Virally Encoded Connectivity Transgenic Overlay RNA sequencing (VECTORseq) defines projection neurons involved in sensorimotor integration
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA784983 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR17078535_1.fastq.gz | Fastqsanger.gz | |||
| SRR17078535_2.fastq.gz | Fastqsanger.gz | |||
| SRR17078536_1.fastq.gz | Fastqsanger.gz | |||
| SRR17078536_2.fastq.gz | Fastqsanger.gz | |||
| SRR17078537_1.fastq.gz | Fastqsanger.gz |
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Project description:Purpose: Establish a high-throughput method to transcriptionally define projection neurons, VECTORseq, that reimagines transgenes expressed by widely used retrogradely infecting viruses as multiplexed RNA barcodes that are detected in single-cell sequencing. Methods: mRNA profiles of adult mouse brains Conclusions: Retrograde viruses express mRNA at levels detectable in single-cell sequencing. Different transgenes can be multiplexed in a single sequencing run. VECTORseq identifies both cortical and subcortical projection neurons. VECTORseq defined new superior colliculus and zona incerta projection populations. Established a high-throughput method to transcriptionally define projection neurons, VECTORseq, that reimagines transgenes expressed by widely used retrogradely infecting viruses as multiplexed RNA barcodes that are detected in single-cell sequencing.
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Project description:Astrocytes, the most abundant cells in the central nervous system, promote synapse formation and help refine neural connectivity. Although they are allocated to spatially distinct regional domains during development, it is unknown whether region-restricted astrocytes are functionally heterogeneous. Here we show that postnatal spinal cord astrocytes express several region-specific genes, and that ventral astrocyte-encoded Semaphorin3a (Sema3a) is required for proper motor neuron and sensory neuron circuit organization. Loss of astrocyte-encoded Sema3a led to dysregulated α−motor neuron axon initial segment orientation, markedly abnormal synaptic inputs, and selective death of α−but not of adjacent γ−motor neurons. Additionally, a subset of TrkA+ sensory afferents projected to ectopic ventral positions. These findings demonstrate that stable maintenance of a positional cue by developing astrocytes influences multiple aspects of sensorimotor circuit formation. More generally, they suggest that regional astrocyte heterogeneity may help to coordinate postnatal neural circuit refinement.
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Project description:Behavior arises from concerted activity throughout the brain. Consequently, a major focus of modern neuroscience is defining the physiology and behavioral roles of projection neurons linking different brain areas. Single-cell RNA sequencing has facilitated these efforts by revealing molecular determinants of cellular physiology and markers that enable genetically targeted perturbations such as optogenetics, but existing methods for sequencing defined projection populations are low throughput, painstaking, and costly. We developed a straightforward, multiplexed approach, virally encoded connectivity transgenic overlay RNA sequencing (VECTORseq). VECTORseq repurposes commercial retrogradely infecting viruses typically used to express functional transgenes (e.g., recombinases and fluorescent proteins) by treating viral transgene mRNA as barcodes within single-cell datasets. VECTORseq is compatible with different viral families, resolves multiple populations with different projection targets in one sequencing run, and identifies cortical and subcortical excitatory and inhibitory projection populations. Our study provides a roadmap for high-throughput identification of neuronal subtypes based on connectivity.
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Project description:Sensorimotor reflex circuits engage distinct neuronal subtypes, defined by precise connectivity, to transform sensation into compensatory behavior. Whether and how motor partner populations shape the subtype fate and connectivity of their pre-motor counterparts remains controversial. Here, we discovered that motor partners are dispensable for proper connectivity across an entire vestibular reflex circuit that stabilizes gaze. We first measured activity following vestibular sensation in pre-motor projection neurons after constitutive loss of their extraocular motor neuron partners.We observed normal responses and topography consistent with unchanged functional connectivity between sensory neurons and projection neurons. Next, we show that projection neurons remain anatomically and molecularly poised to connect appropriately with their motor partners. Lastly, we show that the transcriptional signatures of projection neuron subtypes develop independently of motor partners. Our findings comprehensively overturn a long-standing model: that connectivity in the circuit for gaze stabilization is retrogradely determined by motor partner-derived signals. By defining the contribution of motor neurons to canonical sensorimotor circuit assembly, our work speaks to comparable processes in spinal circuits and advances our understanding of general principles of neural development.
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Project description:Interconnectivity between neocortical areas is critical for sensory integration and sensorimotor transformations. These functions are mediated by heterogeneous interareal cortical projection neurons (ICPN), which send axon branches to distinct cortical areas as well as to subcortical targets. Although ICPN are anatomically diverse, they are molecularly homogeneous and how the diversity of their anatomical and functional features emerge during development remains largely unknown. Here, we address this question by linking connectome and transcriptome in developing single ICPN of the mouse neocortex using a combination of MAPseq mapping (to identify single-neuron axonal projections) and single-cell RNA sequencing (to identify corresponding gene expression). Focusing on neurons of the primary somatosensory cortex (S1), we reveal a protracted unfolding of the molecular and functional differentiation of motor cortex-projecting (M) compared to secondary somatosensory cortex-projecting (S2) ICPN. We identify SOX11 as a temporally differentially expressed transcription factor in M vs. S2 ICPN. Postnatal manipulation of SOX11 expression level in S1 impaired sensorimotor connectivity and selectively disrupted exploratory behavior in freely moving mice. Together, our results reveal that within a single cortical area, different subtypes of ICPN have distinct postnatal molecular differentiation paces, which is then reflected in distinct circuit connectivities and functions. Dynamic differences in expression levels of largely generic set of genes, rather than fundamental differences in the identity of developmental genetic programs, may thus account for emergence of intra-type diversity in cortical neurons.
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Project description:Interconnectivity between neocortical areas is critical for sensory integration and sensorimotor transformations. These functions are mediated by heterogeneous interareal cortical projection neurons (ICPN), which send axon branches to distinct cortical areas as well as to subcortical targets. Although ICPN are anatomically diverse, they are molecularly homogeneous and how the diversity of their anatomical and functional features emerge during development remains largely unknown. Here, we address this question by linking connectome and transcriptome in developing single ICPN of the mouse neocortex using a combination of MAPseq mapping (to identify single-neuron axonal projections) and single-cell RNA sequencing (to identify corresponding gene expression). Focusing on neurons of the primary somatosensory cortex (S1), we reveal a protracted unfolding of the molecular and functional differentiation of motor cortex-projecting (M) compared to secondary somatosensory cortex-projecting (S2) ICPN. We identify SOX11 as a temporally differentially expressed transcription factor in M vs. S2 ICPN. Postnatal manipulation of SOX11 expression level in S1 impaired sensorimotor connectivity and selectively disrupted exploratory behavior in freely moving mice. Together, our results reveal that within a single cortical area, different subtypes of ICPN have distinct postnatal molecular differentiation paces, which is then reflected in distinct circuit connectivities and functions. Dynamic differences in expression levels of largely generic set of genes, rather than fundamental differences in the identity of developmental genetic programs, may thus account for emergence of intra-type diversity in cortical neurons.
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Project description:A virally encoded high resolution screen of cytomegalovirus host dependencies
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Project description:Purpose:This work aimed to identify the genetic profiles of piriform projection neurons and characterize their spatial organization within the piriform cortex. Methods: We microdissected the three layers of pirifrom cortex by laser capture (LMD) and performed RNA deep sequencing in order to identify layer-specific molecular markers, we then validated these data by using RNA in situ hybridization and immunohistochemistry.We next performed anterograde neural tracing experiments to identify piriform target regions, and retrograde neural tracing experiments to analyze how piriform projection neurons are organized within piriform cortex.We then combined the analysis of patterns of gene expression with retrograde tracing experiments to identify molecular signatures of the different subclasses of piriform projecting neurons. Results:We show that layers and sub-layers of the piriform cortex can be discriminated by gene expression patterns in adult piriform cortex. We observe that neurons projecting to distinct target areas are localized in distinct layers and express specific genes. We demonstrate that these molecular signatures of piriform projection neurons are maintained in reeler mice, in which cortical lamination is lost and neural positioning is scrambled, suggesting that piriform output connectivity strictly depends on the molecular programm, rather than a proper lamination of the cortex. Conclusion:These results provide important insights into the principles underling the piriform connectivity.
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