An individual interneuron participates in many kinds of inhibition and spans much of the mouse visual thalamus, bioRxiv, 2019-06-27
SUMMARYIn principle, one way to define the functional role of a neuron would be to identify all the synaptic input it receives and all synaptic output it confers onto its targets. With serial electron microscopy we annotated all the input synapses (862) and output synapses (626) associated with one inhibitory interneuron in the visual thalamus of the mouse. This neuron’s neurites covered a broad swath of lateral geniculate nucleus and spanned multiple functionally distinct regions. Every one of its neurites formed synapses onto hundreds of thalamocortical cells of several different types. All but one small neurite also had dendrite-like properties and received input from retinal ganglion cell axons. Pre- and postsynaptic associations with other inhibitory interneurons were also distributed throughout the interneuron’s territory. We observed a diverse array of local synaptic motifs and three fundamentally different types of inhibitory neurites. Many thalamocortical cells were innervated weakly by this interneuron by single en passant shaft synapses. But a subset of the interneuron’s thalamocortical cell targets received multiple synaptic inputs from targeted inhibitory neurites that climbed along the thalamocortical cell’s dendrite with an assemblage of fasciculated retinal ganglion cell axons. Because of the diverse range of synaptic relationships exhibited by this one neuron, this cell defies a single functional label and seems rather to be using extremely local synaptic processing to participate in many different functions.
biorxiv neuroscience 100-200-users 2019Astrocytes Contribute to Remote Memory Formation by Modulating Hippocampal-Cortical Communication During Learning, bioRxiv, 2019-06-27
ABSTRACTThe consolidation and retrieval of remote memories depend on the coordinated activity of the hippocampus and frontal cortices. However, the exact time at which these regions are recruited to support memory and the interactions between them are still debated. Astrocytes can sense and modify neuronal activity with great precision, but their role in cognitive function has not been extensively explored. To investigate the role of astrocytes in remote memory we expressed the Gi-coupled receptor hM4Di in CA1 astrocytes, allowing their manipulation by a designer drug. We discovered that astrocytic modulation during learning resulted in a specific impairment in remote, but not recent, memory recall, accompanied by decreased neuronal activity in the anterior cingulate cortex (ACC) during retrieval. We revealed a massive recruitment of ACC-projecting neurons in CA1 during memory acquisition, accompanied by activation of ACC neurons. Astrocytic Gi activation disrupted CA3 to CA1 communication in-vivo, and reduced the downstream response in the ACC. This same manipulation in behaving mice induced a projection-specific inhibition of ACC-projecting CA1 neurons during learning, consequently preventing the recruitment of the ACC. Our findings suggest that the foundation of remote memory is established in the ACC during acquisition, engaging a distinct process from the one supporting consolidation of recent memory. Furthermore, the mechanism underlying remote memory involves projection-specific functions of astrocytes in regulating neuronal activity.
biorxiv neuroscience 100-200-users 2019Dissociation of solid tumour tissues with cold active protease for single-cell RNA-seq minimizes conserved collagenase-associated stress responses, bioRxiv, 2019-06-27
AbstractBackgroundSingle-cell RNA sequencing (scRNAseq) is a powerful tool for studying complex biological systems, such as tumour heterogeneity and tissue microenvironments. However, the sources of technical and biological variation in primary solid tumour tissues and patient-derived mouse xenografts for scRNAseq, are not well understood. Here, we used low temperature (6°C) protease and collagenase (37°C) to identify the transcriptional signatures associated with tissue dissociation across a diverse scRNAseq dataset comprising 128,481 cells from patient cancer tissues, patient-derived breast cancer xenografts and cancer cell lines.ResultsWe observe substantial variation in standard quality control (QC) metrics of cell viability across conditions and tissues. From FACS sorted populations gated for cell viability, we identify a sub-population of dead cells that would pass standard data filtering practices, and quantify the extent to which their transcriptomes differ from live cells. We identify a further subpopulation of transcriptomically “dying” cells that exhibit up-regulation of MHC class I transcripts, in contrast with live and fully dead cells. From the contrast between tissue protease dissociation at 37°C or 6°C, we observe that collagenase digestion results in a stress response. We derive a core gene set of 512 heat shock and stress response genes, including FOS and JUN, induced by collagenase (37°C), which are minimized by dissociation with a cold active protease (6°C). While induction of these genes was highly conserved across all cell types, cell type-specific responses to collagenase digestion were observed in patient tissues. We observe that the yield of cancer and non-cancer cell types varies between tissues and dissociation methods.ConclusionsThe method and conditions of tumour dissociation influence cell yield and transcriptome state and are both tissue and cell type dependent. Interpretation of stress pathway expression differences in cancer single cell studies, including components of surface immune recognition such as MHC class I, may be especially confounded. We define a core set of 512 genes that can assist with identification of such effects in dissociated scRNA-seq experiments.
biorxiv genomics 200-500-users 2019DNA-loop extruding condensin complexes can traverse one another, bioRxiv, 2019-06-27
Condensin, a key member of the Structure Maintenance of Chromosome (SMC) protein complexes, has recently been shown to be a motor that extrudes loops of DNA1. It remains unclear, however, how condensin complexes work together to collectively package DNA into the chromosomal architecture. Here, we use time-lapse single-molecule visualization to study mutual interactions between two DNA-loop-extruding yeast condensins. We find that these one-side-pulling motor proteins are able to dynamically change each other’s DNA loop sizes, even when located large distances apart. When coming into close proximity upon forming a loop within a loop, condensin complexes are, surprisingly, able to traverse each other and form a new type of loop structure, which we term Z loop – three double-stranded DNA helices aligned in parallel with one condensin at each edge. These Z-loops can fill gaps left by single loops and can form symmetric dimer motors that reel in DNA from both sides. These new findings indicate that condensin may achieve chromosomal compaction using a variety of looping structures.
biorxiv biophysics 100-200-users 2019Increasing the efficiency of long-read sequencing for hybrid assembly with k-mer-based multiplexing, bioRxiv, 2019-06-25
AbstractHybrid genome assembly has emerged as an important technique in bacterial genomics, but cost and labor requirements limit large-scale application. We present Ultraplexing, a method to improve per-sample sequencing cost and hands-on-time of Nanopore sequencing for hybrid assembly by at least 50%, compared to molecular barcoding while maintaining high assembly quality (Quality Value; QV ≥ 42). Ultraplexing requires the availability of Illumina data and uses inter-sample genetic variability to assign reads to isolates, which obviates the need for molecular barcoding. Thus, Ultraplexing can enable significant sequencing and labor cost reductions in large-scale bacterial genome projects.
biorxiv bioinformatics 100-200-users 2019What is the test-retest reliability of common task-fMRI measures? New empirical evidence and a meta-analysis, bioRxiv, 2019-06-25
AbstractIdentifying brain biomarkers of disease risk is a growing priority in neuroscience. The ability to identify meaningful biomarkers is limited by measurement reliability; unreliable measures are unsuitable for predicting clinical outcomes. Measuring brain activity using task-fMRI is a major focus of biomarker development; however, the reliability of task-fMRI has not been systematically evaluated. We present converging evidence demonstrating poor reliability of task-fMRI measures. First, a meta-analysis of 90 experiments (N=1,008) revealed poor overall reliability (mean ICC=.397). Second, the test-retest reliabilities of activity in a priori regions of interest across 11 common fMRI tasks collected in the context of the Human Connectome Project (N=45) and the Dunedin Study (N=20) were poor (ICCs=.067-.485). Collectively, these findings demonstrate that common task-fMRI measures are not currently suitable for brain biomarker discovery or individual differences research. We review how this state of affairs came to be and highlight avenues for improving task-fMRI reliability.
biorxiv neuroscience 200-500-users 2019