Biology subjects

Wyart, C.

Publications and source records attributed to Wyart, C..

23 records · Page 2Linked to original sources

bioRxiv2019

Sensory neurons contacting the cerebrospinal fluid require the Reissner fiber to detect spinal curvature in vivo

Recent evidence indicate active roles for the cerebrospinal fluid (CSF) on body axis development and morphogenesis of the spine implying CSF-contacting neurons (CSF-cNs) in the spinal cord. CSF-cNs project a ciliated apical extension into the central canal that is enriched in the channel PKD2L1 and enables the detection of spinal curvature in a directional manner. Dorsolateral CSF-cNs ipsilaterally respond to lateral bending while ventral CSF-cNs respond to longitudinal bending. Historically, the implication of the Reissner fiber (RF), a long extracellular thread in the CSF, to CSF-cN sensory functions has remained a subject of debate. Here, we reveal using electron microscopy in zebrafish larvae that the RF is in close vicinity with cilia and microvilli of ventral and dorsolateral CSF-cNs. We investigate in vivo the role of cilia and the Reissner fiber in the mechanosensory functions of CSF-cNs by combining calcium imaging with patch-clamp recordings. We show that disruption of cilia motility affects CSF-cN sensory responses to passive and active curvature of the spinal cord without affecting the Pkd2l1 channel activity. Since ciliary defects alter the formation of the Reissner fiber, we investigated whether the Reissner fiber contributes to CSF-cN mechanosensitivity in vivo. Using a hypomorphic mutation in the scospondin gene that forbids the aggregation of SCO-spondin into a fiber, we demonstrate in vivo that the Reissner fiber per se is critical for CSF-cN mechanosensory function. Our study uncovers that neurons contacting the cerebrospinal fluid functionally interact with the Reissner fiber to detect spinal curvature in the vertebrate spinal cord. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=152 SRC="FIGDIR/small/861344v1_ufig1.gif" ALT="Figure 1"> View larger version (42K): org.highwire.dtl.DTLVardef@3c7baaorg.highwire.dtl.DTLVardef@db094forg.highwire.dtl.DTLVardef@4041c6org.highwire.dtl.DTLVardef@9e3dae_HPS_FORMAT_FIGEXP M_FIG C_FIG eToCThe role of the Reissner fiber, a long extracellular thread running in the cerebrospinal fluid (CSF), has been since its discovery in 1860 a subject of debate. Orts-DelImmagine et al. report that the Reissner fiber plays a critical role in the detection of spinal curvature by sensory neurons contacting the CSF. HighlightsO_LISince its discovery, the role of the Reissner fiber has long been a subject of debate C_LIO_LIMechanoreception in CSF-contacting neurons (CSF-cNs) in vivo requires the Reissner fiber C_LIO_LICSF-cN apical extension is in close vicinity of the Reissner fiber C_LIO_LICSF-cNs and the Reissner fiber form in vivo a sensory organ detecting spinal curvature C_LI

neuroscience↗

bioRxiv2019

Simultaneous fluorescence imaging of tilted focal planes at two depths in thick neural tissue: Implementation with remote focus in a widefield electrophysiological microscope.

Wide-field imaging conventionally results in a single image plane oriented perpendicular to the optical axis. However, in brain slice or in vivo recording, neuronal or circuit morphologies lie in arbitrarily tilted planes. Consequently the spatiotemporal advantages of wide-field non-scanned imaging are lost because of the time required for stepwise focal readjustments to view an entire neuron or network. We describe an application of remote focus that views simultaneously two planes separated by up to 100 {micro}m, each with variable tilt from the conventional image plane. This permits fluorescence detection of ion fluxes or membrane potential across neuronal compartments and their correlation with electrical activity. Further, two fluorophores can be viewed simultaneously in each plane. We show (i) neuronal images tilted to optimise simultaneous aquisition of somatic, dendritic and axonal compartments; (ii) networks viewed simultaneously at 2 depths separated by up to 100 {micro}m, (iii) widefield imaging at 30 Hz of Gcamp5 fluorescence during spontaneous spiking in motoneuron layers of zebrafish spinal cord separated by 30-40 microns.

neuroscience↗

bioRxiv2019

Origin of bidirectionality of cerebrospinal fluid flow and impact on long-range transport between brain and spinal cord

The circulation of cerebrospinal fluid (CSF) plays pivotal roles for body axis formation and brain development. During embryogenesis, CSF is rich in particles and proteins and flows bidirectionally in the central canal. The origins of bidirectional flow and its impact on development are unknown. Experiments combined with modeling and simulations demonstrate that the bidirectionality of CSF flow is generated locally by caudally-polarized motile cilia confined to the ventral wall of the central canal. Such active bidirectional flow of the CSF accelerates the long-range transport of particles propagating rostrally and caudally. In addition, spontaneous muscle contractions increase local CSF flow and consequently enhance long-range transport of extracellular lipidic particles. Focal ablation of the channel connecting brain ventricles to the central canal reduces embryo length, indicating that long-range transport contributes to embryonic growth. Our study also demonstrates that at this early stage, motile cilia ensure the proper formation of the central canal.

biophysics↗

bioRxiv2018

Pkd2l1 is required for mechanoception in cerebrospinal fluid-contacting neurons and maintenance of spine curvature

Flow of cerebrospinal fluid (CSF) may contribute to spine morphogenesis, as mutations affecting both cilia motility and CSF flow lead to scoliosis1. However, the mechanisms underlying detection of the CSF flow in the central canal of the spinal cord remain elusive. Here we used full-field optical coherence tomography (FF-OCT) and bead tracking to demonstrate that CSF flows bidirectionally along the antero-posterior axis in the central canal of zebrafish embryos. In the zebrafish mutant cfap298tm304, previously known as kurly, reduction of cilia motility slows transport down the length of central canal. To investigate downstream mechanisms that could transduce CSF flow, we performed calcium imaging in sensory neurons contacting the CSF (CSF-cNs) and found that disruption in cilia motility impaired the activity of CSF-cNs. CSF-cNs across species express the transient receptor potential channel PKD2L1, also known as TRPP3, which contributes to CSF-cN chemosensory properties. Using calcium imaging and whole-cell patch clamp recordings, we found that the loss of the Pkd2l1 channel in pkd2l1 mutant embryos also abolished CSF-cN activity. Whole-cell recordings further demonstrated that opening of a single channel is sufficient to trigger action potentials in wild type CSF-cNs. Recording from isolated cells in vitro, we showed that CSF-cNs are mechanosensory cells that respond to pressure in a Pkd2l1-dependent manner. Interestingly, adult pkd2l1 mutant zebrafish develop an exaggerated spine curvature, reminiscent of kyphosis in humans. Our study indicates that CSF-cNs are mechanosensory cells whose spontaneous activity reflects CSF flow in vivo. Furthermore, Pkd2l1 in CSF-cNs contributes to the maintenance of the natural curvature of the spine.

neuroscience↗

bioRxiv2017

Multiplexed temporally focused light shaping for high-resolutionmulti-cell targeting

Patterning light at the single-cell level over multiple neurons in the brain is crucial for optogenetic photostimulation that can recapitulate natural activity patterns and, thereby, determine the role of specific components of brain activity in behavior. To this end we have developed a method for projecting three-dimensional, 2-photon excitation patterns that are confined to many individual neurons. The new versatile optical scheme generates multiple extended excitation spots in a large volume with micrometric lateral and axial resolution. Two-dimensional temporally focused shapes are multiplexed several times over selected positions, thanks to the precise spatial phase modulation of the pulsed beam. This permits, under multiple configurations, the generation of tens of axially confined spots in an extended volume, spanning a range in depth of up to 500 m. We demonstrate the potential of the approach by performing multi-cell volumetric excitation of photoactivatable GCaMP in the central nervous system of Drosophila larvae, a challenging structure with densely arrayed and small diameter neurons, and by photoconverting the fluorescent protein Kaede in zebrafish larvae. Our technique paves the way for the optogenetic manipulation of a large number of neurons in intact circuits.

neuroscience↗