SpiCCI

Spinal Cord and Cerebro-Spinal Fluid Interface

Our research project aims at characterizing at the morphological and functional level the properties of a unique neuronal population present around the central canal (CC) and in contact with the Cerebro-Spinal Fluid (CSF) in the spinal cord of mammals. These CSF-contacting neurons (CSF-cNs) had been known for decades, are present in all vertebrate but their physiological role in the mammalian Central Nervous System (CNS) remains largely elusive.

Our ultimate goal is to determine how this unique neuronal population is functionally inserted around the CC and in spinal cord networks and to identify their physiological roles in mammals.

Medullo-spinal CSF-cNs are present along the entire length of the CC axis, are mainly GABAergic and selectively express the Polycystin subtype (also known as polycystin kidney disease 2-like 1, PKD2L1) of the ‘Transient Receptor Potential (TRP)’ superfamily. This channel represents a hallmark for this neuronal population since PKD2L1 expressing CSF-cNs are the only neurons projecting to the CC. CSF-cNs exhibit a typical and conserved morphology with a dendrite projecting to the CC ended in its lumen with a ciliated protrusion (or bud). They are polarized cells with an axon extending through the parenchyma. However, the territories innervated by CSF-cNs axons remain to be identified in mammals.

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Using patch-clamp recording techniques in acute brainstem and spinal cord slices, our group was among the first to characterize the electrophysiological properties of medullo-spinal CSF-cNs in mice. We revealed that CSF-cNs present, in whole-cell configuration, spontaneous PKD2L1 channel activity at the level of the unitary current. This channel was suggested to act as a chemo- and/or mechanoreceptor for CSF-cNs and to be capable of modulating CSF-cNs excitability. Moreover, CSF-cNs express several ionotropic receptors (nicotinic, AMPA/kainate, glycine and GABA) and are inserted in a functional network that need to be characterized. Finally, more recently, spinal CSF-cNs were shown in lower vertebrate (zebrafish and lamprey) to detect variation in CSF flow and composition as well as in the extent of spinal cord torsion (mechanical stimuli) and to act as neuromodulator for motor activity.

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On the other hand, CSF-cNs are found in a region around the spinal CC referred as the ‘spinal cord stem cell niche’ (spinal niche) where dynamic and plastic processes in relation to glio- and neurogenesis involved in spinal cord reparatory processes were reported. Interestingly, CSF-cNs exhibit a peculiar maturation apparently halted in an intermediate rather low maturity status (low NeuN expression along with maintained expression of immaturity markers) even in 1-year-old animals. Thus, CSF-cN intermediate maturity, their strategical localization at the interface between CSF and parenchyma added to the fact that they are capable of detecting circulating signals point to a role in the organization and regulation of the spinal niche. Based on the recent studies, spinal CSF-cNs are suggested to represent a novel population of multimodal sensory neurons in the CNS capable of integrating messages from the extracellular environment and of modulating CNS activity through neuronal networks that need to be identified and characterized. CSF-cNs might also interact with cells within the spinal niche and modulate their activity or reactivity especially in the context of inflammatory or reparatory processes following spinal cord injuries (SCI). To characterize the role of CSF-cNs, our team develops a broad range of complementary techniques from electrophysiological recordings combined with calcium imaging to immunohistochemical, biochemical and molecular approaches.

three major axes

Our research project largely benefits from the PKD2L1-Cre (Cre-Lox system) transgenic mouse model that allows the selective visualization (EGFP, tdTomato), targeting (AAV and rabies viral infection), monitoring (GCaMP models) and manipulation (Optogenetics, DREADDs, genetic deletion) of spinal CSF-cNs. Using complementary experimental models from acute spinal cord slices to in vivo model as well as hemisection and ‘en bloc’ spinal cord preparation, our research activities develop around three major axes:

What are the physiological properties of spinal CSF-cNs along the central canal? CSF-cNs are present along the whole axis of the CC but little is known about their properties (phenotype, activity, modulation) and whether these differ according to the spinal cord segment considered. We conduct electrophysiological studies combined to molecular profiling and immunohistofluorescence to address this question. In particular, PKD2L1, which is highly expressed and has important functional consequences in CSF-cN activity, is only active at the level of the single channel. We are therefore analyzing what are the molecular mechanisms that might underlie its peculiar activity, under which circumstances it can be modulated and what will be the consequences on CSF-cNs physiology.

Characterizing the neuronal network(s) spinal CSF-cNs are inserted in to understand their physiological role in the spinal cord Spinal CSF-cNs appear inserted in a functional neuronal network but neither the territories they innervate, nor their pre-synaptic partners have been identified. Using transgenic mouse models and state-of-the-art virus-based tracing approaches combined to electrophysiological recordings and molecular profiling approaches, we are characterizing CSF-cN circuitry along the CC axis both at a morphological and functional level. This identification of the network CSF-cNs are part of is the prerequisite to better understand their role in the spinal cord activity and will open new lines of research to assess their role as sensory neuron intrinsic to the CNS.

Do CSF-cNs interact with cells within the spinal cord niche and do the participate to reparatory processes following SCI? The region around the CC is known for its regenerative properties and was shown to participate in spinal cord reparatory processes. On the other hand, CSF-cNs are in a strategic position between CSF and parenchyma and are suggested to play a role as sensory neurons. One can therefore ask whether this unique neuronal population interact with cells within the spinal niche, whether they are active players in its organization and whether following spinal cord injury or inflammation they take part to reparatory processes? To address these question, we develop genetic deletion mouse models of CSF-cNs as well as SCI models to determine such a role.

Collaborators

Dr Zampieri: cartography of spinal CSF-cN network using targeted viral infection approaches and behavioral approach to test in vivo the role of CSF-cNs.

Dr Hugnot: CSF-cN molecular profiling and interaction with cells of the spinal cord stem cell niche.

Figure-1_CSF-cN_PKD-EGFP_Sag_SpiCCI Figure-2_CSF-cN_PKD-MAP2_SpiCCI Figure-3_CSF-cN_EGFP-aTub_SpiCCI Figure-4_CSF-cN_EGFP-MAP2_SpiCCI Figure-5_CSF-cN_EGFP-NF_SpiCCI
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