Neocortical interactions with dorsal striatum support many motor and executive functions, and such underlying functional networks are particularly vulnerable to a variety of developmental, neurological, and psychiatric brain disorders, including autism spectrum disorders, Parkinson’s disease, and Huntington’s disease. Relatively little is known about the development of functional corticostriatal interactions, and in particular, virtually nothing is known of molecular mechanisms that control generation of prefrontal cortex-striatal circuits. Here, we used regional and cellular in situ hybridization techniques coupled with neuronal tract tracing to show that Cadherin 8 (Cdh8), a homophilic adhesion protein encoded by a gene associated with autism spectrum disorders and learning disability susceptibility, is enriched within striatal projection neurons in medial prefrontal cortex and in striatal medium spiny neurons forming the direct- or indirect-pathways. Developmental analysis of quantitative RTPCR and Western blot data show that Cdh8 expression peaks in prefrontal cortex and striatum at P10, when cortical projections start to form synapses in the striatum. High-resolution immunoelectron-microscopy shows Cdh8 is concentrated at excitatory synapses in dorsal striatum, and Cdh8 knockdown in cortical neurons impairs dendritic arborization and dendrite self-avoidance. Taken together our findings indicate that Cdh8 delineates developing corticostriatal circuits where it is a strong candidate for regulating the generation of normal cortical projections, neuronal morphology, and corticostriatal synapses.

Target of rapamycin complex 1 (TORC1) is an important regulator of neuronal function. However, whereas a modest activation of the TORC1 signaling pathway has been shown to affect synaptic plasticity, learning and memory, the effect of TORC1 hypo-activation is less clear. This knowledge is particularly important since TORC1 inhibitors may hold great promise for treating a variety of disorders, including developmental disorders, aging-related disorders, epilepsy and cancer. Such treatments are likely to be long lasting and could involve treating young children. Hence, it is pivotal that the effects of sustained TORC1 inhibition on brain development and cognitive function are determined. Here, we made use of constitutive and conditional Rheb1 mutant mice to study the effect of prolonged and specific reduction in the TORC1 pathway. We show that Rheb1 mutant mice show up to 75% reduction in TORC1 signaling, but develop normally and show intact synaptic plasticity and hippocampus-dependent learning and memory. We discuss our findings in light of current literature in which the effect of pharmacological inhibition of TORC1 is studied in the context of synaptic plasticity and learning. We conclude that in contrast to TORC1 hyper-activity, cognitive function is not very sensitive to sustained and specific down-regulation of TORC1 activity.

Background: The most-common strategy for zebrafish Cre/lox-mediated lineage labeling experiments combines ubiquitously expressed, lox-based Switch reporter transgenes with tissue-specific Cre or 4-OH-Tamoxifen-inducible CreERT2 driver lines. Although numerous Cre driver lines have been produced, only a few broadly expressed Switch reporters exist in zebrafish and their generation by random transgene integration has been challenging due to position-effect sensitivity of the lox-flanked recombination cassettes. Here, we compare commonly used Switch reporter lines for their recombination efficiency and reporter expression pattern during zebrafish development. Results: Using different experimental setups, we show that ubi:Switch and hsp70l:Switch outperform current generations of the two additional Switch reporters actb2:BFP-DsRed and actb2:Stop-DsRed. Our comparisons also document preferential Cre-dependent recombination of ubi:Switch and hsp70l: Switch in distinct zebrafish tissues at early developmental stages. To investigate what genomic features may influence Cre accessibility and lox recombination efficiency in highly functional Switch lines, we mapped these transgenes and charted chromatin dynamics at their integration sites. Conclusions: Our data documents the heterogeneity among lox-based Switch transgenes toward informing suitable transgene selection for lineage labeling experiments. Our work further proposes that ubi:Switch and hsp70l:Switch define genomic integration sites suitable for universal transgene or switch reporter knock-in in zebrafish.

The heart is the first functional organ to form during vertebrate development. Congenital heart defects are the most common type of human birth defect, many originating as anomalies in early heart development. The zebrafish model provides an accessible vertebrate system to study early heart morphogenesis and to gain new insights into the mechanisms of congenital disease. Although composed of only two chambers compared with the four-chambered mammalian heart, the zebrafish heart integrates the core processes and cellular lineages central to cardiac development across vertebrates. The rapid, translucent development of zebrafish is amenable to in vivo imaging and genetic lineage tracing techniques, providing versatile tools to study heart field migration and myocardial progenitor addition and differentiation. Combining transgenic reporters with rapid genome engineering via CRISPR-Cas9 allows for functional testing of candidate genes associated with congenital heart defects and the discovery of molecular causes leading to observed phenotypes. Here, we summarize key insights gained through zebrafish studies into the early patterning of uncommitted lateral plate mesoderm into cardiac progenitors and their regulation. We review the central genetic mechanisms, available tools, and approaches for modeling congenital heart anomalies in the zebrafish as a representative vertebrate model.

Endothelial specification is a key event during embryogenesis; however, when, and how, endothelial cells separate from other lineages is poorly understood. In zebrafish, Npas4l is indispensable for endothelial specification by inducing the expression of the transcription factor genes etsrp , tal1 , and lmo2 . We generated a knock-in reporter in zebrafish npas4l to visualize endothelial progenitors and their derivatives in wild-type and mutant embryos. Unexpectedly, we find that in npas4l mutants, npas4l reporter–expressing cells contribute to the pronephron tubules. Single-cell transcriptomics and live imaging of the early lateral plate mesoderm in wild-type embryos indeed reveals coexpression of endothelial and pronephron markers, a finding confirmed by creERT2-based lineage tracing. Increased contribution of npas4l reporter–expressing cells to pronephron tubules is also observed in tal1 and lmo2 mutants and is reversed in npas4l mutants injected with tal1 mRNA. Together, these data reveal that Npas4l/Tal1/Lmo2 regulate the fate decision between the endothelial and pronephron lineages.

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Human homologue of yeast UV excision repair protein Rad23b (HR23B) inclusions are found in a number of neurodegenerative diseases, including frontotemporal dementia (FTD), Huntington’s disease (HD), spinocerebellar ataxia type 3 and 7 (SCA3/7), fragile X associated tremor/ataxia syndrome (FXTAS) and Parkinson’s disease (PD). Here, we describe HR23B pathology in C9ORF72 linked FTD and amyotrophic lateral sclerosis (ALS) cases. HR23B presented in neuropils, intranuclear inclusions and cytoplasmic and perinuclear inclusions and was predominantly found in cortices (frontal, temporal and motor), spinal cord and hippocampal dentate gyrus. HR23B co-localized with poly-GA-, pTDP-43- and p62-positive inclusions in frontal cortex and in hippocampal dentate gyrus, the latter showing higher co-localization percentages. HR23B binding partners XPC, 20S and ataxin-3, which are involved in nucleotide excision repair (NER) and the ubiquitin-proteasome system (UPS), did not show an aberrant distribution. However, C9ORF72 fibroblasts were more sensitive for UV-C damage than healthy control fibroblasts, even though all factors involved in NER localized normally to DNA damage and the efficiency of DNA repair was not reduced. HR23Bs other binding partner NGly1/PNGase, involved in ER-associated degradation (ERAD) of misfolded proteins, was not expressed in the majority of neurons in C9FTD/ALS brain sections compared to non-demented controls. Our results suggest a difference in HR23B aggregation and co-localization pattern with DPRs, pTDP-43 and p62 between different brain areas from C9FTD/ALS cases. We hypothesize that HR23B may play a role in C9ORF72 pathogenesis, possibly by aberrant ERAD functioning.Electronic supplementary materialThe online version of this article (10.1186/s40478-019-0694-6) contains supplementary material, which is available to authorized users.

The hexanucleotide G4C2 repeat expansion in the first intron of the C9ORF72 gene explains the majority of frontotemporal dementia (FTD) and amyotrophic lateral sclerosis (ALS) cases. Numerous studies have indicated the toxicity of dipeptide repeats (DPRs) which are produced via repeat-associated non-AUG (RAN) translation from the repeat expansion and accumulate in the brain of C9FTD/ALS patients. Mouse models expressing the human C9ORF72 repeat and/or DPRs show variable pathological, functional and behavioral characteristics of FTD and ALS. Here, we report a new Tet-on inducible mouse model that expresses 36x pure G4C2 repeats with 100bp upstream and downstream human flanking regions. Brain specific expression causes the formation of sporadic sense DPRs aggregates upon 6 months dox induction but no apparent neurodegeneration. Expression in the rest of the body evokes abundant sense DPRs in multiple organs, leading to weight loss, neuromuscular junction disruption, myopathy and a locomotor phenotype within the time frame of four weeks. We did not observe any RNA foci or pTDP-43 pathology. Accumulation of DPRs and the myopathy phenotype could be prevented when 36x G4C2 repeat expression was stopped after 1 week. After 2 weeks of expression, the phenotype could not be reversed, even though DPR levels were reduced. In conclusion, expression of 36x pure G4C2 repeats including 100bp human flanking regions is sufficient for RAN translation of sense DPRs and evokes a functional locomotor phenotype. Our inducible mouse model highlights the importance of early diagnosis and treatment for C9FTD/ALS patients.Summary statementOnly 36 C9ORF72 repeats are sufficient for RAN translation in a new mouse model for ALS and FTD. Reducing toxic dipeptides can prevent but not reverse the phenotype.