Frédéric Brocard’s research while affiliated with Aix-Marseille University and other places

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Publications (66)


Modified Ashworth scale for spasticity assessment [43]. 453 454 Neurological Evaluations (GCS, ASIA)
Calpain-mediated sodium channel fragments in the monocentric SpasT-SCI-T trial: Biomarkers for central nervous system injuries and spasticity prediction
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February 2025

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Frédéric Brocard

Spinal cord injury and traumatic brain injury are major causes of long-term disability and are often complicated by spasticity, a motor disorder characterized by increased muscle tone and exaggerated reflexes that significantly impair quality of life. Current diagnostic methods lack the sensitivity needed to accurately predict the severity of injury or the onset and progression of spasticity. Trauma-induced calcium dysregulation activates calpains, a family of proteases that cleave sodium channels, disrupting their inactivation and increasing persistent sodium currents. This cascade drives the overexcitability of motoneurons, contributing to the development of spasticity. Consequently, sodium channel fragments have emerged as promising biomarkers that link injury mechanisms to clinical outcomes. The present SpasT-SCI-T clinical trial protocol aims to evaluate sodium channel fragments as blood biomarkers for assessing the severity of spinal cord and traumatic brain injuries, as well as their potential to predict clinical outcomes, including the development of spasticity. This prospective, multicenter, case-control and cohort study involves 40 participants: 20 individuals with spinal cord injury, 10 individuals with traumatic brain injury, and 10 healthy controls. Blood samples are collected within six hours of injury and at follow-up points over six months. Clinical outcomes, including spasticity (assessed using the Modified Ashworth Scale), neurological recovery (measured by the American Spinal Injury Association Impairment Scale and Glasgow Coma Scale), and quality of life (evaluated using the Short Form-36 Health Survey), are analyzed in correlation with biomarker levels. We anticipate that calpain-mediated sodium channel fragments will transform the management of central nervous system injuries by enabling early diagnosis, improving prognostic accuracy, and guiding personalized therapeutic strategies. The clinical trial is registered on ClinicalTrials.gov (NCT06532760, January 10, 2024), with Assistance Publique–Hôpitaux de Marseille as the sponsor.

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Functional contribution of astrocytic Kir4.1 channels to spasticity after spinal cord injury

October 2024

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52 Reads

Spasticity, a prevalent motor issue characterized by network hyperexcitability, causes pain and discomfort, with existing treatments offering limited relief. While past research has focused on neuronal factors, the role of astrocytes in spasticity has been overlooked. This study explores the potential of restoring astrocytic potassium (K ⁺ ) uptake to reduce spasticity following SCI. Astrocytes buffer extracellular K ⁺ via Kir4.1 channels, preventing neuronal hyperexcitability. Following spinal cord injury (SCI), Kir4.1 levels decrease at the injury site, though the consequences and mechanisms of this reduction within the motor output area have not been investigated. Utilizing advanced techniques, we demonstrate that lumbar astrocytes in a juvenile thoracic SCI mouse model switch to reactive phenotype, displaying morpho-functional and pro-inflammatory changes. These astrocytes also experience NBCe1-mediated intracellular acidosis, leading to Kir4.1 dysfunction and impaired K ⁺ uptake. Enhancing Kir4.1 function reduces spasticity in SCI mice, revealing new therapeutic targets for neurological diseases associated with neuronal hyperexcitability. Highlights Lumbar astrocytes adopt a reactive phenotype following a thoracic SCI NBCe1-mediated acidosis in astrocytes disrupts Kir4.1 function post-SCI. Impaired K+ uptake leads to motoneuron hyperexcitability post-SCI. Enhanced astroglial Kir4.1 function reduces spastic-like symptoms in SCI mice.


Effect of size on expression of bistability in mouse spinal motoneurons

February 2024

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88 Reads

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5 Citations

Journal of Neurophysiology

Bistability in spinal motoneurons supports tonic spike activity in the absence of excitatory drive. Earlier work in adult preparations suggested that smaller motoneurons innervating slow antigravity muscle fibers are more likely to generate bistability for postural maintenance. However, whether large motoneurons innervating fast-fatigable muscle fibers display bistability is still controversial. To address this, we examined the relationship between soma size and bistability in lumbar (L4-L5) ventrolateral a-motoneurons of ChAT-GFP and Hb9-GFP mice during the first four weeks of life. We found that as neuron size increases, the prevalence of bistability rises. Smaller a-motoneurons lack bistability, while larger fast a-motoneurons (MMP-9 ⁺ /Hb9 ⁺ ) with a soma area ≥ 400µm ² exhibit significantly higher bistability. Ionic currents associated with bistability, including the persistent Nav1.6 current, thermosensitive Trpm5 Ca ²⁺ -activated Na ⁺ current and the slowly inactivating Kv1.2 current, also scale with cell size. Serotonin evokes full bistability in large motoneurons with partial bistable properties, but not in small motoneurons. Our study provides important insights into the neural mechanisms underlying bistability and how motoneuron size correlates with bistability in mice.



A size principle for bistability in mouse spinal motoneurons

September 2023

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92 Reads

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1 Citation

Bistability in spinal motoneurons supports tonic spike activity in the absence of excitatory drive. Earlier work in adult preparations suggested that smaller motoneurons innervating slow antigravity muscle fibers are more likely to generate bistability for postural maintenance. However, whether large motoneurons innervating fast-fatigable muscle fibers display bistability related to postural tone is still controversial. To address this, we examined the relationship between soma size and bistability in lumbar ventrolateral α-motoneurons of ChAT-GFP and Hb9-GFP mice across different developmental stages: neonatal (P2-P7), young (P7-P14) and mature (P21-P25). We found that as neuron size increases, the prevalence of bistability rises. Smaller α-motoneurons lack bistability, while larger fast α-motoneurons (MMP-9 ⁺ /Hb9 ⁺ ) with a soma area ≥ 400µm ² exhibit significantly higher bistability. Ionic currents associated with bistability, including the persistent Nav1.6 current, thermosensitive Trpm5 Ca ²⁺ -activated Na ⁺ current and the slowly inactivating Kv1.2 current, also scale with cell size. Serotonin evokes full bistability in large motoneurons with partial bistable properties, but not in small motoneurons. Our study provides important insights into the neural mechanisms underlying bistability and how motoneuron size dictates this process. New and Noteworthy Bistability is not a common feature of all mouse spinal motoneurons. It is absent in small, slow motoneurons but present in most large, fast motoneurons. This difference results from differential expression of ionic currents that enable bistability, which are highly expressed in large motoneurons but small or absent in small motoneurons. These results support a possible role for fast motoneurons in maintenance of tonic posture in addition to their known roles in fast movements.


Figure 4. Nav1.6 channels encode I NaP in motoneurons
Persistent Nav1.1 and Nav1.6 currents drive spinal locomotor functions through nonlinear dynamics

September 2023

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129 Reads

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6 Citations

Cell Reports

Persistent sodium current (INaP) in the spinal locomotor network promotes two distinct nonlinear firing patterns: a self-sustained spiking triggered by a brief excitation in bistable motoneurons and bursting oscillations in interneurons of the central pattern generator (CPG). Here, we identify the NaV channels responsible for INaP and their role in motor behaviors. We report the axonal Nav1.6 as the main molecular player for INaP in lumbar motoneurons. The inhibition of Nav1.6, but not of Nav1.1, in motoneurons impairs INaP, bistability, postural tone, and locomotor performance. In interneurons of the rhythmogenic CPG region, both Nav1.6 and Nav1.1 equally mediate INaP. Inhibition of both channels is required to abolish oscillatory bursting activities and the locomotor rhythm. Overall, Nav1.6 plays a significant role both in posture and locomotion by governing INaP-dependent bistability in motoneurons and working in tandem with Nav1.1 to provide INaP-dependent rhythmogenic properties of the CPG.


Persistent Nav1.6 current drives spinal locomotor functions through nonlinear dynamics

April 2023

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61 Reads

Persistent sodium current (INaP) in the spinal locomotor network promotes two distinct nonlinear firing patterns: a self-sustained spiking triggered by a brief excitation in bistable motoneurons and bursting oscillations in interneurons of the central pattern generator (CPG). Here, we identified the NaV channels responsible for INaP and their role in motor behaviors. We report the axonal Nav1.6 as the main molecular player for INaP in lumbar motoneurons. The motoneuronal inhibition of Nav1.6, but not of Nav1.1, impairs INaP, bistability, postural tone and locomotor performance. In interneurons of the CPG region, Nav1.6 with Nav1.1 equally mediate INaP and the inhibition of both channels is required to abolish oscillatory bursting activities and the locomotor rhythm. Overall, Nav1.6 plays a significant role both in posture and locomotion by governing INaP-dependent bistability in motoneurons and working in tandem with Nav1.1 to provide INaP-dependent rhythmogenic properties of the CPG.


Standard protocol items recommended for interventional trials (SPIRIT)
Schedule of enrolment, intervention and assessments. The study schedule will be the same in step 1 and in step 2 and it includes the following visits and clinical examinations. Patients with chronic SCI will undergo a screening period (v1) with a maximum duration of 2 weeks and then they will be randomized to one of the two groups (v2): riluzole twice daily or placebo twice daily for a 2 weeks period. Three follow up visits are planned: at day3 after the beginning of treatment for PK analysis (v3), and at the end of the 2 weeks treatment period (v4) and one week after withdrawal of study medication (v5).
CONSORT diagram
Riluzole for treating spasticity in patients with chronic traumatic spinal cord injury: Study protocol in the phase ib/iib adaptive multicenter randomized controlled RILUSCI trial

January 2023

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79 Reads

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5 Citations

Background Satisfactory treatment is often lacking for spasticity, a highly prevalent motor disorder in patients with spinal cord injury (SCI). Low concentrations of riluzole potently reduce the persistent sodium current, the post-SCI increase in which contributes to spasticity. The repurposing of this drug may therefore constitute a useful potential therapeutic option for relieving SCI patients suffering from chronic traumatic spasticity. Objective RILUSCI is a phase 1b–2b trial designed to assess whether riluzole is a safe and biologically effective means of managing spasticity in adult patients with traumatic chronic SCI. Methods In this multicenter double-blind trial, adults (aged 18–65 years) suffering from spasticity after SCI (target enrollment: 90 participants) will be randomly assigned to be given either a placebo or a recommended daily oral dose of riluzole for two weeks. The latter dose will be previously determined in phase 1b of the study by performing double-blind dose-finding tests using a Bayesian continuous reassessment method. The primary endpoint of the trial will be an improvement in the Modified Ashworth Score (MAS) or the Numerical Rating Score (NRS) quantifying spasticity. The secondary outcomes will be based on the safety and pharmacokinetics of riluzole as well as its impact on muscle spasms, pain, bladder dysfunction and quality of life. Analyses will be performed before, during and after the treatment and the placebo-controlled period. Conclusion To the best of our knowledge, this clinical trial will be the first to document the safety and efficacy of riluzole as a means of reducing spasticity in patients with chronic SCI. Trial registration The clinical trial, which is already in progress, was registered on the ClinicalTrials.gov website on August 9, 2016 under the registration number NCT02859792. Trial sponsor Assistance Publique–Hôpitaux de Marseille.


Astrocytic Kir4.1 channels regulate locomotion by orchestrating neuronal rhythmicity in the spinal network

January 2023

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214 Reads

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7 Citations

Glia

Neuronal rhythmogenesis in the spinal cord is correlated with variations in extracellular K + levels ([K + ] e). Astrocytes play important role in [K + ] e homeostasis and compute neuro-nal information. Yet it is unclear how neuronal oscillations are regulated by astrocytic K + homeostasis. Here we identify the astrocytic inward-rectifying K + channel Kir4.1 (a.k.a. Kcnj10) as a key molecular player for neuronal rhythmicity in the spinal central pattern generator (CPG). By combining two-photon calcium imaging with electrophysiology, immunohistochemistry and genetic tools, we report that astrocytes display Ca 2+ tran-sients before and during oscillations of neighboring neurons. Inhibition of astrocytic Ca 2+-transients with BAPTA decreases the barium-sensitive Kir4.1 current responsible of K + clearance. Finally, we show in mice that Kir4.1 knockdown in astrocytes progressively prevents neuronal oscillations and alters the locomotor pattern resulting in lower motor performances in challenging tasks. These data identify astroglial Kir4.1 channels as key regulators of neuronal rhythmogenesis in the CPG driving locomotion.



Citations (48)


... All experiments were designed to gather data within a stable period (i.e., at least 2 min after establishing whole-cell access). Because (i) the cell size influences neuronal excitability [39] and because (ii) we recorded non-identified neurons, we only compared the electrophysiological parameters of cells with the same size in both conditions to avoid any recording bias related to their morphological features. ...

Reference:

Dorsoventral photobiomodulation therapy safely reduces inflammation and sensorimotor deficits in a mouse model of multiple sclerosis
Effect of size on expression of bistability in mouse spinal motoneurons

Journal of Neurophysiology

... Недавно было показано, что у животных с нарушением целостности спинного мозга инактивация гена кальпаина-1, хорошо известной кальций-зависимой цистеиновой протеазы, предотвращает снижение содержания КСС2 и последующую спастичность мышц [21]. Результаты этого исследования позволяют сделать вывод, что именно кальпаин-зависимый протеолиз является основной причиной снижения содержания KCC2 в поясничном отделе спинного мозга. ...

Knockdown of calpain1 in lumbar motoneurons reduces spasticity after spinal cord injury in adult rats

Molecular Therapy

... These changes help to facilitate motoneuron recruitment and repetitive firing that are needed for sustained motor behaviours such as posture and weight bearing locomotion (Carrascal et al., 2005;Jean-Xavier et al., 2018). One such marked change is the decrease in threshold and increase in amplitude of persistent inward currents (PICs) in the first 3 weeks after birth (Harris-Warrick et al., 2023;Quinlan et al., 2011;Revill et al., 2019;Sharples & Miles, 2021). PICs are mediated by voltage-activated sodium (Na V ) and calcium (Ca V ) channels (Li et al., 2004) and by a calcium-activated sodium conductance (I CaN ) (Bos et al., 2021). ...

A size principle for bistability in mouse spinal motoneurons

... While differences in biophysical properties could also underlie the differential roles of Na V 1.1 and Na V 1.6 to proprioceptive transmission, these channels share many functional similarities. They both rapidly activate and inactivate; can generate peak, persistent, and resurgent currents; and have similar recovery from inactivation kinetics (7,27,28,(45)(46)(47). Prior work in neurons of the central nervous system is consistent with our hypothesis that localization dictates Na V contributions to proprioceptor function. ...

Persistent Nav1.1 and Nav1.6 currents drive spinal locomotor functions through nonlinear dynamics

Cell Reports

... Although the initial assessment fell short of meeting the predetermined criteria for efficacy, further analyses revealed significant functional recovery gains across all subgroups of cervical SCI patients who were treated with riluzole. A phase 2 adaptive, multicenter, placebocontrolled, randomized, double-blind trial, to assess whether riluzole is a safe and biologically effective means of managing spasticity in adult patients with traumatic chronic SCI is already in progress [88]. ...

Riluzole for treating spasticity in patients with chronic traumatic spinal cord injury: Study protocol in the phase ib/iib adaptive multicenter randomized controlled RILUSCI trial

... First, a strong effect of PBMT was observed in the ventral motor networks, where photons struggled to penetrate, compared with the dorsal regions that are more directly exposed to light after spinal glass window implantation. Second, since astrocytes are interconnected by gap junctions into widespread glial networks [47] and modulate neuronal excitability in both the dorsal [44] and ventral [48] spinal cord, dorsal illumination could thus indirectly impact ventral neuronal excitability. However, the effect of PBMT on glial reactivity, both astrocytic and microglial, was also larger in the ventral region than in the dorsal one. ...

Astrocytic Kir4.1 channels regulate locomotion by orchestrating neuronal rhythmicity in the spinal network

Glia

... A consequence of VRK1 pathogenic variants in motor neurons is the impairment of their action potential that are smaller in amplitude, larger in duration, and show a more depolarized threshold [54]. These defects in motor neurons, clinically manifested by neurological symptoms, eventually, are the results of the pathogenic roles of the different variant combinations, among them, the effects that VRK1 variants have on the altered assembly and stability of Cajal bodies, which can play a major pathogenic role [3,7,9,23,37,39]. ...

Altered action potential waveform and shorter axonal initial segment in hiPSC-derived motor neurons with mutations in VRK1

Neurobiology of Disease

... In the motoneurons, TRPM5 is activated by Ca 2+ released via the ryanodine receptor. Silencing TRPM5 in motoneurons leads to hind limb paresis and causes difficulties in executing high-demand locomotor tasks [6]. ...

Trpm5 channels encode bistability of spinal motoneurons and ensure motor control of hindlimbs in mice

... Both INaP and IKM have slow inactivation kinetics and are activated near the resting membrane potential (~-65 mV). 42,43 Previous studies demonstrated their functional interaction as inhibition of IKM results in the INaPmediated burst discharges [44][45][46][47] or a plateau depolarization when extracellular Ca 2+ is reduced. 48,49 The latter might be particularly relevant to the slow ramp membrane depolarization during SD which involves a rapid drop of extracellular Ca 2+ . 1 On the other hand, since Scn8a and Kcnq2 are expressed differentially across cell types and brain regions, mutations in these genes will impair the INaP and IKM balance differently across the brain regions. ...

The M-current works in tandem with the persistent sodium current to set the speed of locomotion

... An intriguing clinical study postulated that loss of RDD in SCI originates from impaired presynaptic mechanisms as motor-evoked potential responses, purportedly reflecting postsynaptic inhibition, were comparable among subjects with and without RDD [49]. However, the pure presynaptic origin theory of spasticity is not unanimously embraced, as other studies implicate glycinergic inhibition of lumbar motor neurons in the mechanisms underlying spasticity after SCI, which also influences RDD [34,50]. Further mechanistic associations were provided by Boulenguez et al. [34] who reported that blocking KCC2 in healthy rats diminished RDD while RDD was also reduced in both KCC2-deficient mice and intact rats following intrathecal delivery of brain-derived neurotrophic factor (BDNF) injection to downregulate KCC2. ...

Alteration of glycinergic receptor expression in lumbar spinal motoneurons is involved in the mechanisms underlying spasticity after spinal cord injury

Journal of Chemical Neuroanatomy