Ventral Horn Neurons Research Articles

A reassessment of spinal cord pathology in severe infantile spinal muscular atrophy: Reassessment of spinal cord pathology.

Spinal muscular atrophy (SMA) is a life-limiting paediatric motor neuron disease characterised by lower motor neuron loss, skeletal muscle atrophy and respiratory failure, if untreated. Revolutionary treatments now extend patient survival. However, a limited understanding of the foundational neuropathology challenges the evaluation of therapeutic success. As opportunities to study treatment-naïve tissue decrease, we have characterised spinal cord pathology in severe infantile SMA using gold-standard techniques, providing a baseline to measure treatment success and therapeutic limitations. Detailed histological analysis, stereology and transmission electron microscopy were applied to post-mortem spinal cord from severe infantile SMA patients to estimate neuron number at the end of life; characterise the morphology of ventral horn, lateral horn and Clarke's column neuron populations; assess cross-sectional spinal cord area; and observe myelinated white matter tracts in the clinically relevant thoracic spinal cord. Ventral horn neuron loss was substantial in all patients, even the youngest cases. The remaining ventral horn neurons were small with abnormal, occasionally chromatolytic morphology, indicating cellular damage. In addition to ventral horn pathology, Clarke's column sensory-associated neurons displayed morphological features of cellular injury, in contrast to the preserved sympathetic lateral horn neurons. Cellular changes were associated with aberrant development of grey and white matter structures that affected the overall dimensions of the spinal cord. We provide robust quantification of the neuronal deficit found at the end of life in SMA spinal cord. We question long-accepted dogmas of SMA pathogenesis and shed new light on SMA neuropathology out with the ventral horn, which must be considered in future therapeutic design.

Cilostazol Ameliorates Motor Dysfunction and Schwann Cell Impairment in Streptozotocin-Induced Diabetic Rats.

This study investigated the effects of cilostazol on motor dysfunction, spinal motor neuron abnormalities, and schwannopathy in rats with diabetes. Diabetes mellitus (DM) was induced in rats via femoral intravenous streptozotocin (STZ) injection (60 mg/kg). After successful DM induction, cilostazol was administered on day 15 via oral gavage (100 mg/kg/day) for 6 weeks until sacrifice. Behavioral assays, including motor function, were performed weekly. The sciatic nerve, L5 spinal cord, and spinal ventral root were collected to evaluate the expression of the glial fibrillary acidic protein (GFAP), myelin protein zero (P0), and choline acetyltransferase (ChAT) by immunofluorescence and Western blotting. DM rats displayed decreased running speeds, running distances, and toe spread but increased foot pressure. In addition, loss of non-myelinating Schwann cells and myelin sheaths was observed in the sciatic nerve and L5 spinal ventral root. Reduced numbers of motor neurons were also found in the L5 spinal ventral horn. Cilostazol administration significantly potentiated running speed and distance; increased hind paw toe spread; and decreased foot pressure. In the sciatic nerve and L5 spinal ventral root, cilostazol treatment significantly improved non-myelinated Schwann cells and increased myelin mass. ChAT expression in motor neurons in the spinal ventral horn was improved, but not significantly. Cilostazol administration may protect sensorimotor function in diabetic rats.

Transsynaptic degeneration of ventral horn motor neurons exists but plays a minor role in lower motor system dysfunction in acute ischemic rats.

As a leading cause of mortality and long-term disability, acute ischemic stroke can produce far-reaching pathophysiological consequences. Accumulating evidence has demonstrated abnormalities in the lower motor system following stroke, while the existence of Transsynaptic degeneration of contralateral spinal cord ventral horn (VH) neurons is still debated. Using a rat model of acute ischemic stroke, we analyzed spinal cord VH neuron counts contralaterally and ipsilaterally after stroke with immunofluorescence staining. Furthermore, we estimated the overall lower motor unit abnormalities after stroke by simultaneously measuring the modified neurological severity score (mNSS), compound muscle action potential (CMAP) amplitude, repetitive nerve stimulation (RNS), spinal cord VH neuron counts, and the corresponding muscle fiber morphology. The activation status of microglia and extracellular signal-regulated kinase 1/2 (ERK 1/2) in the spinal cord VH was also assessed. At 7 days after stroke, the contralateral CMAP amplitudes declined to a nadir indicating lower motor function damage, and significant muscle disuse atrophy was observed on the same side; meanwhile, the VH neurons remained intact. At 14 days after focal stroke, lower motor function recovered with alleviated muscle disuse atrophy, while transsynaptic degeneration occurred on the contralateral side with elevated activation of ERK 1/2, along with the occurrence of neurogenic muscle atrophy. No apparent decrement of CMAP amplitude was observed with RNS during the whole experimental process. This study offered an overview of changes in the lower motor system in experimental ischemic rats. We demonstrated that transsynaptic degeneration of contralateral VH neurons occurred when lower motor function significantly recovered, which indicated the minor role of transsynaptic degeneration in lower motor dysfunction during the acute and subacute phases of focal ischemic stroke.

Effect of Non-invasive Spinal Stimulation on Self-sustained Firing Motoneuron Model: In-Silico Study Using Human Body Model.

Transcutaneous spinal electrical stimulation (tSCS) is a non-invasive neuromodulation approach using a low intensity direct current. Recent developments in the technique have opened the possibility that tSCS can help restore motor function after spinal cord injury (SCI). However, the exact mechanism of action tSCS has on the spinal circuits is still unknown. Due to the complexity of experimental synthesis in a human model to delineate the mechanisms, models that link the stimulation paradigm and circuit behaviors are advantageous. Thus, this study aims to simulate the underlying changes in motor circuit firing rates in response to external stimuli induced by tSCS. Serial stimulations combining a high-fidelity finite element model with the human torso and spinal cord with a lumped motor neuron model is constructed. The parameters for both components of the model were derived from previous studies. We focused our analysis on a lumped motor neuron model that describes sustained firing behavior of the motor neuron driven primarily by persistent inward current (PIC), a signature behavior of the motor neuron after SCI. Modulation of the PIC behaviors was achieved by stimulating voltage-dependent calcium and sodium channels in the dendrite using a tSCS-induced electric field (E-field) expressed at different a spatial locations of the motor neuron in the gray matter. The PIC behaviors of spinal motor neurons in the left ventral horn were suppressed, while for the most part invariant in the right ventral horn. These initial simulations will provide a steppingstone for future examinations that incorporate additional neuronal models of inhibitory and excitatory interneurons to access the circuit-level effect of spinal stimulation.

Therapy of spinal cord injury by zinc pyrogallol modified nanozyme via anti-inflammatory strategies

ROS-mediated treatment is a potential strategy that can reduce the risk of spinal cord injury (SCI)-related obvious adverse effects, but at the cost of therapeutic efficacy. Considering the significant reactive oxygen species (ROS) scavenging ability and biocompatibility of natural enzyme such as superoxide dismutase (SOD) and catalase (CAT), zinc pyrogallol nanozyme (PA-Zn) was developed to treat the injured spinal cord, in which zinc as metal center was coupled to pyrogallol through the Zn-O-C bridge to form a zinc four-coordination structure that can mimic enzyme-like behaviors of SOD and CAT for SCI treatment. In vitro tests demonstrated that PA-Zn could efficiently inhibit the expression of ROS and M1-related markers (IL-1β), and upregulate the expression of M2-related markers (Arg-1). Furthermore, PA-Zn could significantly promote ventral horn neurons survival and locomotor functional recovery in vivo, while inhibit injured spinal cord damage, and decrease the infiltration of macrophages in the lesion. PA-Zn had a better therapeutic effect on SCI animals at extremely low dosage (0.1 mg kg−1) and shorter time (2 weeks). The working dosage of PA-Zn was even 100-fold lower than some previous reported nanomedicines for SCI treatment. The PA-Zn with muti-enzymatic activities and biocompatibility showed reliable and efficient treatment efficiency for not only SCI but also other ROS-mediated diseases.

Psychostimulant Drugs Activate Cell-type Specific and Topographic cFos Expression in the Lumbar Spinal Cord

Psychostimulant drugs, such as cocaine, d-amphetamine and methylphenidate, alter a wide range of behaviors including locomotor activity and somatosensory perception. These altered behaviors are accompanied by the activation of specific neuronal populations within reward-, emotion- and locomotion-related circuits. However, whether such regulation occurs at the level of the spinal cord, a key node for neural circuits integrating and coordinating sensory and motor functions has never been addressed. By evaluating the temporal and spatial expression pattern of the phosphorylated form of the immediate early gene cFos at Ser32 (pS32-cFos), used as a proxy of neuronal activation, we demonstrate that, in adult male mice, d-amphetamine increases pS32-cFos expression in both inhibitory and excitatory neurons in dorsal and ventral horns at the lumbar spinal cord level. Interestingly, a fraction of neurons activated by a first exposure to d-amphetamine can be re-activated following d-amphetamine re-exposure. Similar expression patterns were observed in response to cocaine and methylphenidate, but not following morphine and dozilcipine administration. Finally, the blockade of dopamine reuptake was sufficient to recapitulate the increase in pS32-cFos expression induced by psychostimulant drugs. Our work provides evidence that cFos expression can be activated in lumbar spinal cord in response to acute psychostimulants administration.

Spatiotemporal evolution of pyroptosis and canonical inflammasome pathway in hSOD1G93A ALS mouse model

BackgroundEvidences indicate that inflammasome compounds participate in amyotrophic lateral sclerosis (ALS), a fatal progressive motoneuron degenerative disease. Researchers have observed the expressions of nucleotide oligomerization domain (NOD)-like receptor protein 3 (NLRP3) related inflammasome components in specific regions of the central nervous system in different ALS models, but the cellular spatiotemporal evolution of this canonical inflammasome pathway and pyroptosis during ALS progression are unclear.MethodsThe spinal cords of hSOD1G93A mice (ALS mice) and age-matched littermates (CON mice) were dissected at pre-symptomatic stage (60 d), early- symptomatic stage (95 d), symptomatic stage (108 d) and late-symptomatic stage (122 d) of the disease. By using Nissl staining, double immunofluorescence labelling, qRT-PCR or western blot, we detected morphology change and the expression, cellular location of GSDMD, NLRP3, caspase-1 and IL-1β in the ventral horn of lumbar spinal cords over the course of disease.ResultsNeural morphology changes and GSDMD+/NeuN+ double positive cells were observed in ventral horn from ALS mice even at 60 d of age, even though there were no changes of GSDMD mRNA and protein expressions at this stage compared with CON mice. With disease progression, compared with age-matched CON mice, increased expressions of GSDMD, NLRP3, activated caspase-1 and IL-1β were detected. Double immunofluorescence labeling revealed that NLRP3, caspase-1, IL-1β positive signals mainly localized in ventral horn neurons at pre- and early-symptomatic stages. From symptomatic stage to late-symptomatic stage, robust positive signals were co-expressed in reactive astrocytes and microglia.ConclusionsEarly activation of the canonical NLRP3 inflammasome induced pyroptosis in ventral horn neurons, which may participate in motor neuron degeneration and initiate neuroinflammatory processes during ALS progression.

Activation of M3-AChR and IP3/Ca2+/PKC signaling pathways by pilocarpine increases glycine-induced currents in ventral horn neurons of the spinal cord

Our study aimed to determine the effects of pilocarpine and the mechanisms involving muscarinic acetylcholine receptors (mAChRs) on glycine receptors (GlyRs) in neurons of the spinal cord ventral horn. An enzymatic digestion combined with acute mechanical separation was applied to isolate neurons from the spinal cord ventral horn. Patch-clamp recording was then used to investigate the outcomes of pilocarpine. Our results indicate that pilocarpine increased the glycine currents in a concentration-dependent manner, which was blocked by the M3-AChR selective antagonists 4-DAMP and J104129. Pilocarpine also enhanced the glycine currents in nominally Ca2+-free extracellular solution. Conversely, the enhancement of glycine currents by pilocarpine disappeared when intracellular Ca2+ was chelated by BAPTA. Heparin and Xe-C, which are IP3 receptor antagonists, also totally abolished the pilocarpine effect. Furthermore, Bis-IV, a PKC inhibitor, eliminated the pilocarpine effect. Additionally, PMA, a PKC activator, mimicked the pilocarpine effect. These results indicate that pilocarpine may increase the glycine currents by activating the M3-AChRs and IP3/Ca2+/PKC pathways.

Induction of Survival of Motor Neuron (SMN) Protein Deficiency in Spinal Astrocytes by Small Interfering RNA as an In Vitro Model of Spinal Muscular Atrophy.

Spinal muscular atrophy (SMA) is a motor neuron disorder leading to progressive loss of ventral horn neurons resulting in muscle wasting. Here we investigate the contribution of spinal astrocytes to the pathogenesis of late-onset SMA forms using a mouse model. Furthermore, we generated SMA-like astrocytes using survival of motor neuron (SMN) siRNA transfection techniques. In the SMA mouse model, the activation of spinal astrocytes and the reduction of the inward rectifier potassium channel Kir4.1 and excitatory amino acid transporter 1 (EAAT1) were observed at postnatal day (P) 28, preceding the loss of spinal motor neurons appearing earliest at P42. Using SMA-like astrocytes, we could mimic the modulation of spinal astrocytes of the mouse model in a dish and perform electrophysiological assessments and functional assays. In SMA-like astrocytes, glutamate uptake was diminished due to a reduction in EAAT1. Furthermore, patch-clamp measurements revealed reduced potassium uptake into astrocytes with membrane depolarization. Additionally, exposure of healthy spinal motor neurons to a conditioned medium of SMA-like astrocytes resulted in increased firing frequency. These data demonstrate spinal astrocytes’ crucial role in the late-onset SMA forms’ pathogenesis.

Coherence analysis of the calcium activity of putative astrocytic and neuronal cells on the L5 ventral horn and neural output in activated lumbar CPG networks

Astrocytes are thought to play a crucial role in providing structure to the spinal cord and maintaining efficient synaptic function and metabolism because their fine processes envelop the synapses of neurons and form many neuronal networks within the central nervous system (CNS). To investigate whether putative astrocytes and putative neurons distributed on the ventral horn play a role in the modulation of lumbar locomotor central pattern generator (CPG) networks, we used extracellular recording and optical imaging techniques and recorded the neural output from the left L5 ventral root and the calcium activity of putative astrocytes and neurons in the L5 ventral horn at the same time when activating an isolated L1–L5 spinal cord preparation from rats aged 0–2 days. Optical measurements detected cells that showed a fluorescence intensity change under all experimental conditions, namely, (1) 5-HT + NMDA, (2) TTX, and (3) TTX + Low K+. These cells were semiautomatically identified using an in-house MATLAB-based program, as putative astrocytes and neurons according to the cell classification, i.e., increased or decreased fluorescence intensity change (ΔF/F0), and subjective judgment based on their soma size. Coherence and its phase were calculated according to the calcium activity of the putative astrocytes and putative neurons, and neural output was calculated during fictive locomotion with in-house MATLAB-based programs. We found that the number of putative astrocytes activated by applying low K+ tends not to differ from that activated by applying the protease-activated receptor 1 (PAR1) selective agonist TFLLR-NH2 (TFLLR). Moreover, the calcium activity of several putative astrocytes and neurons synchronized with locomotor-like activity at a frequency range below 0.5 Hz and the time lag between peaks of cellular calcium activity and locomotor-like activity ranged from −1000 to + 1000 ms. These findings presumably indicates that these putative astrocytes and neurons in the left L5 ventral horn require −1000 to + 1000 ms to communicate with lumbar CPG networks and maintain efficient synaptic function and metabolism in activated lumbar CPG networks. This finding suggests the possibility that putative astrocytic and neuronal cells in the L5 ventral horn contribute to generating the rhythms and patterns of locomotor-like activity by activated CPG networks in the first to fifth lumbar spinal cord.

Fetal and Perinatal Expression Profiles of Proinflammatory Cytokines in the Neuroplacodes of Rats with Myelomeningoceles: A Contribution to the Understanding of Secondary Spinal Cord Injury in Open Spinal Dysraphism.

The cellular and molecular mechanisms that presumably underlie the progressive functional decline of the myelomeningocele (MMC) placode are not well understood. We previously identified key players in post-traumatic spinal cord injury cascades in human MMC tissues obtained during postnatal repair. In this study, we conducted experiments to further investigate these mediators in the prenatal time course under standardized conditions in a retinoic acid-induced MMC rat model. A retinoic acid MMC model was established using time-dated Sprague-Dawley rats, which were gavage-fed with all-trans retinoic acid (RA; 60 mg/kg) dissolved in olive oil at E10. Control animals received olive oil only. Fetuses from both groups were obtained at E16, E18, and E22. The spinal cords (SCs) of both groups were formalin-fixed or snap-frozen. Tissues were screened by real-time reverse transcription polymerase chain reaction for the expression of cytokines and chemokines known to play a role in the lesion cascades of the central nervous system after trauma. MMC placodes exhibited inflammatory cells and glial activation in the later gestational stages. At the messenger RNA (mRNA) level, interleukin-1 beta, tumor necrosis factor alpha, and tumor necrosis factor receptor type 1 exhibited significant induction at E22. interleukin-1 beta receptor type 1 mRNA was induced significantly at E16 and E22. Double labeling experiments confirmed the co-staining of these cytokines and their receptors with ionized calcium-binding adapter molecule 1 (i.e., inflammatory cells), vimentin, and nestin in different anatomical SC areas and neuronal nuclear protein in ventral horn neurons. C-X-C motif chemokine 12 mRNA was elevated in control and MMC animals at E16 compared with E18 and E22. C-X3-C motif ligand 1 mRNA was lower in MMC tissues than in control tissues on E16. The presented findings contribute to the concept that pathophysiological mechanisms, such as cytokine induction in the neuroplacode, in addition to the "first hit," promote secondary spinal cord injury with functional loss in the late fetal time course. Further, these mediators should be taken into consideration in the development of new therapeutic approaches for open spinal dysraphism.

Schmallenberg virus infection suspected in a calf born to an imported heifer.

Schmallenberg virus infection suspected in a calf born to an imported heifer.

Orexin-A inhibits γ-aminobutyric acid current of neonatal rat spinal cord ventral horn neurons by activating OX1R, OX2R and Ca2+-independent PKC

To investigate the effect of orexin-A on the functionality of ionotropic γ-aminobutyric acid (GABA) receptors in spinal cord ventral horn neurons and its mechanisms. The spinal cord containing the lumbosacral enlargement was isolated from neonatal SD rats (7-12 days old) and sliced. The slices were digested with papain (in 0.18 g/30 mL artificial cerebrospinal fluid) for 40-60 min, and the ventral horn neurons were separated acutely using fire-polished Pasteur pipettes. After the cells adhered to the bottom of Petri dishes, patch-clamp experiments combined with pharmacological methods were performed to test the effects of orexin-A on GABA currents of the neurons treated with SB334867 (a selective OX1R antagonist), TCSOX229 (a selective OX2R antagonist), Bis-Ⅳ (a PKC inhibitor), PMA (a PKC agonist), Rp-cAMP (a PKA inhibitor), or BAPTA (Ca2+ chelator). The isolated neurons maintained good morphologies with diverse shapes of cell body and long protrusions. Treatment with orexin-A significantly inhibited the amplitude of GABA-induced current (P < 0.001, n=49) with an inhibition rate of (67.48±12.50)%. SB334867 and TCSOX229, when applied simultaneously, completely abolished the suppressive effect of orexin-A on the GABA currents (P=0.93, n=6), and their separate use partially relieved the suppressive effect of orexin-A (P=0.001, n=8; P=0.02, n=8). The application of Bis-Ⅳ also abolished the suppressive effect of orexin-A on GABA currents (P=0.31, n=5). PMA mimicked the effect of orexin-A in these neurons and significantly inhibited GABA currents with an inhibition rate of (60.79±10.94)%, and the application of orexin-A did not cause further suppression of GABA currents in PMA-treated neurons (P=0.15, n=6). Orexin-A was still capable of suppressing GABA currents in Rp-cAMP-treated neurons (P=0.001, n=5). The extracellular Ca2+-free solution (P=0.004, n=8) or the presence of BAPTA (P=0.04, n=7) did not significantly affect the inhibitory effect of orexin-A on GABA currents. Orexin-A inhibits GABA currents in the ventral horn neurons of rat spinal cord probably by activating OX1R, OX2R and Ca2+-independent PKC.

This Month in Anesthesiology

This Month in Anesthesiology

Inhibition of ERK1/2 phosphorylation attenuates spinal cord injury induced astrocyte activation and inflammation through negatively regulating aquaporin-4 in rats

The extracellular signal-regulated kinase (ERK) pathway has been reported to play a pivotal role in mediating spinal cord injury (SCI) progression. The present study aimed to investigate the effects of phosphorylated ERK1/2 (p-ERK1/2) inhibition on SCI-induced astrocyte activation and inflammation and its possible mechanism in rats. Here, female Sprague-Dawley rats were randomly assigned to four groups: (1) Sham group, (2) SCI group, (3) TGN-020 group (aquaporin-4, AQP4, blocking agent), (4) PD98059 group (ERK blocking agent). A well SCI model was established by compressing the thoracic vertebra 10 level (weight 35 g, time 5 min) in rats. Western blotting and immunofluorescence staining were used to measure the expression of associated proteins after SCI. HE staining and Nissl staining were performed to detect the morphological changes of spinal cords and the number of surviving neurons following SCI, respectively. The Basso-Beattie-Bresnahan open-field rating scale was used to evaluate functional locomotor recovery following SCI in rats. Our results demonstrated that SCI significantly induced the upregulation of aquaporin-4, p-ERK1/2, glial fibrillary acidic protein, proliferating cell nuclear antigen, and proinflammatory cytokines (tumor necrosis factor-α, interleukin-6 and interleukin-1β). However, treatment with TGN-020 or PD98059 could effectively inhibit astrocyte proliferation and proinflammatory cytokine release, preserve the number of surviving ventral horn neurons, and subsequently improve the locomotor function of rats after SCI. Interestingly, the SCI-induced elevation of AQP4 expression was downregulated by p-ERK1/2 inhibition, suggesting that blocking ERK1/2 phosphorylation could attenuate astrocyte activation and inflammatory processes through negative regulation of AQP4. Therefore, p-ERK1/2 blockade may be employed as a therapeutic target for SCI.

Orexin-A potentiates glycine currents by activating OX1R and IP3/Ca2+/PKC signaling pathways in spinal cord ventral horn neurons

Orexin-A/B modulates multiple physical functions by activating their receptors (OX1R and OX2R), but its effects in the spinal cord motor control remain unknown. Using acute separation (by digestive enzyme) of cells and patch-clamp recordings, we aimed to investigate the effect and mechanisms of orexin-A on the glycine receptors in the spinal cord ventral horn neurons. Orexin-A potentiated the glycine currents by activating OX1R. In Ca2+-free extracellular solution, orexin-A still increased the glycine currents. While, the orexin-A-induced potentiation was blocked when Ca2+ was chelated by internal infusion of BAPTA, and the orexin-A effect was abolished by the IP3 receptor antagonists heparin and Xe-C. The PKC inhibitor Bis-IV nullified the orexin-A effect. In addition, orexin-A did not cause a further enhancement of the glycine currents after bath application of the PKC activator PMA. In conclusion, after OX1R is activated, a distinct IP3/Ca2+-dependent PKC signaling pathway, is likely responsible for the orexin-A potentiation on glycine currents in the spinal cord ventral horn neurons.

Intrathecal administration of the extracellular vesicles derived from human Wharton’s jelly stem cells inhibit inflammation and attenuate the activity of inflammasome complexes after spinal cord injury in rats

Activation of inflammasome complexes during spinal cord injury (SCI) lead to conversion of pro-inflammatory cytokines, interleukin-1beta (IL-1β) and interleukin-18 (IL-18) to their active form to initiates the neuroinflammation. Mesenchymal stem cells (MSCs) showed anti-inflammatory properties through their extracellular vehicles (EVs). We investigated immunomodulatory potential of human Wharton's jelly mesenchymal stem cells derived extracellular vesicles (hWJ-MSC-EVs) on inflammasome activity one week after SCI in rats. The gene expression and protein level of IL-1β, IL-18, tumor necrosis factor alpha (TNF-α) and caspase1, were assessed by QPCR and western blotting. Immunohistochemistry (IHC) was done to measure the glial fibrillary acidic protein (GFAP) and Nestin expression. Cell death, histological evaluation and hind limb locomotion was studied by TUNEL assay, Nissl staining and Basso, Beattie, Bresnaham (BBB), respectively. Our finding represented that intrathecally administrated of hWJ-MSC-EVs significantly attenuated expression of the examined factors in both mRNA (P < 0.05 and P ≤ 0.01) and protein levels (P < 0.05 and P ≤ 0.01), decreased GFAP and increased Nestin expression (P < 0.05), reduced cell death and revealed the higher number of typical neurons in ventral horn of spinal cord. Consequently, progress in locomotion. We came to the conclusion that hWJ-MSC-EVs has the potential to control the inflammasome activity after SCI in rats. Moreover, EVs stimulated the neural progenitor cells and modulate the astrocyte activity.

Etomidate reduces excitability of the neurons and suppresses the function of nAChR ventral horn in the spinal cord of neonatal rats

To investigate the effects of etomidate on electrophysiological properties and nicotinic acetylcholine receptors (nAChRs) of ventral horn neurons in the spinal cord. The spinal cord containing lumbosacral enlargement was isolated from 19 neonatal SD rats aged 7-12 days. The spinal cord were sliced and digested with papain (0.18 g/30 mL artificial cerebrospinal fluid) and incubated for 40 min. At the ventral horn, acute mechanical separation of neurons was performed with fire-polished Pasteur pipettes, and perforated patch-clamp recordings combined with pharmacological methods were employed on the adherent healthy neurons. In current-clamp mode, the spontaneous action potential (AP) of the ventral horn neurons in the spinal cord was recorded. The effects of pretreatment with different concentrations of etomidate on AP recorded in the ventral horn neurons were examined. In the voltage-clamp mode, nicotine was applied to induce inward currents in the ventral horn neurons, and the effect of pretreatment with etomidate on the inward currents induced by nicotine were examined with different etomidate concentrations, different holding potentials and different use time. The isolated ventral horn neurons were in good condition with large diverse somata and intact processes. The isolated spinal ventral horn neurons (n=21) had spontaneous action potentials, and were continuously perfused for 2 min with 0.3, 3.0 and 30.0 μmol/L etomidate. Compared with those before administration, the AP amplitude, spike potential amplitude and overshoot were concentration-dependently suppressed (P &lt; 0.01), and spontaneous discharge frequency was obviously reduced (P &lt; 0.01, n=12). The APs of the other 9 neurons were completely abolished by etomidate at 3.0 or 30 μmol/L. At the same holding potential (VH=-70 mV), pretreatment with 0.3, 3.0 or 30.0 μmol/L etomidate for 2 min concentration-dependently suppressed the current amplitude induced by 0.4 mmol/L nicotine (P &lt; 0.01, n=7). At the holding potentials of - 30, - 50, and - 70 mV, pretreatment with 30.0 μmol/L etomidate for 2 min voltage-dependently suppressed the current amplitude induced by 0.4 mmol/L nicotine (P &lt; 0.01, n=6 for each holding potential). During the 6 min of 30.0 μmol/L etomidate pretreatment, the clamped cells were exposed to 0.4 mmol/L nicotine for 4 times at 0, 2, 4, and 6 min (each exposure time was 2 s), and the nicotinic current amplitude decreased gradually as the number of exposures increased. But at the same concentration, two nicotine exposures (one at the beginning and the other at the end of the 6 min pretreatment) resulted in a significantly lower inhibition rate compared with 4 nicotine exposures (P &lt; 0.01, n=6). etomidate reduces the excitability of the spinal ventral neurons in a concentration-dependent manner and suppresses the function of nAChR in a concentration-, voltage-, and use-dependent manner.

Blocking CTGF/CCN2 reverses neural fibrosis and sensorimotor declines in a rat model of overuse-induced median mononeuropathy.

Encapsulation of median nerves is a hallmark of overuse‐induced median mononeuropathy and contributes to functional declines. We tested if an antibody against CTGF/CCN2 (termed FG‐3019 or Pamrevlumab) reduces established neural fibrosis and sensorimotor declines in a clinically relevant rodent model of overuse in which median mononeuropathy develops. Young adult female rats performed a high repetition high force (HRHF) lever‐pulling task for 18 weeks. Rats were then euthanised at 18 weeks (HRHF untreated), or rested and systemically treated for 6 weeks with either an anti‐CCN2 monoclonal antibody (HRHF‐Rest/FG‐3019) or IgG (HRHF‐Rest/IgG), with results compared with nontask control rats. Neuropathology was evident in HRHF‐untreated and HRHF‐Rest/IgG rats as increased perineural collagen deposition and degraded myelin basic protein (dMBP) in median nerves, and increased substance P in lower cervical dorsal root ganglia (DRG), compared with controls. Both groups showed functional declines, specifically, decreased sensory conduction velocity in median nerves, noxious cold temperature hypersensitivity, and grip strength declines, compared with controls. There were also increases of ATF3‐immunopositive nuclei in ventral horn neurons in HRHF‐untreated rats, compared with controls (which showed none). FG‐3019‐treated rats showed no increase above control levels of perineural collagen or dMBP in median nerves, Substance P in lower cervical DRGs, or ATF3‐immunopositive nuclei in ventral horns, and similar median nerve conduction velocities and thermal sensitivity, compared with controls. We hypothesize that neural fibrotic processes underpin the sensorimotor declines by compressing or impeding median nerves during movement, and that inhibiting fibrosis using an anti‐CCN2 treatment reverses these effects.

Expression and regulation of FRMD6 in mouse DRG neurons and spinal cord after nerve injury

FRMD6, a member of the group of FERM-domain proteins, is involved both in communication between cells, interactions with extracellular matrix, cellular apoptotic and regenerative mechanisms. FRMD6 was first discovered in the rodent sciatic nerve, and in the present immunohistochemical study we investigated the distribution of FRMD6 in the dorsal root ganglia (DRGs), sciatic nerve and spinal cord following sciatic nerve injury. FRMD6-immunoreactivity was found in the cytoplasm, nucleus or both, and in a majority of DRG neurons. FRMD6-immunoreactivity co-existed with several well-known neuronal markers, including calcitonin gene-related peptide, isolectin B4 and neurofilament 200 in mouse DRGs. After peripheral nerve injury, the FRMD6 mRNA levels and the overall percentage of FRMD6-positive neuron profiles (NPs) were decreased in ipsilateral lumbar DRGs, the latter mainly affecting small size neurons with cytoplasmic localization. Conversely, the proportion of NPs with nuclear FRMD6-immunoreactivity was significantly increased. In the sciatic nerve, FRMD6-immunoreactivity was observed in non-neuronal cells and in axons, and accumulated proximally to a ligation of the nerve. In the spinal cord FRMD6-immunoreactivity was detected in neurons in both dorsal and ventral horns, and was upregulated in ipsilateral dorsal horn after peripheral nerve axotomy. Our results demonstrate that FRMD6 is strictly regulated by peripheral nerve injury at the spinal level.