Biology subjects

Vroman, R.

Publications and source records attributed to Vroman, R..

5 recordsLinked to original sources

bioRxiv2024

Reduced Capsaicin-Induced Mechanical Allodynia and Neuronal Responses in the DRG in the Presence of Shp1 Overexpression

Transient Receptor Potential Vanilloid 1 (TRPV1) is a nonselective cation channel expressed by pain-sensing neurons and has been an attractive target for the development of drugs to treat pain. Recently, Src homology region 2 domain-containing phosphatase-1 (SHP-1) was shown to dephosphorylate TRPV1 in dorsal root ganglia (DRG) neurons, which was linked with alleviating different pain phenotypes. These previous studies were performed in male rodents only and did not directly investigate the role of SHP-1 in TRPV-1 mediated sensitization. Therefore, our goal was to determine the impact of Shp1 overexpression on TRPV1-mediated neuronal responses and capsaicin-induced pain behavior in mice of both sexes. Twelve-week-old male and female mice overexpressing Shp1 (Shp1-Tg) and their wild type (WT) littermates were used. Shp1 overexpression was confirmed in the DRG of Shp1-Tg mice by RNA in situ hybridization and RT-qPCR. Trpv1 and Shp1 were found to be co-expressed in DRG sensory neurons in both genotypes. Functionally, this overexpression resulted in lower magnitude intracellular calcium responses to 200 nM capsaicin stimulation in DRG cultures from Shp1-Tg mice compared to WTs. In vivo, we tested the effects of Shp1 overexpression on capsaicin-induced pain through a model of capsaicin footpad injection. While capsaicin injection evoked nocifensive behavior (paw licking) and paw swelling in both genotypes and sexes, only WT mice developed mechanical allodynia after capsaicin injection. We observed similar level of TRPV1 protein expression in the DRG of both genotypes, however, a higher amount of tyrosine phosphorylated TRPV1 was detected in WT DRG. These experiments suggest that, while SHP-1 does not mediate the acute swelling and nocifensive behavior induced by capsaicin, it does mediate a protective effect against capsaicin-induced mechanical allodynia in both sexes. The protective effect of SHP-1 might be mediated by TRPV1 dephosphorylation in capsaicin-sensitive sensory neurons of the DRG.

neuroscience↗

bioRxiv2022

The matrisome of the murine and human dorsal root ganglion: a transcriptomal approach

The extracellular matrix (ECM) is a dynamic structure composed of a large number of molecules that can be divided into six different categories and are collectively called the matrisome. The ECM plays pivotal roles in physiological and pathological processes in many tissues, including the nervous system. Intriguingly, alterations in ECM molecules/pathways are often associated with painful human conditions and murine experimental pain models. Nevertheless, mechanistic insight into the interplay of normal or defective ECM and pain is largely lacking. To expand the knowledge on ECM composition and synthesis in the peripheral nervous system, we used a transcriptomal approach to investigate the expression and cellular origin of matrisome genes in murine and human dorsal root ganglia (DRG), containing the cell bodies of sensory neurons. Bulk RNA sequencing data showed that over 60% of all matrisome genes were expressed in both murine and human DRG, with proportionally more core matrisome genes (glycoproteins, collagens, and proteoglycans) expressed compared to matrisome-associated genes (ECM-affiliated genes, ECM regulators and secreted factors). Examination of the cellular origin of matrisome expression by single cell RNA sequencing on murine DRG revealed that core matrisome genes, especially collagens, were expressed by vascular leptomeningeal-like (fibroblast) cell types whereas matrisome-associated genes were mainly expressed by neuronal cell types. We analyzed cell-cell communication networks with the CellChat R package and predicted an important role for the Collagen signaling pathway in connecting vascular cell types and nociceptors in murine tissue, which we confirmed by analysis of spatial transcriptomic data from human DRG. RNAscope in situ hybridization and immunohistochemistry confirmed expression of collagens in fibroblasts surrounding nociceptors in human DRG. This study supports the idea that the DRG matrisome may contribute to neuronal signaling in both mouse and human. The identification of the cellular distribution of murine and human matrisome genes provides a framework to study the role of the ECM in peripheral nervous tissue and its effects on pain signaling. HighlightsO_LITranscriptomal analyses of mouse and human dorsal root ganglia (DRG) revealed that over 60% of matrisome genes are expressed by murine and human dorsal root ganglia (DRG), with over 85% of the genes with orthologues overlapping between both species. C_LIO_LIMatrisome-associated genes had the highest expression in both species and included conserved expression of annexins, S100 calcium binding proteins and cathepsins. C_LIO_LICollagens and collagen receptors are expressed by distinct cell types in murine and human DRG, suggesting that the collagen signaling pathway could be involved in cell-cell signaling. C_LI

neuroscience↗

bioRxiv2021

The output of interneurons in the primary visual cortex is best reflected by pre-synaptic activity, not somatic activity

Recent advances in 2-photon calcium-imaging in awake mice have made it possible to study the effect of different behavioural states on cortical circuitry. Many studies assume that somatic activity can be used as a measure for neuronal output. We set out to test the validity of this assumption by comparing somatic activity with the pre-synaptic activity of VIP (Vasoactive intestinal peptide)- and SST (Somatostatin)-positive interneurons in layer 2/3 of the primary visual cortex (V1). We used mice expressing genetically encoded calcium indicators in VIP/SST-interneurons across the whole cell (VIP/SST:GCaMP6f) or confined to pre-synapses (VIP/SST:SyGCaMP5). Mice were exposed to a full-field visual stimulation protocol consisting of 60-second-long presentations of moving Gabor gratings (0.04 cpd, 2 Hz) alternated by 30 seconds of grey screen. During imaging, mice were placed on an air-suspended Styrofoam ball, allowing them to run voluntarily. We compared neural activity during three 4-second time-windows: Before visual stimulation (-4 to 0 sec), during the initial onset (1 to 5 sec) and at the end of the stimulation (56 to 60 sec.). These were further compared while the mice were stationary and while they were voluntarily locomoting. Unlike VIP-somas, VIP-pre-synapses showed strong suppressive responses to the visual stimulus. Furthermore, VIP-somas were positively correlated with locomotion, whereas in VIP-synapses we observed a split between positive and negative correlations. In addition, a similar but weaker distinction was found between SST-somas and pre-synapses. The excitatory effect of locomotion in VIP-somas increased over the course of the visual stimulus but this property was only shared with the positively correlated VIP-pre-synapses. The remaining negatively correlated pre-synapses showed no relation to the overall activity of the Soma. Our results suggest that when making statements about the involvement of interneurons in V1 layer 2/3 circuitry it is crucial to measure from synaptic terminals as well as from somas.

neuroscience↗

bioRxiv2019

Correction of z-motion artefacts to allow population imaging of synaptic activity in awake behaving mice

Functional imaging of head-fixed, awake, behaving mice using two-photon imaging of fluorescent activity reporters has become a powerful tool in the studying the function of the brain. Motion artefacts are an inevitable problem during such experiments and are routinely corrected for in x and y dimensions. However, axial (z) shifts of several microns can also occur, leading to intensity fluctuations in structures such as synapses that are small compared to the axial point-spread function of the microscope. Here we present a simple strategy to correct z-motion artefacts arising over the course of a time-series experiment in a single optical plane. Displacement in z was calculated using dye-filled blood vessels as an anatomical marker, providing high contrast images and accuracy to within [~]0.1 {micro}m. The axial profiles of ROIs corresponding to synapses were described using a Moffat function and this \"ROI-spread function\" used to correct activity traces on an ROI-by-ROI basis. We demonstrate the accuracy and utility of the procedures in simulation experiments using fluorescent beads and then apply them to correcting measurements of synaptic activity in populations of vasoactive-intestinal peptide (VIP) interneurons expressing the synaptic reporter SyGCaMP6f. Correction of z-motion artefacts had a substantial impact on the apparent correlation between synaptic activity and running speed, demonstrating the importance of correcting for these artefacts for the interpretation of in vivo imaging experiments in awake mice.\n\nSummary of Key PointsO_LIMotion artefacts associated with motor behaviour are an inevitable problem of multiphoton imaging in awake behaving animals, particularly imaging synapses.\nC_LIO_LICorrection of axial motion usually requires volumetric imaging resulting in slower rates of acquisition.\nC_LIO_LIWe describe a method that is easy to implement to correct z-motion artefacts that allows population imaging of synaptic activity while scanning a single plane in a standard multiphoton microscope.\nC_LIO_LIThe method uses a reference volume acquired in two colour channels - an activity reporter and an anatomical marker of blood vessels. The procedure estimates the z-displacement in every frame and applies an intensity correction in which the z point-spread function for each synapse is modelled as a Moffat function.\nC_LIO_LIWe demonstrate that the method allows synaptic calcium activity signals to be collected from populations of synaptic boutons in mouse primary visual cortex during locomotion.\nC_LI

neuroscience↗

bioRxiv2019

Disentangling neuronal inhibition and inhibitory pathways in the lateral habenula

The lateral habenula (LHb) is hyperactive in depression, and thus potentiating inhibition of this structure makes an interesting target for future antidepressant therapies. However, the circuit mechanisms mediating inhibitory signalling within the LHb are not well-known. We addressed this issue by studying LHb neurons expressing either parvalbumin (PV), neuron-derived neurotrophic factor (Ndnf) or somatostatin (SOM), three markers of particular sub-classes of neocortical inhibitory neurons. While we report that Ndnf is not representative of any particular sub-population of LHb neuron, we find that both PV and SOM are expressed by physiologically distinct sub-classes. Furthermore, we describe multiple sources of inhibitory input to the LHb arising from both local PV-positive neurons, and from PV-positive neurons in the medial dorsal thalamic nucleus, and from SOM-positive neurons in the ventral pallidum. These findings hence provide new insight into inhibitory control within the LHb, and highlight that this structure is more neuronally diverse than previously thought. SummaryThe lateral habenula receives inhibitory input from three distinct sources: from local PV-positive neurons, from PV-positive neurons in the medial dorsal thalamic nucleus (MDT); and from SOM-positive neurons in the ventral pallidum (VP). O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=178 SRC="FIGDIR/small/633271v2_ufig1.gif" ALT="Figure 1"> View larger version (49K): org.highwire.dtl.DTLVardef@8df117org.highwire.dtl.DTLVardef@cf226aorg.highwire.dtl.DTLVardef@dc94f5org.highwire.dtl.DTLVardef@1b76a7f_HPS_FORMAT_FIGEXP M_FIG C_FIG Significance statementThe circuitry by which inhibitory signalling is processed within the lateral habenula is currently not well understood; yet this is an important topic as inhibition of the lateral habenula has been shown to have antidepressant efficacy. We therefore investigated inhibitory signalling mechanisms within the lateral habenula by studying input neurons expressing markers commonly associated with inhibitory identity. We identity sources of inhibitory input from both local neurons, and arising from neurons in the medial dorsal thalamic nucleus and ventral pallidum. ContributionsJ.F.W. performed the experiments. R.V. contributed to experiments. J.F.W. analysed the data. K.B. and P.W. designed and performed the in situ hybridisation experiments. C.W. designed and supervised the study, and helped J.F.W write the manuscript. R.V. and S.S. contributed to the manuscript and discussions.

neuroscience↗