Restoration of Visual Function by Enhancing Conduction in Regenerated Axons.

Although a number of repair strategies have been shown to promote axon outgrowth following neuronal injury in the mammalian CNS, it remains unclear whether regenerated axons establish functional synapses and support behavior. Here, in both juvenile and adult mice, we show that either PTEN and SOCS3 co-deletion, or co-overexpression of osteopontin (OPN)/insulin-like growth factor 1 (IGF1)/ciliary neurotrophic factor (CNTF), induces regrowth of retinal axons and formation of functional synapses in the superior colliculus (SC) but not significant recovery of visual function. Further analyses suggest that regenerated axons fail to conduct action potentials from the eye to the SC due to lack of myelination. Consistent with this idea, administration of voltage-gated potassium channel blockers restores conduction and results in increased visual acuity. Thus, enhancing both regeneration and conduction effectively improves function after retinal axon injury.

Design and evolution of an enzyme with a non-canonical organocatalytic mechanism.

The combination of computational design and laboratory evolution is a powerful and potentially versatile strategy for the development of enzymes with new functions1-4. However, the limited functionality presented by the genetic code restricts the range of catalytic mechanisms that are accessible in designed active sites. Inspired by mechanistic strategies from small-molecule organocatalysis5, here we report the generation of a hydrolytic enzyme that uses Nδ-methylhistidine as a non-canonical catalytic nucleophile. Histidine methylation is essential for catalytic function because it prevents the formation of unreactive acyl-enzyme intermediates, which has been a long-standing challenge when using canonical nucleophiles in enzyme design6-10. Enzyme performance was optimized using directed evolution protocols adapted to an expanded genetic code, affording a biocatalyst capable of accelerating ester hydrolysis with greater than 9,000-fold increased efficiency over free Nδ-methylhistidine in solution. Crystallographic snapshots along the evolutionary trajectory highlight the catalytic devices that are responsible for this increase in efficiency. Nδ-methylhistidine can be considered to be a genetically encodable surrogate of the widely employed nucleophilic catalyst dimethylaminopyridine11, and its use will create opportunities to design and engineer enzymes for a wealth of valuable chemical transformations.

Mechanism of 4-aminopyridine inhibition of the lysosomal channel TMEM175.

Transmembrane protein 175 (TMEM175) is an evolutionarily distinct lysosomal cation channel whose mutation is associated with the development of Parkinson's disease. Here, we present a cryoelectron microscopy structure and molecular simulations of TMEM175 bound to 4-aminopyridine (4-AP), the only known small-molecule inhibitor of TMEM175 and a broad K+ channel inhibitor, as well as a drug approved by the Food and Drug Administration against multiple sclerosis. The structure shows that 4-AP, whose mode of action had not been previously visualized, binds near the center of the ion conduction pathway, in the open state of the channel. Molecular dynamics simulations reveal that this binding site is near the middle of the transmembrane potential gradient, providing a rationale for the voltage-dependent dissociation of 4-AP from TMEM175. Interestingly, bound 4-AP rapidly switches between three predominant binding poses, stabilized by alternate interaction patterns dictated by the twofold symmetry of the channel. Despite this highly dynamic binding mode, bound 4-AP prevents not only ion permeation but also water flow. Together, these studies provide a framework for the rational design of novel small-molecule inhibitors of TMEM175 that might reveal the role of this channel in human lysosomal physiology both in health and disease.

Involvement of GABAergic Interneuron Subtypes in 4-Aminopyridine-Induced Seizure-Like Events in Mouse Entorhinal Cortex in Vitro.

Single-unit recordings performed in temporal lobe epilepsy patients and in models of temporal lobe seizures have shown that interneurons are active at focal seizure onset. We performed simultaneous patch-clamp and field potential recordings in entorhinal cortex slices of GAD65 and GAD67 C57BL/6J male mice that express green fluorescent protein in GABAergic neurons to analyze the activity of specific interneuron (IN) subpopulations during acute seizure-like events (SLEs) induced by 4-aminopyridine (4-AP; 100 µm). IN subtypes were identified as parvalbuminergic (INPV, n = 17), cholecystokinergic (INCCK), n = 13], and somatostatinergic (INSOM, n = 15), according to neurophysiological features and single-cell digital PCR. INPV and INCCK discharged at the start of 4-AP-induced SLEs characterized by either low-voltage fast or hyper-synchronous onset pattern. In both SLE onset types, INSOM fired earliest before SLEs, followed by INPV and INCCK discharges. Pyramidal neurons became active with variable delays after SLE onset. Depolarizing block was observed in ∼50% of cells in each INs subgroup, and it was longer in IN (∼4 s) than in pyramidal neurons (<1 s). As SLE evolved, all IN subtypes generated action potential bursts synchronous with the field potential events leading to SLE termination. High-frequency firing throughout the SLE occurred in one-third of INPV and INSOM We conclude that entorhinal cortex INs are very active at the onset and during the progression of SLEs induced by 4-AP. These results support earlier in vivo and in vivo evidence and suggest that INs have a preferential role in focal seizure initiation and development.SIGNIFICANCE STATEMENT Focal seizures are believed to result from enhanced excitation. Nevertheless, we and others demonstrated that cortical GABAergic networks may initiate focal seizures. Here, we analyzed for the first time the role of different IN subtypes in seizures generated by 4-aminopyridine in the mouse entorhinal cortex slices. We found that in this in vitro focal seizure model, all IN types contribute to seizure initiation and that INs precede firing of principal cells. This evidence is in agreement with the active role of GABAergic networks in seizure generation.

Pharmacological management of secondary chronic spinal cord injury: a systematic review.

INTRODUCTION: Spinal cord injury (SCI) may bring lifelong consequences for affected patients and a high financial burden to the health care system. SOURCE OF DATA: Published peer-reviewed scientific articles identified from EMBASE, Google Scholar, PubMed and Scopus. AREAS OF AGREEMENT: Surgery and blood pressure management are the main targets in acute SCI to avoid secondary damage. AREAS OF CONTROVERSY: The management of secondary chronic SCI is challenging, with unpredictable outcomes. GROWING POINTS: Given the lack of consensus on pharmacological therapy for acute and secondary chronic SCI, the present study analyses the currently available drugs and treatment options to manage secondary chronic SCI. AREAS TIMELY FOR DEVELOPING RESEARCH: Different approaches exist for the pharmacological management of secondary chronic SCI. One of the most investigated drugs, 4-aminopyridine, improves central motor conduction and shows improvement in neurological signs. Positive results in different areas have been observed in patients receiving the anti-spastic drugs tizanidine and baclofen or Granulocyte colony-stimulating factor. Growth hormone showed only minimal or no significant effects, and the therapy of secondary chronic SCI with riluzole has been poorly researched to date.

Cell-specific switch for epileptiform activity: critical role of interneurons in the mouse subicular network.

During epileptic seizures, neuronal network activity is hyper synchronized whereby GABAergic parvalbumin-interneurons may have a key role. Previous studies have mostly utilized 4-aminopyridine to induce epileptiform discharges in brain slices from healthy animals. However, it is not clear if the seizure-triggering ability of parvalbumin-interneurons also holds true without the use of external convulsive agents. Here, we investigate whether synchronized activation of parvalbumin-interneurons or principal cells can elicit epileptiform discharges in subiculum slices of epileptic mice. We found that selective synchronized activation of parvalbumin-interneurons or principal cells with optogenetics do not result in light-induced epileptiform discharges (LIEDs) neither in epileptic nor in normal brain slices. Adding 4-aminopyridine to slices, activation of parvalbumin-interneurons still failed to trigger LIEDs. In contrast, such activation of principal neurons readily generated LIEDs with features resembling afterdischarges. When GABAA receptor blocker was added to the perfusion medium, the LIEDs were abolished. These results demonstrate that in subiculum, selective synchronized activation of principal excitatory neurons can trigger epileptiform discharges by recruiting a large pool of downstream interneurons. This study also suggests region-specific role of principal neurons and interneurons in ictogenesis, opening towards differential targeting of specific brain areas for future treatment strategies tailored for individual patients with epilepsy.

Performance Measures and Plasma Biomarker Levels in Patients with Multiple Sclerosis after 14 Days of Fampridine Treatment: An Explorative Study.

Peripheral cytokine levels may serve as biomarkers for treatment response and disease monitoring in patients with multiple sclerosis (pwMS). The objectives were to assess changes in plasma biomarkers in PwMS after 14 days of fampridine treatment and to explore correlations between changes in performance measures and plasma biomarkers. We included 27 PwMS, 14 women and 13 men, aged 52.0 ± 11.6 years, with a disease duration of 17 ± 8.5 years, and an Expanded Disability Status Scale of 6 [IQR 5.0/6.5]. Gait and hand function were assessed using performance tests completed prior to fampridine and after 14 days of treatment. Venous blood was obtained, and chemiluminescence analysis conducted to assess plasma cytokines and neurodegenerative markers. All performance measures demonstrated improvements. Biomarkers showed decreased tumor necrosis factor (TNF) receptor-2 levels. Associations were found between change scores in (i) Six Spot Step Test and Interleukin (IL)-2, IL-8, and IL-17 levels; (ii) timed 25-foot walk and interferon-γ, IL-2, IL-8, TNF-α, and neurofilament light levels, and (iii) 12-Item Multiple Sclerosis Walking Scale and IL-17 levels. The associations may reflect increased MS-related inflammatory activity rather than a fampridine-induced response or that a higher level of inflammation induces a better response to fampridine.

Potassium channels at the crossroads of neuroinflammation and myelination in experimental models of multiple sclerosis.

Multiple sclerosis (MS) is a chronic demyelinating disease of the central nervous system (CNS), characterized by the presence of localized demyelinating lesions accompanied by an inflammatory reaction, evidently leading to neurodegeneration. A number of ion channels have been implicated in the progression of MS, most notably in cell types involved in the immune response. In the present study, we investigated the implication of two ion channel isoforms, Kv1.1 and Kv1.3, in experimental models of neuroinflammation and demyelination. Immunohistochemical staining of brain sections from the mouse cuprizone model displayed high levels Kv1.3 expression. In an astroglial cellular model of inflammation, stimulation with LPS resulted also in a higher expression of Kv1.1 and Kv1.3, while the introduction of 4-Aminopyridine (4-AP) exacerbated the release of pro-inflammatory chemokine CXCL10. In the oligodendroglial cellular model of demyelination, the alteration in expression levels of Kv1.1 and Kv1.3 may be correlated with that of MBP levels. Indirect co-culture was attempted to further understand the communication between astrocytes and oligodendrocytes, The addition of reactive astrocytes' secretome significantly inhibited the production of MBP, this inhibition was accompanied by an alteration in the expression of Kv1.1 and Kv1.3. The addition of 4-AP in this case did not alleviate the decrease in MBP production. In conclusion, the use of 4-AP generated controversial results, suggesting 4-AP may be used in the early stages of the disease or in the remission phases to stimulate myelination, yet in induced toxic inflammatory environment, 4-AP exacerbated this effect.

Cell-specific NFIA upregulation promotes epileptogenesis by TRPV4-mediated astrocyte reactivity.

BACKGROUND: The astrocytes in the central nervous system (CNS) exhibit morphological and functional diversity in brain region-specific pattern. Functional alterations of reactive astrocytes are commonly present in human temporal lobe epilepsy (TLE) cases, meanwhile the neuroinflammation mediated by reactive astrocytes may advance the development of hippocampal epilepsy in animal models. Nuclear factor I-A (NFIA) may regulate astrocyte diversity in the adult brain. However, whether NFIA endows the astrocytes with regional specificity to be involved in epileptogenesis remains elusive. METHODS: Here, we utilize an interference RNA targeting NFIA to explore the characteristics of NFIA expression and its role in astrocyte reactivity in a 4-aminopyridine (4-AP)-induced seizure model in vivo and in vitro. Combined with the employment of a HA-tagged plasmid overexpressing NFIA, we further investigate the precise mechanisms how NIFA facilitates epileptogenesis. RESULTS: 4-AP-induced NFIA upregulation in hippocampal region is astrocyte-specific, and primarily promotes detrimental actions of reactive astrocyte. In line with this phenomenon, both NFIA and vanilloid transient receptor potential 4 (TRPV4) are upregulated in hippocampal astrocytes in human samples from the TLE surgical patients and mouse samples with intraperitoneal 4-AP. NFIA directly regulates mouse astrocytic TRPV4 expression while the quantity and the functional activity of TRPV4 are required for 4-AP-induced astrocyte reactivity and release of proinflammatory cytokines in the charge of NFIA upregulation. NFIA deficiency efficiently inhibits 4-AP-induced TRPV4 upregulation, weakens astrocytic calcium activity and specific astrocyte reactivity, thereby mitigating aberrant neuronal discharges and neuronal damage, and suppressing epileptic seizure. CONCLUSIONS: Our results uncover the critical role of NFIA in astrocyte reactivity and illustrate how epileptogenic brain injury initiates cell-specific signaling pathway to dictate the astrocyte responses.

Metabolic Stability of the Demyelination Positron Emission Tomography Tracer [18F]3-Fluoro-4-Aminopyridine and Identification of Its Metabolites.

[18F]3-fluoro-4-aminopyridine ([18F]3F4AP) is a positron emission tomography (PET) tracer for imaging demyelination based on the multiple sclerosis drug 4-aminopyridine (4AP, dalfampridine). This radiotracer was found to be stable in rodents and nonhuman primates imaged under isoflurane anesthesia. However, recent findings indicate that its stability is greatly decreased in awake humans and mice. Since both 4AP and isoflurane are metabolized primarily by cytochrome P450 enzymes, particularly cytochrome P450 family 2 subfamily E member 1 (CYP2E1), we postulated that this enzyme may be responsible for the metabolism of 3F4AP. Here, we investigated the metabolism of [18F]3F4AP by CYP2E1 and identified its metabolites. We also investigated whether deuteration, a common approach to increase the stability of drugs, could improve its stability. Our results demonstrate that CYP2E1 readily metabolizes 3F4AP and its deuterated analogs and that the primary metabolites are 5-hydroxy-3F4AP and 3F4AP N-oxide. Although deuteration did not decrease the rate of the CYP2E1-mediated oxidation, our findings explain the diminished in vivo stability of 3F4AP compared with 4AP and further our understanding of when deuteration may improve the metabolic stability of drugs and PET ligands. SIGNIFICANCE STATEMENT: The demyelination tracer [18F]3F4AP was found to undergo rapid metabolism in humans, which could compromise its utility. Understanding the enzymes and metabolic products involved may offer strategies to reduce metabolism. Using a combination of in vitro assays and chemical syntheses, this report shows that cytochrome P450 enzyme CYP2E1 is likely responsible for [18F]3F4AP metabolism, that 4-amino-5-fluoroprydin-3-ol (5-hydroxy-3F4AP, 5OH3F4AP) and 4-amino-3-fluoropyridine 1-oxide (3F4AP N-oxide) are the main metabolites, and that deuteration is unlikely to improve the stability of the tracer in vivo.

In vitro fertilization with frozen embryo transfer increased histamine-mediated contractile sensitivity via PKCß in human umbilical vein.

OBJECTIVE: In vitro fertilization-embryo transfer (IVF-ET) technologies (especially frozen ET) have been widely used, which might affect maternal and fetal health. Information regarding influence of IVF-ET on the vasoconstriction of human umbilical vein (HUV) is limited. This study determined effects of frozen ET on histamine-mediated vascular responses in HUV and related mechanisms. METHODS AND RESULTS: HUVs were collected from frozen ET conceived pregnancy and spontaneously conceived pregnancy (control). Histamine concentration in umbilical plasma was higher in frozen ET group than the control. Histamine-mediated contractile response curve was left-shifted in the frozen ET group when comparing with the control. In isolated HUV rings, H1R showed a critical role in regulating vascular constriction, while H2R played little roles in regulating vessel tone. Iberiotoxin and 4-aminopyridine didn't significantly change histamine-mediated constriction in HUVs. Histamine-induced vasoconstrictions were significantly decreased by nifedipine, KN93, or GF109203X, while the inhibitory effects were significantly greater in the frozen ET group in comparison to the control. The constrictions by Bay K8644, phenylephrine, or PDBu were stronger in frozen ET, respectively. There was a decrease in the protein expressions of H1R and H2R, an increase in protein expressions of BKCaα and PKCß. CONCLUSIONS: Histamine-induced constriction in HUV was mainly via H1R. The increased sensitivity to histamine in HUV following frozen ET cycles were linked to the enhanced PKCß protein expression and function. The new data and findings in this study provide important insight into influences of frozen ET on fetal vessel development and potential influence in long-term.

Excitatory-inhibitory mismatch shapes node recruitment in an epileptic network.

OBJECTIVE: Focal epilepsy is thought to be a network disease, in which epileptiform activity can spread noncontiguously through the brain via highly interconnected nodes, or hubs, within existing networks. Animal models confirming this hypothesis are scarce, and our understanding of how distant nodes are recruited is also lacking. Whether interictal spikes (IISs) also create and reverberate through a network is not well understood. METHODS: We injected bicuculline into the S1 barrel cortex and employed multisite local field potential and Thy-1 and parvalbumin (PV) cell mesoscopic calcium imaging during IISs to monitor excitatory and inhibitory cells in two monosynaptically connected nodes and one disynaptically connected node: ipsilateral secondary motor area (iM2), contralateral S1 (cS1), and contralateral secondary motor area (cM2). Node participation was analyzed with spike-triggered coactivity maps. Experiments were repeated with 4-aminopyridine as an epileptic agent. RESULTS: We found that each IIS reverberated throughout the network, differentially recruiting both excitatory and inhibitory cells in all connected nodes. The strongest response was found in iM2. Paradoxically, node cM2, which was connected disynaptically to the focus, was recruited more intensely than node cS1, which was connected monosynaptically. The explanation for this effect could be found in node-specific excitatory/inhibitory (E/I) balance, as cS1 demonstrated greater PV inhibitory cell activation compared with cM2, where Thy-1 excitatory cells were more heavily recruited. SIGNIFICANCE: Our data show that IISs spread noncontiguously by exploiting fiber pathways that connect nodes in a distributed network and that E/I balance plays a critical role in node recruitment. This multinodal IIS network model can be used to investigate cell-specific dynamics in the spatial propagation of epileptiform activity.

4-Aminopyridine Protects Nigral Dopaminergic Neurons in the MPTP Mouse Model of Parkinson's Disease.

Various pharmacological blockers targeting K+ channel have been identified to be related to the treatment of Parkinson's disease (PD). Previous studies showed that 4-Aminopyridine (4-AP), a wide-spectrum K+ channel blocker, was able to attenuate apomorphine-induced rotation in parkinsonism rats, indicating the possible beneficial effects in attenuation of PD motor symptoms. However, it is unclear whether 4-AP exhibits neuroprotective effects against the neurodegeneration of substantia nigra (SN)-striatum system in PD. In this study, the 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated mouse model was employed to evaluate the neuroprotective effects of 4-AP. Results showed that 4-AP inhibited MPTP-induced dopaminergic neuronal loss in the SN as well as dopamine depletion in the striatum. Behavior indexes of open field test and rotarod test confirmed that 4-AP attenuated MPTP-induced motor deficits. We also showed that 4-AP treatment could significantly attenuate the MPTP-induced increase in malonaldehyde (MDA) levels and decrease in superoxide dismutase (SOD) levels. Additionally, MPTP significantly reduced the Bcl-2 expression and promoted the Caspase-3 activation; 4-AP protected dopaminergic neurons against MPTP-induced neurotoxicity by reversing these changes. These results indicate that 4-AP exerts a neuroprotective effect on dopaminergic neurons against MPTP by decreasing oxidative stress and apoptosis. This provides a promising therapeutic target for the treatment of PD.

Elabela relaxes rat pulmonary artery and trachea via BKCa, KV, and KATP channels.

OBJECTIVE: Elabela is a newly discovered peptide hormone. This study aimed to determine the functional effects and mechanisms of action of elabela in rat pulmonary artery and trachea. MATERIALS AND METHODS: Vascular rings isolated from the pulmonary arteries of male Wistar Albino rats were placed in chambers in the isolated tissue bath system. The resting tension was set to 1 g. After the equilibration period, the pulmonary artery rings were contracted with 10-6 M phenylephrine. Once a stable contraction was achieved, elabela was applied cumulatively (10-10-10-6 M) to the vascular rings. To determine the vasoactive effect mechanisms of elabela, the specified experimental protocol was repeated after the incubation of signaling pathway inhibitors and potassium channel blockers. The effect and mechanisms of action of elabela on tracheal smooth muscle were also determined by a similar protocol. RESULTS: Elabela exhibited a concentration-dependent relaxation in the precontracted rat pulmonary artery rings (p < .001). Maximal relaxation level was 83% (pEC50: 7.947 CI95(7.824-8.069)). Removal of the endothelium, indomethacin incubation, and dideoxyadenosine incubation significantly decreased the vasorelaxant effect levels of elabela (p < .001). Elabela-induced vasorelaxation levels were significantly reduced after iberiotoxin, glyburide, and 4-Aminopyridine administrations (p < .001). L-NAME, methylene blue, apamin, TRAM-34, anandamide, and BaCl2 administrations did not cause a significant change in the vasorelaxant effect level of elabela (p = 1.000). Elabela showed a relaxing effect on precontracted tracheal rings (p < .001). Maximal relaxation level was 73% (pEC50: 6.978 CI95(6.791-7.153)). The relaxant effect of elabela on tracheal smooth muscle was decreased significantly after indomethacin, dideoxyadenosine, iberiotoxin, glyburide, and 4-Aminopyridine incubations (p < .001). CONCLUSIONS: Elabela exerted a prominent relaxant effect in the rat pulmonary artery and trachea. Intact endothelium, prostaglandins, cAMP signaling pathway, and potassium channels (BKCa, KV, and KATP channels) are involved in the vasorelaxant effect of elabela. Prostaglandins, cAMP signaling pathway, BKCa channels, KV channels, and KATP channels also contribute to elabela-induced tracheal smooth muscle relaxant effect.

Fampridine for gait imbalance in patients with multiple sclerosis (MS): a systematic review and meta-analysis.

BACKGROUND: Gait imbalance is one of the frequent complications in subjects with multiple sclerosis (MS). Fampridine (4-aminopyridine) is a potassium-channel blocker that is administered for gait imbalance in MS. Different studies showed the effects of fampridine on gait status based on various tests in subjects with MS. Some showed significant improvement after treatment, and others did not. So, we designed this systematic review, and meta-analysis to estimate the pooled effects of fampridine on gait status in patients with MS. METHODS: The main goal is the evaluation of times of different gait test pre and post fampridine treatment. Two independent expert researchers conducted a systematic and comprehensive search in PubMed, Scopus, EMBASE, Web of Science, and Google Scholar and also gray literature, including references of the references and conference abstracts. The search was done on September 16, 2022. Before-after studies trials reporting scores of the walking tests. We extracted data regarding the total number of participants, first author, publication year, country of origin, mean age, Expanded Disability Status Scale (EDSS), and the results of walking tests. RESULTS: The literature search revealed 1963 studies; after deleting duplicates, 1098 studies remained. Seventy-seven full texts were evaluated. Finally, 18 studies were included for meta-analysis, while most of them were not placebo-controlled trials. The most frequent country of origin was Germany, and the mean age and EDSS ranged between 44 and 56 years and 4 and 6, respectively. The studies were published between 2013 and 2019. The pooled standardized mean difference (SMD) (after-before) of the MS Walking Scale (MSWS-12) was - 1.97 (95%CI: - 1.7, - 1.03) (I2 = 93.1%, P < 0.001). The pooled SMD (after-before) of the six-minute walk test (6MWT) was 0.49 (95%CI: 0.22, - 0.76) (I2 = 0%, P = 0.7). The pooled SMD (after-before) of T Timed 25-Foot Walk (T25FW) was - 0.99(95%CI: - 1.52, - 0.47) (I2 = 97.5%, P < 0.001). CONCLUSION: This systematic review and meta-analysis show that fampridine improves gait imbalance in patients with MS.

Fampridine in multiple sclerosis patients with acute phase of cervical transverse myelitis: a double-blind, randomized placebo-controlled trial.

BACKGROUND: Fampridine is a potassium channel blocker drug used to improve walking ability in patients with multiple sclerosis (MS). We evaluated the effect of fampridine in patients with MS in the acute phase of transverse myelitis. METHODS: In a randomized, placebo-controlled trial, 30 patients who had their first episode of cervical myelitis with quadriparesis presentation, with the final diagnosis of MS, were randomly divided into two equal groups. The intervention group received intravenous methylprednisolone (IVMP) for 7 days plus fampridine. The placebo group received IVMP for 7 days plus placebo. To compare the treatment results, we compared the Barthel index (BI) scores of the groups at the start of the trial and the 21st day after the start of treatment. RESULTS: There was no significant difference in baseline characteristics between the intervention and placebo groups in terms of mean age, sex, and mean admission BI (p > 0.05). Mean (SD) admission BI in placebo and intervention groups was 27.20 (7.341) and 27.87(5.78), respectively (p = 0.784). The measured mean (SD) BI after treatment was 48.73 (15.54) in the placebo and 64.93 (11.81) in the intervention group (p = 0.003) after 3 weeks. CONCLUSION: Using fampridine plus IVMP in the acute phase of transverse myelitis in MS patients improved the disease's symptoms and increased the daily activity ability of patients.

Effects of 4-Aminopyridine on Combined Nerve and Muscle Injury and Bone Loss.

PURPOSE: Musculoskeletal injuries are common, and peripheral nerve injury (PNI) causes significant muscle and bone loss within weeks. After PNI, 4-aminopyridine (4-AP) improves functional recovery and muscle atrophy. However, it is unknown whether 4-AP has any effect on isolated traumatic muscle injury and PNI-induced bone loss. METHODS: A standardized crush injury was performed on the sciatic nerve and muscles in mice, and the mice were assigned to receive normal saline or 4-AP treatment daily for 21 days. The postinjury motor and sensory function recovery was assessed, injured muscles were processed for histomorphometry, and the tibial bone was scanned for bone density. RESULTS: 4-Aminopyridine significantly accelerated the postinjury motor and sensory function recovery, improved muscle histomorphometry, increased muscle satellite cell numbers, and shifted muscle fiber types after combined nerve and muscle injury. Importantly, the 4-AP treatment significantly reduced PNI-induced bone loss. In contrast, in the case of isolated muscle injury, 4-AP had no effect on functional recovery and bone density, but it improved muscle-specific histomorphometry to a limited extent. CONCLUSIONS: These findings demonstrate the potential beneficial effects of 4-AP on the recovery of muscle morphology and bone density after combined muscle and nerve injury. CLINICAL RELEVANCE: Nerve injuries frequently involve muscle and result in rapid muscle and bone atrophy. In this scenario, 4-AP, in addition to accelerating nerve functional recovery, might work as an adjunctive agent to improve the recovery of injured muscle and attenuate PNI-induced bone loss.

Plantainoside D Reduces Depolarization-Evoked Glutamate Release from Rat Cerebral Cortical Synaptosomes.

Inhibiting the excessive release of glutamate in the brain is emerging as a promising therapeutic option and is efficient for treating neurodegenerative disorders. The aim of this study is to investigate the effect and mechanism of plantainoside D (PD), a phenylenthanoid glycoside isolated from Plantago asiatica L., on glutamate release in rat cerebral cortical nerve terminals (synaptosomes). We observed that PD inhibited the potassium channel blocker 4-aminopyridine (4-AP)-evoked release of glutamate and elevated concentration of cytosolic Ca2+. Using bafilomycin A1 to block glutamate uptake into synaptic vesicles and EDTA to chelate extracellular Ca2+, the inhibitory effect of PD on 4-AP-evoked glutamate release was prevented. In contrast, the action of PD on the 4-AP-evoked release of glutamate in the presence of dl-TBOA, a potent nontransportable inhibitor of glutamate transporters, was unaffected. PD does not alter the 4-AP-mediated depolarization of the synaptosomal membrane potential, suggesting that the inhibitory effect of PD on glutamate release is associated with voltage-dependent Ca2+ channels (VDCCs) but not the modulation of plasma membrane potential. Pretreatment with the Ca2+ channel blocker (N-type) ω-conotoxin GVIA abolished the inhibitory effect of PD on the evoked glutamate release, as did pretreatment with the protein kinase C inhibitor GF109203x. However, the PD-mediated inhibition of glutamate release was eliminated by applying the mitochondrial Na+/Ca2+ exchanger inhibitor CGP37157 or dantrolene, which inhibits Ca2+ release through ryanodine receptor channels. These data suggest that PD mediates the inhibition of evoked glutamate release from synaptosomes primarily by reducing the influx of Ca2+ through N-type Ca2+ channels, subsequently reducing the protein kinase C cascade.

Refractory Convulsive Status Epilepticus Provoked by Intoxication with Dalfampridine in a Patient with Multiple Sclerosis and Depression Disorder: A Case Report and Literature Review.

PURPOSE: Dalfampridine (DFP) is used to improve motor functions in patients with multiple sclerosis (MS). Overdose of DFP can occur for a variety of reasons and can lead to a state of epilepsy. CASE REPORT: A 24-year-old woman with MS was admitted to hospital with severe sweating and delirium after attempting suicide by overdosing on DFP. At the time of hospitalization, she developed a tonic-clonic seizure that did not respond to immediate intravenous (IV) diazepam injection, followed by intravenous sodium valproate. Therefore, according to the hospital protocol of the neurology department, the patient was intubated and IV infusion of midazolam was started, Due to the persistence of seizures, sodium thiopental began and the patient was admitted to the intensive care unit (ICU). In the ICU, she received an infusion of sodium thiopental and intravenous sodium valproate, monitored by a daily electroencephalogram (EEG). The patient was discharged after four days due to her stable medical condition. CONCLUSION: Epilepsy in case of overuse of DFP should be considered as a life-threatening side effect and timely treatment should be done to prevent damage to the nervous system.

Chronic Pharmacological Increase of Neuronal Activity Improves Sensory-Motor Dysfunction in Spinal Muscular Atrophy Mice.

Dysfunction of neuronal circuits is an important determinant of neurodegenerative diseases. Synaptic dysfunction, death, and intrinsic activity of neurons are thought to contribute to the demise of normal behavior in the disease state. However, the interplay between these major pathogenic events during disease progression is poorly understood. Spinal muscular atrophy (SMA) is a neurodegenerative disease caused by a deficiency in the ubiquitously expressed protein SMN and is characterized by motor neuron death, skeletal muscle atrophy, as well as dysfunction and loss of both central and peripheral excitatory synapses. These disease hallmarks result in an overall reduction of neuronal activity in the spinal sensory-motor circuit. Here, we show that increasing neuronal activity by chronic treatment with the FDA-approved potassium channel blocker 4-aminopyridine (4-AP) improves motor behavior in both sexes of a severe mouse model of SMA. 4-AP restores neurotransmission and number of proprioceptive synapses and neuromuscular junctions (NMJs), while having no effects on motor neuron death. In addition, 4-AP treatment with pharmacological inhibition of p53-dependent motor neuron death results in additive effects, leading to full correction of sensory-motor circuit pathology and enhanced phenotypic benefit in SMA mice. Our in vivo study reveals that 4-AP-induced increase of neuronal activity restores synaptic connectivity and function in the sensory-motor circuit to improve the SMA motor phenotype.SIGNIFICANCE STATEMENT Spinal muscular atrophy (SMA) is a neurodegenerative disease, characterized by synaptic loss, motor neuron death, and reduced neuronal activity in spinal sensory-motor circuits. However, whether these are parallel or dependent events is unclear. We show here that long-term increase of neuronal activity by the FDA-approved drug 4-aminopyridine (4-AP) rescues the number and function of central and peripheral synapses in a SMA mouse model, resulting in an improvement of the sensory-motor circuit and motor behavior. Combinatorial treatment of pharmacological inhibition of p53, which is responsible for motor neuron death and 4-AP, results in additive beneficial effects on the sensory-motor circuit in SMA. Thus, neuronal activity restores synaptic connections and improves significantly the severe SMA phenotype.