Project description:Primary objectives: Characterization of the macrophage population subset that is modulated by enteric neurons
Primary endpoints: Characterization of the macrophage population subset that is modulated by enteric neurons via RNA sequencing
Project description:In this project, we cultured rat hippocampal neurons in mechanically different environments and studied electrical maturation. We compared WT neurons with two different Piezo1 knockdown conditions. These two knockdown conditions were generated in two independent CRISPR-Cas9 KD assays. Each assay is based on four different CRISPR-Cas9 guides targeting the Piezo1 gene. RNA sequencing was used to analyse the pathway leading to the stiffness dependent maturation behaviour.
Project description:Recent findings have revealed the complexity of the transcriptional landscape in mammalian cells. One recently described class of novel transcripts are the Cytoplasmic Intron-sequence Retaining Transcripts (CIRTs), hypothesized to confer post-transcriptional regulatory function. For instance, the neuronal CIRT KCNMA1i16 contributes to the firing properties of hippocampal neurons. We hypothesized that CIRTs may be present in a broad set of transcripts and comprise novel signals for post-transcriptional regulation. We carried out a transcriptome-wide survey of CIRTs by sequencing micro-dissected subcellular RNA fractions. Two batches of 150-300 individually dissected dendrites from primary cultures of hippocampal neurons in rat and three batches from mouse hippocampal neurons were sequenced. After statistical processing to minimize artifacts, we found a broad prevalence of CIRTs in the neurons in both species (44-60% of the expressed transcripts). The analysis for CIRTs was also carried out by sequencing single cells from mouse brown adipose tissue and mouse cardiomyocytes. There was widespread prevalence of CIRTs in all of the cell types. Single cell samples were aRNA amplified and sequenced using Illumina GA Analyzer II and Illumina Hiseq 2000
Project description:Poly(A) RNA profiling upon Gld2 knockdown in cultured hippocampal neurons Neurons transduced with scrambled and Gld2 knowdown shRNA
Project description:We conducted RNA-sequencing of lidocaine hydrochloride in treating rat dorsal root ganglion neurons to detect lidocaine’s effect of transcriptome profiling changes compared with control.
Project description:We established a neuron-specific Argonaute2:GFP-RNA immunoprecipitation followed by high throughput sequencing (AGO2-RIP-seq) to analyse the regulatory role of miRNAs in mouse hippocampal neurons. Using this technique, we identified more than two thousand miRNA target genes in hippocampal neurons, regulating essential neuronal features such as axon guidance and transcription. Furthermore, we found that stable inhibition of the highly expressed miR-124 in hippocampal neurons led to significant changes in the AGO2 binding of target mRNAs, resulting in subsequent upregulation of numerous miRNA target genes. Our data suggest that target redundancies are common among microRNA families. Together, these findings greatly enhance our understanding of the mechanisms and dynamics through which miRNAs regulate their target genes in neurons. Analysis of the miRNA targetome in hippocampal neurons after inhibition of 2 different miRNAs. AAV5 injections into the hippocampus of adult C57BL/6 mice producing either of the following under a synapsin promoter: GFP only (Samples beginning with 'GFP124…' or 'GFP125…'), GFP-miR124sp (Samples beginning with 'miR124…'), GFP-miR125sp (Samples beginning with 'miR125…'), GFP-AGO2-miR292sponge (samples ending with '…292'), GFP-AGO2-miR124sponge (samples ending with '…124'), GFP-AGO2-miR125sponge (samples ending with '…125'). All other samples were sham-injected.
Project description:We report that local application of Abeta(1-42) for 24h triggers changes in the localized transcriptome, while the transcriptome in cell bodies remains largely unchanged. Axonal and somatic mRNA profiles of rat embryonic hippocampal neurons grown in microfluidic chambers were generated by RNA-Seq in duplicates using an Illumina MiSeq. Axons only were treated for 24 h with soluble oligomeric Abeta(1-42) or vehicle control.
