ABSTRACT: Using microdissected dendrites from primary cultures of hippocampal neurons of two mouse strains (C57BL/6 and Balb/c) and one rat strain (Sprague-Dawley), we investigate via microarrays, subcellular localization of mRNAs in dendrites of neurons to assay the evolutionary differences in subcellular dendritic transcripts localization.
Project description:Using microdissected dendrites from primary cultures of hippocampal neurons of two mouse strains (C57BL/6 and Balb/c) and one rat strain (Sprague-Dawley), we investigate via microarrays, subcellular localization of mRNAs in dendrites of neurons to assay the evolutionary differences in subcellular dendritic transcripts localization.
Project description:Using microdissected dendrites from primary cultures of hippocampal neurons of two mouse strains (C57BL/6 and Balb/c) and one rat strain (Sprague-Dawley), we investigate via microarrays, subcellular localization of mRNAs in dendrites of neurons to assay the evolutionary differences in subcellular dendritic transcripts localization. To assess neuronal dendrite expression divergence between mice and rats, we used the Affymetrix array platform to assay the transcriptomes of micro-dissected individual dendrites of hippocampal neurons in dispersed primary cell cultures from Sprague-Dawley rat (9 biological replicates), C57BL/6 mouse (14 biological replicates), and Balb/c mouse (5 biological replicates)
Project description:Using microdissected dendrites from primary cultures of hippocampal neurons of two mouse strains (C57BL/6 and Balb/c) and one rat strain (Sprague-Dawley), we investigate via microarrays, subcellular localization of mRNAs in dendrites of neurons to assay the evolutionary differences in subcellular dendritic transcripts localization. To assess neuronal dendrite expression divergence between mice and rats, we used the Affymetrix array platform to assay the transcriptomes of micro-dissected individual dendrites of hippocampal neurons in dispersed primary cell cultures from Sprague-Dawley rat (9 biological replicates), C57BL/6 mouse (14 biological replicates), and Balb/c mouse (5 biological replicates)
Project description:To assess neuronal expression divergence between mice and rats, we used the Affymetrix array platform to assay the transcriptomes of micro-dissected individual soma and pool of dendrites of hippocampal neurons in dispersed primary cell cultures from rat and mouse. Using microdissected soma and dendrites from primary cultures of hippocampal neurons of two mouse strains (C57BL/6 and Balb/c) and one rat strain (Sprague-Dawley), we investigate via microarrays, subcellular localization of mRNAs in neurons
Project description:To assess neuronal expression divergence between mice and rats, we used the Affymetrix array platform to assay the transcriptomes of micro-dissected individual soma and pool of dendrites of hippocampal neurons in dispersed primary cell cultures from rat and mouse. Using microdissected soma and dendrites from primary cultures of hippocampal neurons of two mouse strains (C57BL/6 and Balb/c) and one rat strain (Sprague-Dawley), we investigate via microarrays, subcellular localization of mRNAs in neurons
Project description:In this study we performed high throughput microfluidic qPCR using BioMark on RNA extracted from single neurons microdissected from DMV and NA tissue of one male Sprague Dawley rat.
Project description:Neurons transport a variety of mRNAs to their dendritic and axonal compartments where they are locally translated. These processes are essential for the functional specialization of these cellular structures. To explore both the intercellular variability and intracellular dynamics of neuronal compartments we characterized the somatic and dendritic transcriptome of single neurons. We used Laser Capture Microdissection to isolate the dendrites and soma of individual neurons from primary hippocampal cultures, followed by scRNA-seq to profile the mRNA content of these compartments. In total, we sequence 276 dendrites and their respective somata, 227 somata without the dendrites, and 16 negative controls (LCM cuts of areas without cellular content). We observed less cell-type specific differences in the dendritic transcriptome than in the somatic transcriptome. However, the dendritic localization of a subset of mRNAs does vary in a cell-type specific manner. Furthermore, we observed that the relation between somatic and dendritic abundance varies according to gene and is strongly influenced by cellular function. This study provides a framework for the regulation of the dendritic transcriptome within and across individual neurons.
Project description:To compare the RNAs present in dendrites and somas of individual neurons, we manually separated the dendrites and soma of primary mouse hippocampal neurons using a micropipette and performed RNA-sequencing on each subcellular fraction such that we obtained the subcellular transcriptomes of the same cell.
Project description:The subcellular localization and translation of messenger RNA (mRNA) supports functional differentiation between cellular compartments. In neuronal dendrites, local translation of mRNA provides a rapid and specific mechanism for synaptic plasticity and memory formation, and might be involved in the pathophysiology of certain brain disorders. Despite the importance of dendritic mRNA translation, little is known about which mRNAs can be translated in dendrites in vivo and when their translation occurs. Here we collect ribosome-bound mRNA from the dendrites of CA1 pyramidal neurons in the adult mouse hippocampus. We find that dendritic mRNA rapidly associates with ribosomes following a novel experience consisting of a contextual fear conditioning trial. High throughput RNA sequencing followed by machine learning classification reveals an unexpected breadth of ribosome-bound dendritic mRNAs, including mRNAs expected to be entirely somatic. Our findings are in agreement with a mechanism of synaptic plasticity that engages the acute local translation of functionally diverse dendritic mRNAs. RNA-Seq of ribosome-bound mRNA immunoprecipitated from dendrites and soma of CA1 pyramidal neurons in the mouse hippocampus
Project description:Mammalian target of rapamycin (mTOR) is implicated in synaptic plasticity and local translation in dendrites. Here we found that the mTOR inhibitor, rapamycin, increased the Kv1.1 voltage-gated potassium channel protein in hippocampal neurons and promoted Kv1.1 surface expression on dendrites without altering its axonal expression. Moreover, endogenous Kv1.1 mRNA was detected in dendrites. Using Kv1.1 fused to the photo-convertible fluorescence protein Kaede as a reporter for local synthesis, we observed Kv1.1 synthesis in dendrites upon inhibition of mTOR or the N-methyl-D-aspartate (NMDA) glutamate receptor. Thus, synaptic excitation may cause local suppression of dendritic Kv1 channels by reducing their local synthesis. Keywords: comparison of synaptosomal mRNA to total hippocampal mRNA