using several neurobehavioral and physiological assays. Here, we report a multitude of phenotypes in female Mecp2(+/-) mice, some presenting as early as 5 weeks of life. We demonstrate that Mecp2(+/-) mice recapitulate several aspects of typical RTT and show that mosaic expression of MeCP2 does not preclude the use of female mice in behavioral and molecular studies. Importantly, we uncover several behavioral abnormalities that are present in two genetic backgrounds and report on phenotypes that are unique to one background. These findings provide a framework for pre-clinical studies aimed at improving the constellation of phenotypes in a mouse model of RTT.

Rett syndrome (RTT) is a postnatal neurological disorder caused by mutations in MECP2, encoding the epigenetic regulator methyl-CpG-binding protein 2 (MeCP2). The onset of RTT symptoms during early life together with findings suggesting neurodevelopmental abnormalities in RTT and mouse models of RTT raised the question of whether maintaining MeCP2 function exclusively during early life might protect against disease. We show by using an inducible model of RTT that deletion of Mecp2 in adult mice recapitulates the germline knock-out phenotype, underscoring the ongoing role of MeCP2 in adult neurological function. Moreover, unlike the effects of other epigenetic instructions programmed during early life, the effects of early MeCP2 function are lost soon after its deletion. These findings suggest that therapies for RTT must be maintained throughout life.

Rett syndrome (RTT) is characterized by specific motor, cognitive, and behavioral deficits. Because several of these abnormalities occur in other disease states associated with alterations in aminergic neurotransmitters, we investigated the contribution of such alterations to RTT pathogenesis. We found that both individuals with RTT and Mecp2 -null mice have lower-than-normal levels of aminergic metabolites and content. Deleting Mecp2 from either TH-positive dopaminergic and noradrenergic neurons or PET1-positive serotonergic neurons in mice decreased corresponding neurotransmitter concentration and specific phenotypes, likely through MeCP2 regulation of rate-limiting enzymes involved in aminergic neurotransmitter production. These data support a cell-autonomous, MeCP2-dependent mechanism for the regulation of aminergic neurotransmitter synthesis contributing to unique behavioral phenotypes.

SummaryRett Syndrome (RTT) is an autism spectrum disorder caused by mutations in the X-linked gene encoding methyl-CpG binding protein 2 (MeCP2). In order to map the neuroanatomic origins of the complex neuropsychiatric behaviors observed in patients with RTT and to uncover endogenous functions of MeCP2 in the hypothalamus, we removed Mecp2 from Sim1-expressing neurons in the hypothalamus using Cre-loxP technology. Loss of MeCP2 in Sim1-expressing neurons resulted in mice that recapitulated the abnormal physiological stress response that is seen upon MeCP2 dysfunction in the entire brain. Surprisingly, we also uncovered a role for MeCP2 in the regulation of social and feeding behaviors since the Mecp2 conditional knockout (CKO) mice were aggressive, hyperphagic, and obese. This study demonstrates that deleting Mecp2 in a defined brain region is an excellent approach to map the neuronal origins of complex behaviors and provides new insight about the function of MeCP2 in specific neurons.

Spinocerebellar ataxia type 1 is caused by expansion of a translated CAG repeat in ataxin1 (ATXN1). The level of the polyglutamine-expanded protein is one of the factors that contributes to disease severity. Here we found that miR-19, miR-101 and miR-130 co-regulate ataxin1 levels and that their inhibition enhanced the cytotoxicity of polyglutamine-expanded ATXN1 in human cells. We provide a new candidate mechanism for modulating the pathogenesis of neurodegenerative diseases sensitive to protein dosage.

Spinocerebellar ataxia type 1 (SCA1) is a dominantly inherited neurodegenerative disease caused by expansion of a glutamine tract in ataxin-1 (ATXN1). SCA1 pathogenesis studies support a model in which the expanded glutamine tract causes toxicity by modulating the normal activities of ATXN1. To explore native interactions that modify the toxicity of ATXN1, we generated a targeted duplication of the mouse ataxin-1-like (Atxn1l, also known as Boat) locus, a highly conserved paralog of SCA1, and tested the role of this protein in SCA1 pathology. Using a knock-in mouse model of SCA1 that recapitulates the selective neurodegeneration seen in affected individuals, we found that elevated Atxn1l levels suppress neuropathology by displacing mutant Atxn1 from its native complex with Capicua (CIC). Our results provide genetic evidence that the selective neuropathology of SCA1 arises from modulation of a core functional activity of ATXN1, and they underscore the importance of studying the paralogs of genes mutated in neurodegenerative diseases to gain insight into mechanisms of pathogenesis.

Fox (forkhead) genes encode transcription factors that play important roles in the regulation of embryonic patterning as well as in tissue specific gene expression. Mutations in the human FOXP2 gene cause abnormal speech development. Here we report the structure and expression pattern of zebrafish FoxP2. In zebrafish, this gene is first expressed at the 20-somite stage in the presumptive telencephalon. At this stage there is a significant overlap of FoxP2 expression with the expression of the emx homeobox genes. However, in contrast to emx1, FoxP2 is not expressed in the pineal gland or in the pronephric duct. After 72 hours of development, the expression of zebrafish FoxP2 becomes more complex in the brain. The developing optic tectum becomes the major area of FoxP2 expression. In the adult brain, the highest concentrations of the FoxP2 transcript can be observed in the optic tectum. In the cerebellum, only the caudal lobes show high levels of Foxp2 expression. These regions correspond to the vestibulocerebellum of mammals. Several other regions of the brain also show high levels of Foxp2 expression.

Rett syndrome (RTT) is a neurodevelopmental disorder caused by mutations in MECP2, encoding methyl-CpG-binding protein 2 (MeCP2). Although MECP2 is ubiquitously transcribed, MeCP2 expression is developmentally regulated and heterogeneous in neuronal subpopulations, defined as MeCP2(lo) and MeCP2(hi). To test the hypothesis that pathways affecting MeCP2 expression changes may be defective in RTT, autism and other neurodevelopmental disorders without MECP2 mutations, a high-throughput quantitation of MeCP2 expression was performed on a tissue microarray containing frontal cortex samples from 28 different patients with neurodevelopmental disorders and age-matched controls. Combined quantitative analyses of MeCP2 protein and alternatively polyadenylated transcript levels were performed by laser scanning cytometry and tested for significant differences from age-matched controls. Normal cerebral samples showed an increase in total MeCP2 expression and the percentage of MeCP2(hi) cells with age that could be explained by increased MECP2 transcription within the MeCP2(hi) population. A significant decrease in the relative usage of the long transcript in the MeCP2(lo) population was observed in postnatal compared to fetal brain, but alternate polyadenylation did not correlate with MeCP2 expression changes at the single cell level. Brain samples from several related neurodevelopmental disorders, including autism, pervasive developmental disorder, Prader-Willi and Angelman syndromes showed significant differences in MeCP2 expression from age-matched controls by apparently different transcriptional and post-transcriptional mechanisms. These results suggest that multiple pathways regulate the complex developmental expression of MeCP2 and are defective in autism-spectrum disorders in addition to RTT.

Hypothesis: Perihepatic packs used to control hemorrhage after liver injury increase the risk of complications and this risk increases the longer packs are left in place. Design: Retrospective case series. Setting: University level I trauma center. Patients: Consecutive patients with hepatic injury. Main Outcome Measures: Liver-related complications (biliary leak and abscess), rebleeding, and mortality. Results: One hundred twenty-nine of 804 patients with liver injuries were treated with perihepatic packing. Of the 69 who survived more than 24 hours, 75% lived to hospital discharge. Mortality rates were 14% and 30% in patients with and without liver complications, respectively (P =.23). Liver complication rates were similar (P = .83) when packs were removed within 36 hours (early [33%]) or between 36 and 72 hours (late [29%]) after they were placed; the rebleeding rate was greater in the early group (21% vs 4%; P