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Obesity, a growing global health concern, is closely linked to depression. However, the neural mechanism of association between obesity and depression remains poorly understood. In this study, neural-specific WFS1 deficiency exacerbates the vicious cycle of obesity and depression in mice fed a high-fat diet (HFD), positioning WFS1 as a crucial factor in this cycle. Through human pluripotent stem cells (hESCs) neural differentiation, it is demonstrated that WFS1 regulates Zn 2+ homeostasis and the apoptosis of neural progenitor cells (NPCs) and cerebral organoids by inhibiting the zinc transporter ZnT3 under the situation of dysregulated lipid metabolism. Notably, riluzole regulates ZnT3 expression to maintain zinc homeostasis and protect NPCs from lipotoxicity-induced cell death. Importantly, riluzole, a therapeutic molecule targeting the nervous system, in vivo administration prevents HFD-induced obesity and associated depression. Thus, a WFS1-ZnT3-Zn 2+ axis critical is demonstrated for the vicious cycle of obesity and depression and that riluzole may have the potential to reverse this process against obesity and depression., (© 2024 The Author(s). Advanced Science published by Wiley‐VCH GmbH.)

Background: Ketamine is an intravenous anesthetic that acts as a channel blocker on the N-methyl- d -aspartate (NMDA) receptor, a glutamate receptor subtype. MK-801 is the most potent compound among noncompetitive NMDA receptor antagonists. Ketamine induces loss of the righting reflex (LORR) in rodents, which is one of the indicators of unconsciousness, whereas high doses of MK-801 produce ataxia, but not LORR. In contrast, we previously reported that MK-801 combined with a low dose of the dopamine receptor antagonist haloperidol-induced LORR in mice. To assess a neurophysiologically distinct brain state and demonstrate unconsciousness, electroencephalograms (EEG) need to be examined together with LORR. Therefore, we herein investigated EEG changes after the systemic administration of MK-801 alone or in combination with haloperidol, and compared them with those induced by ketamine, the glutamate release inhibitor riluzole, and the γ-aminobutyric acid type A receptor agonist propofol., Methods: All drugs were intraperitoneally administered to adult male ddY mice (n = 168). General anesthesia was evaluated based on the righting reflex test. Animals who exhibited no righting for more than 30 seconds were considered to have LORR. In a separate group of mice, EEG of the primary visual cortex was recorded before and after the administration of MK-801 (3.0 mg/kg) alone or in combination with haloperidol (0.2 mg/kg), ketamine (150 mg/kg), riluzole (30 mg/kg), or propofol (240 mg/kg). The waveforms recorded were analyzed using EEG power spectra and spectrograms., Results: The high dose of MK-801 alone did not induce LORR, whereas MK-801 combined with haloperidol produced LORR in a dose-dependent manner. Ketamine, riluzole, and propofol also dose-dependently induced LORR. In the EEG study, MK-801 alone induced a significant increase in δ power, while MK-801 plus haloperidol exerted similar effects on not only δ, but also θ and α power during LORR, suggesting that increases in δ, θ, and α power were necessary for LORR. The results obtained on MK-801 plus haloperidol were similar to those on ketamine in the behavioral and EEG studies, except for an increase in γ power by ketamine during LORR. Propofol significantly increased δ, θ, α, and β power during LORR. However, the EEG results obtained using riluzole, which produced a unique pattern of lower amplitude activity spanning most frequencies, markedly differed from those with the other drugs., Conclusions: This study revealed differences in EEG changes induced by various sedatives. The results obtained on MK-801 alone and MK-801 plus haloperidol suggest the importance of dopamine transmission in maintaining the righting reflex., Competing Interests: The authors declare no conflicts of interest., (Copyright © 2024 The Author(s). Published by Wolters Kluwer Health, Inc. on behalf of the International Anesthesia Research Society.)

Pancreatic ductal adenocarcinoma (PDAC) remains the most lethal cancer type. PDAC is characterized by fibrotic, hypoxic, and presumably acidic tumor microenvironment (TME). Acidic TME is an important player in tumor development, progression, aggressiveness, and chemoresistance. The dysregulation of ductal ion transporters/channels might contribute to extracellular pH (pH e ) acidification and PDAC progression. Our aim was to test whether H + /K + -ATPases and pH-sensitive K + channels contribute to these processes and could be targeted by clinically approved drugs. We used human pancreatic cancer cells adapted to various pH e conditions and grown in monolayers and spheroids. First, we created cells expressing pHoran4 at the outer plasma membrane and showed that pantoprazole, the H + /K + -ATPase inhibitor, alkalinized pH e . Second, we used FluoVolt to monitor the membrane voltage (V m ) and showed that riluzole hyperpolarized V m , most likely by opening of pH-sensitive K + channels such as TREK-1. Third, we show that pantoprazole and riluzole inhibited cell proliferation and viability of monolayers and spheroids of cancer cells adapted to various pH e conditions. Most importantly, combination of the two drugs had significantly larger inhibitory effects on PDAC cell survival. We propose that co-targeting H + /K + -ATPases and pH-sensitive K + channels by re-purposing of pantoprazole and riluzole could provide novel acidosis-targeted therapies of PDAC., (© 2024 The Author(s). International Journal of Cancer published by John Wiley & Sons Ltd on behalf of UICC.)

Riluzole, the first clinically approved treatment for amyotrophic lateral sclerosis (ALS), represents a successful example of a drug endowed with a multimodal mechanism of action. In recent years, different series of riluzole-based compounds have been reported, including several agents acting as Multi-Target-Directed Ligands (MTLDs) endowed with neuroprotective effects. Aiming at identical twin structures inspired by riluzole (2a-c), a synthetic procedure was planned, but the reactivity of the system took a different path, leading to the serendipitous isolation of benzo[b][1,4]thiazepines 3a-c and expanded intermediates N-cyano-benzo[b][1,4]thiazepines 4a-c, which were fully characterized. The newly obtained structures 3a-c, bearing riluzole key elements, were initially tested in an in vitro ischemia/reperfusion injury protocol, simulating the cerebral stroke. Results identified compound 3b as the most effective in reverting the injury caused by an ischemia-like condition, and its activity was comparable, or even higher than that of riluzole, exhibiting a concentration-dependent neuroprotective effect. Moreover, derivative 3b completely reverted the release of Lactate Dehydrogenase (LDH), lowering the values to those of the control slices. Based on its very promising pharmacological properties, compound 3b was then selected to assess its effects on voltage-dependent Na + and K + currents. The results indicated that derivative 3b induced a multifaceted inhibitory effect on voltage-gated currents in SH-SY5Y differentiated neurons, suggesting its possible applications in epilepsy and stroke management, other than ALS. Accordingly, brain penetration was also measured for 3b, as it represents an elegant example of a MTDL and opens the way to further ex-vivo and/or in-vivo characterization., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Neuronal hyperexcitability is a key element of many neurodegenerative disorders including the motor neuron disease Amyotrophic Lateral Sclerosis (ALS), where it occurs associated with elevated late sodium current (I NaL ). I NaL results from incomplete inactivation of voltage-gated sodium channels (VGSCs) after their opening and shapes physiological membrane excitability. However, dysfunctional increases can cause hyperexcitability-associated diseases. Here we reveal the atypical binding mechanism which explains how the neuroprotective ALS-treatment drug riluzole stabilises VGSCs in their inactivated state to cause the suppression of I NaL that leads to reversed cellular overexcitability. Riluzole accumulates in the membrane and enters VGSCs through openings to their membrane-accessible fenestrations. Riluzole binds within these fenestrations to stabilise the inactivated channel state, allowing for the selective allosteric inhibition of I NaL without the physical block of Na + conduction associated with traditional channel pore binding VGSC drugs. We further demonstrate that riluzole can reproduce these effects on a disease variant of the non-neuronal VGSC isoform Nav1.4, where pathologically increased I NaL is caused directly by mutation. Overall, we identify a model for VGSC inhibition that produces effects consistent with the inhibitory action of riluzole observed in models of ALS. Our findings will aid future drug design and supports research directed towards riluzole repurposing., (© 2024. The Author(s).)

Background: The metastasis of tumors into bone tissue typically leads to intractable pain that is both very disabling and particularly difficult to manage. We investigated here whether riluzole could have beneficial effects for the treatment of prostate cancer-induced bone pain and how it could influence the development of bone metastasis., Methods: We used a bone pain model induced by intratibial injection of human PC3 prostate cancer cells into male SCID mice treated or not with riluzole administered in drinking water. We also used riluzole in vitro to assess its possible effect on PC3 cell viability and functionality, using patch-clamp., Results: Riluzole had a significant preventive effect on both evoked and spontaneous pain involving the TREK-1 potassium channel. Riluzole did not interfere with PC3-induced bone loss or bone remodeling in vivo. It also significantly decreased PC3 cell viability in vitro. The antiproliferative effect of riluzole is correlated with a TREK-1-dependent membrane hyperpolarization in these cells., Conclusion: The present data suggest that riluzole could be very useful to manage evoked and spontaneous hypersensitivity in cancer-induced bone pain and has no significant adverse effect on cancer progression., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. Conflict of interest statement The authors have declared that no conflict of interest exists., (Copyright © 2024 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

Objective: To examine the effect of combined treatment with Bojungikgi-tang (BJIGT, Buzhong Yiqi Decoction) and riluzole (RZ) in transactive response DNA-binding protein 43 (TDP-43) stress granule (SG) cells, a amyotrophic lateral sclerosis (ALS) cell line using transcriptomic and molecular techniques., Methods: TDP-43 SG cells were pretreated with BJIGT (100 µg/mL), RZ (50 µmol/L), and combined BJIGT (100 µg/mL)/RZ (50 µmol/L) for 6 h before treatment with lipopolysaccharide (LPS, 200 µmol/L). Cell viability assay was performed to elucidate cell toxicity in TDP-43 SC cells using a cell-counting kit-8 (CCK8) assay kit. The expression levels of cell death-related proteins, including Bax, caspase 1, cleaved caspase 3 and DJ1 in TDP-43 SG cells were examined by Western blot analysis. The autophagy-related proteins, including pmTOR/mTOR, LC3b, P62, ATG7 and Bcl-2-associated athanogene 3 (Bag3) were investigated using immunofluorescence and immunoblotting assays., Results: Cell viability assay and Western blot analysis showed that combined treatment with BJIGT and RZ suppressed LPS-induced cell death and expression of cell death-related proteins, including Bax, caspase 1, and DJ1 (P<0.05 or P<0.01). Immunofluorescence and immunoblotting assays showed that combined treatment with BJIGT and RZ reduced LPS-induced formation of TDP-43 aggregates and regulated autophagy-related protein levels, including p62, light chain 3b, Bag3, and ATG7, in TDP-43-expressing cells (P<0.05 or P<0.01)., Conclusion: The combined treatment of BJIGT and RZ might reduce inflammation and regulate autophagy dysfunction in TDP-43-induced ALS., (© 2023. The Chinese Journal of Integrated Traditional and Western Medicine Press and Springer-Verlag GmbH Germany, part of Springer Nature.)

Abstract: Objective: This study aimed to explore the impact of heat stress (HS) on glutamate transmission-dependent expression levels of interleukin-1β (IL-1β) and IL-18 in BV-2 microglial cells. Methods: BV-2 microglial cells were cultured in vitro , with cells maintained at 37°C serving as the control. The HS group experienced incubation at 40°C for 1 h, followed by further culturing at 37°C for 6 or 12 h. The experimental group was preincubated with glutamate, the glutamate antagonist riluzole, or the mGluR5 agonist, 2-chloro-5-hydroxyphenylglycine (CHPG), before HS. Glutamate content in BV-2 culture supernatant was assessed using colorimetric assay. Moreover, mRNA expression levels of EAAT3 and/or mGluR5 in BV-2 cells were determined via quantitative polymerase chain reaction. Interleukins (IL-1β and IL-18) in cell culture supernatant were measured using enzyme-linked immunosorbent assay. Western blot analysis was employed to assess protein levels of IL-1β and IL-18 in BV-2 cells. Results: HS induced a significant release of glutamate and increased the expression levels of mGluR5 and EAAT3 in BV-2 cells. It also triggered the expression levels and release of proinflammatory factors, such as IL-1β and IL-18, synergizing with the effects of glutamate treatment. Preincubation with both riluzole and CHPG significantly reduced HS-induced glutamate release and mitigated the increased expression levels and release of IL-1β and IL-18 induced by HS. Conclusion: The findings confirmed that microglia could be involved in HS primarily through glutamate metabolisms, influencing the expression levels and release of IL-1β and IL-18., Competing Interests: The authors report no conflicts of interest., (Copyright © 2024 The Author(s). Published by Wolters Kluwer Health, Inc. on behalf of the Shock Society.)

Background/aim: Glioblastoma multiforme (GBM)-induced oedema is a major cause of morbidity and mortality among patients with GBM. Dexamethasone (Dex) is the most common corticosteroid used pre-operatively to control cerebral oedema in patients with GBM. Dex is associated with many side effects, and shorter overall survival and progression-free survival of patients with GBM. These negative effects of Dex highlight the need for combinational therapy. Riluzole (Ril), a drug used to treat amyotrophic lateral sclerosis (ALS), is thought to have potential as a treatment for various cancers, with clinical trials underway. Here, we investigated whether Ril could reverse some of the undesirable effects of Dex., Materials and Methods: The effect of Dex, Ril, and Ril-Dex treatment on cell migration was monitored using the xCELLigence system. Cell viability assays were performed using 3-(4, 5-dimethylthiazol)-2, 5-diphenyltetrazolium bromide (MTT). The expression of genes involved in migration, glucose metabolism, and stemness was examined using real-time polymerase chain reaction (PCR)., Results: Pre-treating GBM cells with Ril reduced Dex-induced cell migration and altered Dex-induced effects on cell invasion, stem cell, and glucose metabolism markers. Furthermore, Ril remained effective in killing GBM cells in combination with Dex., Conclusion: Ril, which acts as an anti-tumorigenic drug, mediates some of the negative effects of Dex; therefore, it could be a potential drug to manage the side effects of Dex therapy in GBM., (Copyright © 2024 International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.)

The development of new molecules for the treatment of calmodulin related cardiovascular or neurodegenerative diseases is an interesting goal. In this work, we introduce a novel strategy with four main steps: (1) chemical synthesis of target molecules, (2) Förster Resonance Energy Transfer (FRET) biosensor development and in vitro biological assay of new derivatives, (3) Cheminformatics models development and in vivo activity prediction, and (4) Docking studies. This strategy is illustrated with a case study. Firstly, a series of 4-substituted Riluzole derivatives 1-3 were synthetized through a strategy that involves the construction of the 4-bromoriluzole framework and its further functionalization via palladium catalysis or organolithium chemistry. Next, a FRET biosensor for monitoring Ca 2+ -dependent CaM-ligands interactions has been developed and used for the in vitro assay of Riluzole derivatives. In particular, the best inhibition (80%) was observed for 4-methoxyphenylriluzole 2b. Besides, we trained and validated a new Networks Invariant, Information Fusion, Perturbation Theory, and Machine Learning (NIFPTML) model for predicting probability profiles of in vivo biological activity parameters in different regions of the brain. Next, we used this model to predict the in vivo activity of the compounds experimentally studied in vitro. Last, docking study conducted on Riluzole and its derivatives has provided valuable insights into their binding conformations with the target protein, involving calmodulin and the SK4 channel. This new combined strategy may be useful to reduce assay costs (animals, materials, time, and human resources) in the drug discovery process of calmodulin inhibitors., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

Cancer-related cognitive impairments (CRCI) are neurological complications associated with cancer treatment, and greatly affect cancer survivors' quality of life. Brain-derived neurotrophic factor (BDNF) plays an essential role in neurogenesis, learning and memory. The reduction of BDNF is associated with the decrease in cognitive function in various neurological disorders. Few pre-clinical studies have reported on the effects of chemotherapy and medical stress on BDNF levels and cognition. The present study aimed to compare the effects of medical stress and cisplatin on serum BDNF levels and cognitive function in 9-month-old female Sprague Dawley rats to age-matched controls. Serum BDNF levels were collected longitudinally during cisplatin treatment, and cognitive function was assessed by novel object recognition (NOR) 14 weeks post-cisplatin initiation. Terminal BDNF levels were collected 24 weeks after cisplatin initiation. In cultured hippocampal neurons, we screened three neuroprotective agents, riluzole (an approved treatment for amyotrophic lateral sclerosis), as well as the ampakines CX546 and CX1739. We assessed dendritic arborization by Sholl analysis and dendritic spine density by quantifying postsynaptic density-95 (PSD-95) puncta. Cisplatin and exposure to medical stress reduced serum BDNF levels and impaired object discrimination in NOR compared to age-matched controls. Pharmacological BDNF augmentation protected neurons against cisplatin-induced reductions in dendritic branching and PSD-95. Ampakines (CX546 and CX1739) and riluzole did not affect the antitumor efficacy of cisplatin in vitro. In conclusion, we established the first middle-aged rat model of cisplatin-induced CRCI, assessing the contribution of medical stress and longitudinal changes in BDNF levels on cognitive function, although future studies are warranted to assess the efficacy of BDNF enhancement in vivo on synaptic plasticity. Collectively, our results indicate that cancer treatment exerts long-lasting changes in BDNF levels, and support BDNF enhancement as a potential preventative approach to target CRCI with therapeutics that are FDA approved and/or in clinical study for other indications., Competing Interests: Declaration of competing interest The authors do not have any potential conflicts of interest to disclose., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Riluzole is commonly used as a neuroprotective agent for treating traumatic spinal cord injury (SCI), which works by blocking the influx of sodium and calcium ions and reducing glutamate activity. However, its clinical application is limited because of its poor solubility, short half-life, potential organ toxicity, and insufficient bioabilities toward upregulated inflammation and oxidative stress levels. To address this issue, epigallocatechin gallate (EGCG), a natural polyphenol, was employed to fabricate nanoparticles (NPs) with riluzole to enhance the neuroprotective effects. The resulting NPs demonstrated good biocompatibility, excellent antioxidative properties, and promising regulation effects from the M1 to M2 macrophages. Furthermore, an in vivo SCI model was successfully established, and NPs could be obviously aggregated at the SCI site. More interestingly, excellent neuroprotective properties of NPs through regulating the levels of oxidative stress, inflammation, and ion channels could be fully demonstrated in vivo by RNA sequencing and sophisticated biochemistry evaluations. Together, the work provided new opportunities toward the design and fabrication of robust and multifunctional NPs for oxidative stress and inflammation-related diseases via biological integration of natural polyphenols and small-molecule drugs.

Background: Despite several hundred clinical trials of drugs that initially showed promise, there has been limited clinical improvement in Alzheimer's disease (AD). This may be attributed to the existence of at least 25 abnormal cellular pathways that underlie the disease. It is improbable for a single drug to address all or most of these pathways, thus even drugs that show promise when administered alone are unlikely to produce significant results. According to previous studies, eight drugs, namely, dantrolene, erythropoietin, lithium, memantine, minocycline, piracetam, riluzole, and silymarin, have been found to target multiple pathways that are involved in the development of AD. Among these drugs, riluzole is currently indicated for the treatment of medical conditions in both adult patients and children and has gained increased attention from scientists due to its potential in the excitotoxic hypothesis of neurodegenerative diseases., Objective: The aim of this study was to investigate the effects of drugs on AD based on cellular and molecular mechanisms., Methods: The literature search for this study utilized the Scopus, ScienceDirect, PubMed, and Google Scholar databases to identify relevant articles., Results: Riluzole exerts its effects in AD through diverse pathways including the inhibition of voltage-dependent sodium and calcium channels, blocking AMPA and NMDA receptors and inhibiting the release of glutamic acid release and stimulation of EAAT1-EAAT2., Conclusion: In this review article, we aimed to review the neuroprotective properties of riluzole, a glutamate modulator, in AD, which could benefit patients with the disease., (© 2023 Société Française de Pharmacologie et de Thérapeutique. Published by John Wiley & Sons Ltd.)

Glutamate is the main excitatory neurotransmitter in the brain and plays a leading role in degenerative diseases, such as motor neuron diseases. Riluzole is a glutamate regulator and a therapeutic drug for motor neuron diseases. In this work, the interaction between glutamate and riluzole was studied using cyclic voltammetry and square-wave voltammetry at a glassy carbon electrode (GCE). It was shown that glutamate underwent a two-electron transfer reaction on the GCE surface, and the electrochemical detection limits of glutamate and riluzole were 483 μmol/L and 11.47 μmol/L, respectively. The results confirm that riluzole can promote the redox reaction of glutamate. This work highlights the significance of electrochemical technology in the sensing detection of the interaction between glutamate and related psychotropic drugs., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH, DE part of Springer Nature.)

Background: Unbalanced activity of medium spiny neurons (MSNs) of the direct and indirect pathways mediates reward-related behaviors induced by addictive drugs. Prelimbic (PL) input to MSNs in the nucleus accumbens core (NAcC) plays a key role in cocaine-induced early locomotor sensitization (LS). However, the adaptive plastic changes at PL-to-NAcC synapses underlying early LS remain unclear., Methods: Using transgenic mice and retrograde tracing, we identified NAcC-projecting pyramidal neurons (PNs) in the PL cortex based on the expression of dopamine receptor types (D1R or D2R). To examine cocaine-induced alterations in PL-to-NAcC synapses, we measured excitatory postsynaptic current amplitudes evoked by optostimulation of PL afferents to MSNs. Riluzole was chosen to test the effects of PL excitability on cocaine-induced changes of PL-to-NAcC synapses., Results: NAcC-projecting PNs were segregated into D1R- and D2R-expressing PNs (D1- and D2-PNs, respectively), and their excitability was opposingly regulated by respective dopamine agonists. Both D1- and D2-PNs exhibited balanced innervation of direct MSNs and indirect MSNs in naïve animals. Repeated cocaine injections resulted in biased synaptic strength toward direct MSNs through presynaptic mechanisms in both D1- and D2-PNs, although D2R activation reduced the D2-PN excitability. Under group 1 metabotropic glutamate receptors coactivation, however, D2R activation enhanced the D2-PN excitability. The cocaine-induced rewiring accompanied LS, and both rewiring and LS were precluded by PL infusion of riluzole, which reduced the intrinsic excitability of PL neurons., Conclusions: These findings indicate that cocaine-induced rewiring of PL-to-NAcC synapses correlates well with early behavioral sensitization and that rewiring and LS can be prevented by riluzole-induced reduction of excitability of PL neurons., (Copyright © 2023 Society of Biological Psychiatry. Published by Elsevier Inc. All rights reserved.)

Avulsion injury results in motoneuron death due to the increased excitotoxicity developing in the affected spinal segments. This study focused on possible short and long term molecular and receptor expression alterations which are thought to be linked to the excitotoxic events in the ventral horn with or without the anti-excitotoxic riluzole treatment. In our experimental model the left lumbar 4 and 5 (L4, 5) ventral roots of the spinal cord were avulsed. Treated animals received riluzole for 2 weeks. Riluzole is a compound that acts to block voltage-activated Na + and Ca 2+ channels. In control animals the L4, 5 ventral roots were avulsed without riluzole treatment. Expression of astrocytic EAAT-2 and that of KCC2 in motoneurons on the affected side of the L4 spinal segment were detected after the injury by confocal and dSTORM imaging, intracellular Ca 2+ levels in motoneurons were quantified by electron microscopy. The KCC2 labeling in the lateral and ventrolateral parts of the L4 ventral horn was weaker compared with the medial part of L4 ventral horn in both groups. Riluzole treatment dramatically enhanced motoneuron survival but was not able to prevent the down-regulation of KCC2 expression in injured motoneurons. In contrast, riluzole successfully obviated the increase of intracellular calcium level and the decrease of EAAT-2 expression in astrocytes compared with untreated injured animals. We conclude that KCC2 may not be an essential component for survival of injured motoneurons and riluzole is able to modulate the intracellular level of calcium and expression of EAAT-2., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier GmbH.. All rights reserved.)

Nanocarriers, and especially nanostructured lipid carriers (NLC), represent one of the most effective systems for topical drug administration. NLCs are biodegradable, biocompatible and provide a prolonged drug release. The glutamate release inhibitor Riluzole (RLZ) is a drug currently used for amyotrophic lateral sclerosis (ALS), with anti-proliferative effects potentially beneficial for diseases with excessive cell turnover. However, RLZ possesses low water solubility and high light-sensibility. We present here optimized NLCs loaded with RLZ (RLZ-NLCs) as a potential topical treatment. RLZ-NLCs were prepared by the hot-pressure homogenization method using active essential oils as liquid lipids, and optimized using the design of experiments approach. RLZ-NLCs were developed obtaining optimal properties for dermal application (mean size below 200 nm, negative surface charge and high RLZ entrapment efficacy). In vitro release study demonstrates that RLZ-NLCs allow the successful delivery of RLZ in a sustained manner. Moreover, RLZ-NLCs are not angiogenic and are able to inhibit keratinocyte cell proliferation. Hence, a NLCs delivery system loading RLZ in combination with natural essential oils constitutes a promising strategy against keratinocyte hyperproliferative conditions.

Cancer-related cognitive impairment (CRCI) considerably affects the quality of life of millions of cancer survivors. Brain-derived neurotrophic factor (BDNF) has been shown to promote survival, differentiation, and maintenance of in vivo dentate neurogenesis, and chemotherapy induces a plethora of physiological and cellular alterations, including a decline in neurogenesis and increased neuroinflammation linked with cognitive impairments. In our clinical studies, breast cancer patients treated with doxorubicin (Adriamycin ® , ADR) experienced a significant reduction in the blood levels of BDNF that was associated with a higher risk of CRCI. Our past rodent studies in CRCI have also shown a significant reduction in dentate neurogenesis accompanied by cognitive impairment. In this study, using a female mouse model of ADR-induced cognitive decline, we tested the impact of riluzole (RZ), an orally active BDNF-enhancing medication that is FDA-approved for amyotrophic lateral sclerosis. ADR-treated mice receiving RZ in the drinking water for 1 month showed significant improvements in hippocampal-dependent learning and memory function (spatial recognition), fear extinction memory consolidation, and reduced anxiety-like behavior. RZ prevented chemotherapy-induced reductions of BDNF levels in the hippocampus. Importantly, RZ mitigated chemotherapy-induced loss of newly born, immature neurons, dentate neurogenesis, and neuroinflammation. In conclusion, this data provides pre-clinical evidence for a translationally feasible approach to enhance the neuroprotective effects of RZ treatment to prevent CRCI., (© 2023. The Author(s).)

Few attempts have so far been made to define the mechanisms underlying the hour-long effects of trans-spinal stimulation combined with epidural polarization. In the present study, we investigated the potential involvement of non-inactivating sodium channels in afferent fibres. To this end, riluzole, a blocker of these channels, was administered locally to the dorsal columns close to the site of the excitation of afferent nerve fibres by epidural stimulation in deeply anaesthetized rats in vivo. Riluzole did not prevent the induction of the polarization-evoked sustained increase in the excitability of dorsal column fibres but tended to weaken it. It likewise weakened but did not abolish the sustained polarization-evoked shortening of the refractory period of these fibres. These results lead to the conclusion that the persistent sodium current may contribute to the sustained post-polarization-evoked effects but is only partly involved in both the induction and the expression of these effects., (© 2023 The Authors. European Journal of Neuroscience published by Federation of European Neuroscience Societies and John Wiley & Sons Ltd.)

Dysregulated neurite outgrowth and synapse formation underlie many psychiatric disorders, which are also manifested by wolfram syndrome (WS). Whether and how the causative gene WFS1 deficiency affects synapse formation remain elusive. By mirroring human brain development with cerebral organoids, WFS1-deficient cerebral organoids not only recapitulate the neuronal loss in WS patients, but also exhibit significantly impaired synapse formation and function associated with reduced astrocytes. WFS1 deficiency in neurons autonomously delays neuronal differentiation with altered expressions of genes associated with psychiatric disorders, and impairs neurite outgrowth and synapse formation with elevated cytosolic calcium. Intriguingly, WFS1 deficiency in astrocytes decreases the expression of glutamate transporter EAAT2 by NF-κB activation and induces excessive glutamate. When co-cultured with wildtype neurons, WFS1-deficient astrocytes lead to impaired neurite outgrowth and increased cytosolic calcium in neurons. Importantly, disrupted synapse formation and function in WFS1-deficient cerebral organoids and impaired neurite outgrowth affected by WFS1-deficient astrocytes are efficiently reversed with Riluzole treatment, by restoring EAAT2 expression in astrocytes. Furthermore, Riluzole rescues the depressive-like behavior in the forced swimming test and the impaired recognition and spatial memory in the novel object test and water maze test in Wfs1 conditional knockout mice. Altogether, our study provides novel insights into how WFS1 deficiency affects synapse formation and function, and offers a strategy to treat this disease., (© 2023. The Author(s).)

Cancer treatment requires the participation of multiple targets/pathways, and single approach is hard to effectively curb the proliferation and metastasis of carcinoma cells. In this work, we conjugated FDA-approved riluzole and platinum(II) drugs into a series of unreported riluzole-Pt(IV) compounds, which were designed to simultaneously target DNA, the solute carrier family 7 member 11 (SLC7A11, xCT), and the human ether a go-go related gene 1 (hERG1), to exert synergistic anticancer effect. Among them, c,c,t-[PtCl 2 (NH 3 ) 2 (OH)(glutarylriluzole)] (compound 2) displayed excellent antiproliferative activity with IC 50 value of 300-times lower than that of cisplatin in HCT-116, and optimal selectivity index between carcinoma and human normal liver cells (LO2). Mechanism studies indicated that compound 2 released riluzole and active Pt(II) species after entering cells to exhibit a prodrug behavior against cancer, which obviously increased DNA-damage and cell apoptosis, as well as suppressed metastasis in HCT-116. Compound 2 persisted in the xCT-target of riluzole and blocked the biosynthesis of glutathione (GSH) to trigger oxidative stress, which could boost the killing to cancer cells and reduce Pt-drug resistance. Meanwhile, compound 2 significantly inhibited invasion and metastasis of HCT-116 cells by targeting hERG1 to interrupt the phosphorylation of phosphatidylinositide 3-kinases/proteinserine-threonine kinase (PI3K/Akt), and reverse epithelial-mesenchymal transformation (EMT). Based on our results, the riluzole-Pt(IV) prodrugs studied in this work could be regarded as a new class of very promising candidates for cancer treatment compared to traditional platinum drugs., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Masson SAS. All rights reserved.)

Current medications for Alzheimer's disease help manage symptoms and behavioral problems. Nevertheless, they do not slow the progression of cognitive decline or dementia. A potential approach for treating Alzheimer's disease is to target neurons that are sensitive to disease pathobiology such as glutamatergic neurons. Several patents disclosed methods for treating Alzheimer's disease by administering riluzole or its prodrugs. Clinical trials revealed that 6 months treatment using riluzole or troriluzole is associated with a slower decline in the tomographic measures of the positron emissions of cerebral glucose metabolism in Alzheimer's patients. The proposed strategy claims to prevent and/or slow the cognitive decline of Alzheimer's patients and to enhance global functioning. These claims may also pave the way for other glutamate modulators to be used for Alzheimer's disease.

Epileptogenic seizures, or status epilepticus (SE), leads to excitotoxic injury in hippocampal and limbic neurons in the kainic acid (KA) animal model of temporal lobe epilepsy (TLE). Here, we have further characterized neural activity regulated methylaminoisobutryic acid (MeAIB)/glutamine transport activity in mature rat hippocampal neurons in vitro that is inhibited by riluzole (IC 50  = 1 μM), an anti-convulsant benzothiazole agent. We screened a library of riluzole derivatives and identified SKA-41 followed by a second screen and synthesized several novel chlorinated aminothiazoles (SKA-377, SKA-378, SKA-379) that are also potent MeAIB transport inhibitors in vitro, and brain penetrant following systemic administration. When administered before KA, SKA-378 did not prevent seizures but still protected the hippocampus and several other limbic areas against SE-induced neurodegeneration at 3d. When SKA-377 - 379, (30 mg/kg) were administered after KA-induced SE, acute neural injury in the CA3, CA1 and CA4/hilus was also largely attenuated. Riluzole (10 mg/kg) blocks acute neural injury. Kinetic analysis of SKA-378 and riluzoles' blockade of Ca 2+ -regulated MeAIB transport in neurons in vitro indicates that inhibition occurs via a non-competitive, indirect mechanism. Sodium channel Na V 1.6 antagonism blocks neural activity regulated MeAIB/Gln transport in vitro (IC 50  = 60 nM) and SKA-378 is the most potent inhibitor of Na V 1.6 (IC 50  = 28 μM) compared to Na V 1.2 (IC 50  = 118 μM) in heterologous cells. However, pharmacokinetic analysis suggests that sodium channel blockade may not be the predominant mechanism of neuroprotection here. Riluzole and our novel aminothiazoles are agents that attenuate acute neural hippocampal injury following KA-induced SE and may help to understand mechanisms involved in the progression of epileptic disease., Competing Interests: Declaration of competing interest The authors have no conflict of interests., (Copyright © 2022 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Tetrodotoxin (TTX) potently inhibits TTX-sensitive voltage-gated sodium (Na V ) channels in nerve and muscle cells, potentially resulting in depressed neurotransmission, paralysis and death from respiratory failure. Since a wide range of pharmaceutical drugs is known to also act on Na V channels, the use of medicines could predispose individuals to a higher susceptibility towards TTX toxicity. We therefore first assessed the inhibitory effect of selected medicines that act on TTX-sensitive (Riluzole, Chloroquine, Fluoxetine, Valproic acid, Lamotrigine, Lidocaine) and TTX-resistant (Carbamazepine, Mexiletine, Flecainide) Na V channels on spontaneous neuronal activity of rat primary cortical cultures grown on microelectrode arrays (MEA). After establishing concentration-effect curves, binary mixtures of the medicines with TTX at calculated NOEC, IC 20 and IC 50 values were used to determine if pharmacodynamic interactions occur between TTX and these drugs on spontaneous neuronal activity. At IC 20 and IC 50 values, all medicines significantly increased the inhibitory effect of TTX on spontaneous neuronal activity of rat cortical cells in vitro. Subsequent experiments using human iPSC-derived neuronal co-cultures grown on MEAs confirmed the ability of selected medicines (Carbamazepine, Flecainide, Riluzole, Lidocaine) to inhibit spontaneous neuronal activity. Despite the need for additional experiments using human iPSC-derived neuronal co-cultures, our combined data already highlight the importance of identifying and including vulnerable risk groups in the risk assessment of TTX., Competing Interests: Declaration of Competing Interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Remco Westerink reports financial support was provided by NVWA., (Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved.)

The present study was initiated with PDB selection and validation where 11 acetylcholinesterase (AChE) and 4 N-methyl-D-aspartate receptor (NMDAR) proteins were considered for docking with Rivastigmine and Riluzole respectively. Out of the 15 proteins, selected significant binding was observed for AChE, with 5FPQ, and NMDA receptors with 5I2K. Molecular docking studies of 5FPQ/Rivastigmine complex displayed a binding score of -8.6 kcal/mol, and the predicted inhibitory concentration (Ki) was found to be 31 nM, whereas the 5I2K/Riluzole complex showed a binding score of -9.6 kcal/mol, with an inhibitory concentration (Ki) of 21 nM. Riluzole in complex with 5I2K formed predominant π-π stacking interactions with Tyr144, pi-alkyl interaction with Pro129, and conventional hydrogen bond with Phe130. In contrast, Rivastigmine in a complex with 5FPQ formed a hydrogen bond with Gln413 and pi-alkyl with Pro537. Molecular dynamics simulation study of both complexes 5FPQ/Rivastigmine and 5I2K/Riluzole exhibited stable RMSD, RMSF, Rg, and significant numbers of hydrogen bonds. From free energy landscape (FEL) analysis both complexes were observed to achieve global minima. Overall, molecular docking and MD simulation with subsequent binding free energies studies (MM-PBSA) elucidate the binding conformations and stability of these reprogrammed drugs in the AChE and NMDAR targets. From these in-silico predictions, it can be suggested that both Rivastigmine and Riluzole combination may provide better insights as a starting point combination therapy for the treatment of Alzheimer's disease.Communicated by Ramaswamy H. Sarma.

Background: Prior research supports a strong link between Alzheimer's disease (AD) and metabolic dysfunction that involves a multi-directional interaction between glucose, glutamatergic homeostasis, and amyloid pathology. Elevated soluble amyloid-β (Aβ) is an early biomarker for AD-associated cognitive decline that contributes to concurrent glutamatergic and metabolic dyshomeostasis in humans and male transgenic AD mice. Yet, it remains unclear how primary time-sensitive targeting of hippocampal glutamatergic activity may impact glucose regulation in an amyloidogenic mouse model. Previous studies have illustrated increased glucose uptake and metabolism using a neuroprotective glutamate modulator (riluzole), supporting the link between glucose and glutamatergic homeostasis., Objective: We hypothesized that targeting early glutamatergic hyperexcitation through riluzole treatment could aid in attenuating co-occurring metabolic and amyloidogenic pathologies with the intent of ameliorating cognitive decline., Methods: We conducted an early intervention study in male and female transgenic (AβPP/PS1) and knock-in (APPNL - F/NL - F) AD mice to assess the on- and off-treatment effects of prodromal glutamatergic modulation (2-6 months of age) on glucose homeostasis and spatial cognition through riluzole treatment., Results: Results indicated a sex- and genotype-specific effect on glucose homeostasis and spatial cognition with riluzole intervention that evolved with disease progression and time since treatment., Conclusion: These findings support the interconnected nature of glucose and glutamatergic homeostasis with amyloid pathology and petition for further investigation into the targeting of this relationship to improve cognitive performance.

Amyotrophic lateral sclerosis (ALS) is a rare, cumulative neurological deteriorating disease that disturbs the neurons (nerve cells) that control voluntary muscle movement (those muscles we choose to move). Currently, the Food and Drug Administration (FDA) approved drugs such as Radicava, Rilutek, Tiglutik, Exservan, and Nuedexta to treat ALS. Given the wide range of pharmaceutical applications of heterocyclic compounds, especially those containing the nitrogen ring systems such as pyridine, pyrimidine, and indole. These molecular frameworks have piqued the interest of medicinal chemists for further investigation in a variety of diseases. We have found several review works done on this research topic. Until now, no reviews published on the nitrogen heterocycles for treating ALS. This review examines the major causes of ALS, a brief history of medications that have been used to treat it so far, and the most recent breakthroughs in nitrogen ring systems for treating ALS. The novelty of this study provides insights on several effective synthetic techniques for nitrogen-based heterocyclic medications that operate as potent anti-inflammatory treatments and guard against ALS. KEY POINTS: • Pharmacological activity of heterocyclic compounds. • Neurodegenerative diseases and their drawbacks are discussed in detail. • Recent survey of nitrogen heterocycles in treating ALS disease are highlighted., (© 2022. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Riluzole (RLZ) is a neuroprotective drug indicated for amyotrophic lateral sclerosis. To examine the feasibility of RLZ for repositioning as an anti-inflammatory bowel disease (IBD) drug, RLZ (2, 5, and 10 mg/kg) was administered orally to rats with colitis induced by 2,4-dinitrobenzenesulfonic acid. Oral RLZ was effective against rat colitis in a dose-dependent manner, which was statistically significant at doses over 5 mg/kg. To address safety issues upon repositioning and further improve anti-colitic effectiveness, RLZ was coupled with salicylic acid (SA) via an azo-bond to yield RLZ-azo-SA (RAS) for the targeted colonic delivery of RLZ. Upon oral gavage, RAS (oral RAS) was efficiently delivered to and activated to RLZ in the large intestine, and systemic absorption of RLZ was substantially reduced. Oral RAS ameliorated colonic damage and inflammation in rat colitis and was more effective than oral RLZ and sulfasalazine, a current anti-IBD drug. Moreover, oral RAS potently inhibited glycogen synthase kinase 3β (GSK3β) in the inflamed distal colon, leading to the suppression of NFκB activity and an increase in the level of the anti-inflammatory cytokine interleukin-10. Taken together, RAS, which enables RLZ to be delivered to and inhibit GSK3β in the inflamed colon, may facilitate repositioning of RLZ as an anti-IBD drug.

Polypharmacology is a new trend in amyotrophic lateral sclerosis (ALS) therapy and an effective way of addressing a multifactorial etiology involving excitotoxicity, mitochondrial dysfunction, oxidative stress, and microglial activation. Inspired by a reported clinical trial, we converted a riluzole ( 1 )-rasagiline ( 2 ) combination into single-molecule multi-target-directed ligands. By a ligand-based approach, the highly structurally integrated hybrids 3 - 8 were designed and synthesized. Through a target- and phenotypic-based screening pipeline, we identified hit compound 6 . It showed monoamine oxidase A (MAO-A) inhibitory activity (IC 50 = 6.9 μM) rationalized by in silico studies as well as in vitro brain permeability. By using neuronal and non-neuronal cell models, including ALS-patient-derived cells, we disclosed for 6 a neuroprotective/neuroinflammatory profile similar to that of the parent compounds and their combination. Furthermore, the unexpected MAO inhibitory activity of 1 (IC 50 = 8.7 μM) might add a piece to the puzzle of its anti-ALS molecular profile.

Cadmium (Cd) damages plant photosynthesis, affects roots and leaves growth, and triggers molecular responses. Riluzole (RIL), which protected neuronal damage via inhibiting excess Glu release in animals, has been found to improve Cd tolerance in duckweed in this study. Firstly, RIL treatment alleviated leaf chlorosis by protecting chlorophyll and decreased root abscission under Cd stress. Secondly, RIL declines Cd accumulation by alleviating excess Glu release during Cd shock. RIL mitigate Glu outburst in duckweed during Cd stress by a decline in Glu in roots. The Cd 2+ influx was repressed by RIL addition with Cd shock. Finally, differentially expressed genes (DEGs) of duckweed under Cd stress with RIL have been investigated. 2141 genes were substantially up-regulated and 3282 genes were substantially down-regulated with RIL addition. RIL down-regulates the genes related to the Glu synthesis, and genes related to DNA repair have been up-regulated with RIL treatment under Cd stress. These results provide new insights into the possibility of RIL to reduce Cd accumulation and increase Cd tolerance in duckweed, and lay the foundation for decreasing Cd accumulation in crops., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

Most cancer cells rely on aerobic glycolysis to support uncontrolled proliferation and evade apoptosis. However, pancreatic cancer cells switch to glutamine metabolism to survive under hypoxic conditions. Activation of the Wnt/β-catenin pathway induces aerobic glycolysis by activating enzymes required for glucose metabolism and regulating the expression of glutamate transporter and glutamine synthetase. The results demonstrate that riluzole inhibits pancreatic cancer cell growth and has no effect on human pancreatic normal ductal epithelial cells. RNA-seq experiments identified the involvement of Wnt and metabolic pathways by riluzole. Inhibition of Wnt-β-catenin/TCF-LEF pathway by riluzole suppresses the expression of PDK, MCT1, cMyc, AXIN, and CyclinD1. Riluzole inhibits glucose transporter 2 expression, glucose uptake, lactate dehydrogenase A expression, and NAD + level. Furthermore, riluzole inhibits glutamate release and glutathione levels, and elevates reactive oxygen species. Riluzole disrupts mitochondrial homeostasis by inhibiting Bcl-2 and upregulating Bax expression, resulting in a drop of mitochondrial membrane potential. Finally, riluzole inhibits pancreatic cancer growth in KPC (Pdx1-Cre, LSL-Trp53 R172H , and LSL-Kras G12D ) mice. In conclusion, riluzole can inhibit pancreatic cancer growth by regulating glucose and glutamine metabolisms and can be used to treat pancreatic cancer., (© 2022. The Author(s).)

Alzheimer's disease (AD) is a neurodegenerative disorder leading to memory loss and impaired cognition. Despite several decades of research, AD therapeutic is not available. In this study, we have investigated the impact of a chronic intervention of riluzole on memory and neurometabolism in the AβPP-PS1 mouse model of AD. The 10-month-old AβPP-PS1 mice were administered 30 doses of riluzole (6 mg/kg, intragastrically) on an alternate day for two months. The memory was assessed using Morris Water Maze, while neurometabolism was evaluated by 1 H-[ 13 C]-NMR spectroscopy together with an intravenous infusion of [1,6- 13 C 2 ]glucose. The normal saline-treated AβPP-PS1 mice exhibited a decrease in learning and memory that were restored to the control level following riluzole treatment. Most interestingly, the reduced 13 C labeling of Glu C4 and Asp C3 from [1,6- 13 C]glucose in the AβPP-PS1 mice was restored to the control level following riluzole intervention. As a consequence, chronic riluzole treatment improved metabolic activity of glutamatergic neurons in AβPP-PS1 mice. Together these data suggest that riluzole may be useful for improving cognition in AD., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Neuropathic pain since early diabetes swamps patients' lives, and diabetes mellitus has become an increasingly worldwide epidemic. No agent, so far, can terminate the ongoing diabetes. Therefore, strategies that delay the process and the further complications are preferred, such as diabetic neuropathic pain (DNP). Dysfunction of ion channels is generally accepted as the central mechanism of diabetic associated neuropathy, of which hyperpolarization-activated cyclic nucleotide-gated 2 (HCN2) ion channel has been verified the involvement of neuropathic pain in dorsal root ganglion (DRG) neurons. Riluzole is a benzothiazole compound with neuroprotective properties on intervention to various ion channels, including hyperpolarization-activated voltage-dependent channels. To investigate the effect of riluzole within lumbar (L3-5) DRG neurons from DNP models, streptozocin (STZ, 70 mg/kg) injection was recruited subcutaneously followed by paw withdrawal mechanical threshold (PWMT) and paw withdrawal thermal latency (PWTL), which both show significant reduction, whilst relieved by riluzole (4 mg/kg/d) administration, which was performed once daily for 7 consecutive days for 14 days. HCN2 expression was also decreased in line with alleviated behavioral tests. Our results indicate riluzole as the alleviator to STZ-induced DNP with involvement of downregulated HCN2 in lumbar DRG by continual systemic administration in rats., Competing Interests: The authors declare that they have no conflicts of interest., (Copyright © 2022 Dong-mei Zhou et al.)

After spinal cord injury (SCI), emergency treatment intervention can minimize tissue damage, which is closely related to the recovery of long-term function. Here, we examined whether the administration of a single dose of riluzole (6 mg/kg) immediately after SCI was a critical window for the drug to exert its regulatory effect and limit long-term neurological deficits. The animals were sacrificed 1 day after administration for investigation of neuronal survival and a potential neuroinflammatory response, and sacrificed in the 6th week for assessment of neurological function. Riluzole applied in a single dose immediately post-SCI decreased the mRNA level of interleukin-1β at 6 h, reduced the destruction of neurons, and reduced the activation of microglia/macrophage M1 expression at day 1 post-SCI. Additionally, riluzole-treated rats showed higher expressions of interleukin-33 and its receptor ST2 in microglia/macrophages of the spinal cord than vehicle-treated rats, suggesting that this signaling pathway might be involved in microglia/macrophage-mediated inflammation. At 6 weeks, riluzole-treated rats exhibited higher motor function scores than vehicle-treated controls. In addition, riluzole-treated rats exhibited higher expression of GAP43 protein and shorter N1 peak latency and larger N1-P1 amplitude in motor-evoked potentials, compared to vehicle-treated rats. Together, these data suggested that early application of riluzole after SCI could be crucial for long-term functional recovery, so it may represent a promising therapeutic candidate within the critical therapeutic window for acute SCI., (© 2021. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Even though amyotrophic lateral sclerosis (ALS) is a disease of the upper and lower motor neurons, to date none of the compounds in clinical trials have been tested for improving the health of diseased upper motor neurons (UMNs). There is an urgent need to develop preclinical assays that include UMN health as a readout. Since ALS is a complex disease, combinatorial treatment strategies will be required to address the mechanisms perturbed in patients. Here, we describe a novel in vitro platform that takes advantage of an UMN reporter line in which UMNs are genetically labeled with fluorescence and have misfolded SOD1 toxicity. We report that NU-9, an analog of the cyclohexane-1,3-dione family of compounds, improves the health of UMNs with misfolded SOD1 toxicity more effectively than riluzole or edaravone, -the only two FDA-approved ALS drugs to date-. Interestingly, when NU-9 is applied in combination with riluzole or edaravone, there is an additive effect on UMN health, as they extend longer axons and display enhanced branching and arborization, two important characteristics of healthy UMNs in vitro., (© 2022. The Author(s).)

Amyotrophic Lateral Sclerosis is a neurodegenerative disease caused by progressive loss of motor neurons, which severely compromises skeletal muscle function. Evidence shows that muscle may act as a molecular powerhouse, whose final signals generate in patients a progressive loss of voluntary muscle function and weakness leading to paralysis. This pathology is the result of a complex cascade of events that involves a crosstalk among motor neurons, glia, and muscles, and evolves through the action of converging toxic mechanisms. In fact, mitochondrial dysfunction, which leads to oxidative stress, is one of the mechanisms causing cell death. It is a common denominator for the two existing forms of the disease: sporadic and familial. Other factors include excitotoxicity, inflammation, and protein aggregation. Currently, there are limited cures. The only approved drug for therapy is riluzole, that modestly prolongs survival, with edaravone now waiting for new clinical trial aimed to clarify its efficacy. Thus, there is a need of effective treatments to reverse the damage in this devastating pathology. Many drugs have been already tested in clinical trials and are currently under investigation. This review summarizes the already tested drugs aimed at restoring muscle-nerve cross-talk and on new treatment options targeting this tissue.

Spinal cord injury (SCI) is a disastrous event that often leads to permanent neurological deficits involving motor, sensory, and autonomic dysfunctions in patients. Accumulating research has demonstrated that riluzole may play crucial roles in the process of spinal tissue repair, but the underlying mechanisms remain elusive. This study verified the effectiveness of riluzole and speculated that a riluzole-afforded protection mechanism may be associated with the glycogen synthase kinase-3 beta (GSK-3β)/collapsin response mediator protein-2 (CRMP-2) pathway in rats after spinal cord injury. Here, a modified Allen's weight dropping model was generated and riluzole at 4 mg/kg was injected intraperitoneally after surgery and twice a day for 7 consecutive days. At 6 weeks after SCI, we found that riluzole treatment reduced the central cavity size of the spinal cord and improved neurological functions. Meanwhile, riluzole-treated rats exhibited shorter latency and larger amplitude in motor evoked potentials and somatosensory evoked potentials, compared with vehicle-treated rats. Furthermore, Western blotting and immunofluorescence data revealed that the expression levels of GSK-3β and phosphorylated-GSK-3β were lower in riluzole-treated SCI rats compared with vehicle-treated rats. We next detected the expression CRMP-2 and phosphorylated CRMP-2 and found that the expression of CRMP-2 showed no difference between the riluzole-treated and vehicle-treated groups; however, administration of riluzole downregulated phosphorylated CRMP-2 expression. The current findings suggest that after SCI, administration of riluzole promotes neurological functional restoration, which may be associated, in part, with its activation of the GSK-3β/CRMP-2 signaling pathway.

Acute intoxication with tetramethylenedisulfotetramine (TETS) can trigger status epilepticus (SE) in humans. Survivors often exhibit long-term neurological effects, including electrographic abnormalities and cognitive deficits, but the pathogenic mechanisms linking the acute toxic effects of TETS to chronic outcomes are not known. Here, we use advanced in vivo imaging techniques to longitudinally monitor the neuropathological consequences of TETS-induced SE in two different mouse strains. Adult male NIH Swiss and C57BL/6J mice were injected with riluzole (10 mg/kg, i.p.), followed 10 min later by an acute dose of TETS (0.2 mg/kg in NIH Swiss; 0.3 mg/kg, i.p. in C57BL/6J) or an equal volume of vehicle (10% DMSO in 0.9% sterile saline). Different TETS doses were administered to trigger comparable seizure behavior between strains. Seizure behavior began within minutes of TETS exposure and rapidly progressed to SE that was terminated after 40 min by administration of midazolam (1.8 mg/kg, i.m.). The brains of vehicle and TETS-exposed mice were imaged using in vivo magnetic resonance (MR) and translocator protein (TSPO) positron emission tomography (PET) at 1, 3, 7, and 14 days post-exposure to monitor brain injury and neuroinflammation, respectively. When the brain scans of TETS mice were compared to those of vehicle controls, subtle and transient neuropathology was observed in both mouse strains, but more extensive and persistent TETS-induced neuropathology was observed in C57BL/6J mice. In addition, one NIH Swiss TETS mouse that did not respond to the midazolam therapy, but remained in SE for more than 2 h, displayed robust neuropathology as determined by in vivo imaging and confirmed by FluoroJade C staining and IBA-1 immunohistochemistry as readouts of neurodegeneration and neuroinflammation, respectively. These findings demonstrate that the extent of injury observed in the mouse brain after TETS-induced SE varied according to strain, dose of TETS and/or the duration of SE. These observations suggest that TETS-intoxicated humans who do not respond to antiseizure medication are at increased risk for brain injury., (Copyright © 2021 The Author(s). Published by Elsevier B.V. All rights reserved.)

Riluzole, a glutamate release inhibitor, has been in use for the treatment of amyotrophic lateral sclerosis for over two decades since its approval by the Food and Drug Administration. Recently, riluzole has been evaluated in cancer cells and indicated to block cell proliferation and/or induce cell death. Riluzole has been proven effective as an anti‑neoplastic drug in cancers of various tissue origins, including the skin, breast, pancreas, colon, liver, bone, brain, lung and nasopharynx. While cancer cells expressing glutamate receptors frequently respond to riluzole treatment, numerous types of cancer cell lacking glutamate receptors unexpectedly responded to riluzole treatment as well. Riluzole was demonstrated to interfere with glutamate secretion, growth signaling pathways, Ca 2+ homeostasis, glutathione synthesis, reactive oxygen species generation and integrity of DNA, as well as autophagic and apoptotic pathways. Of note, riluzole is highly effective in inducing cell death in cisplatin‑resistant lung cancer cells. Furthermore, riluzole pretreatment sensitizes glioma and melanoma to radiation therapy. In addition, in triple‑negative breast cancer, colorectal cancer, melanoma and glioblastoma, riluzole has synergistic effects in combination with select drugs. In an effort to highlight the therapeutic potential of riluzole, the current study reviewed the effect and outcome of riluzole treatment on numerous cancer types investigated thus far. The mechanism of action and the various molecular pathways affected by riluzole are discussed.

Our lab has previously demonstrated Riluzole to be an effective drug in inhibiting proliferation and inducing apoptosis in both human and mouse osteosarcoma. Yes-associated protein is a transcription co-activator, known to be involved in cell proliferation or apoptosis depending on its protein partner. In the present study we investigated the role of YAP in apoptosis in osteosarcoma, we hypothesized that YAP may be activated by Riluzole to induce apoptosis in osteosarcoma. By knocking down the expression of YAP, we have demonstrated that Riluzole failed to induce apoptosis in YAP deficient osteosarcoma cells. Riluzole caused translocation of YAP from the cytoplasm to the nucleus, indicating YAP's role in apoptosis. Both Riluzole-induced phosphorylation of YAP at tyrosine 357 and Riluzole-induced apoptosis were blocked by inhibitors of c-Abl kinase. In addition, knockdown of c-Abl kinase prevented Riluzole-induced apoptosis in LM7 cells. We further demonstrated that Riluzole promoted interaction between YAP and p73, while c-Abl kinase inhibitors abolished the interaction. Subsequently, we demonstrated that Riluzole enhanced activity of the Bax promoter in a luciferase reporter assay and enhanced YAP/p73 binding on endogenous Bax promoter in a ChIP assay. Our data supports a novel mechanism in which Riluzole activates c-Abl kinase to regulate pro-apoptotic activity of YAP in osteosarcoma., (© 2021. The Author(s).)

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease commonly treated with riluzole, a small molecule that may act via modulation of glutamatergic neurotransmission. However, riluzole only modestly extends lifespan for people living with ALS, and its precise mechanisms of action remain unclear. Most ALS cases are characterised by accumulation of cytoplasmic TAR DNA binding protein of 43 kDa (TDP-43), and understanding the effects of riluzole in models that closely recapitulate TDP-43 pathology may provide insights for development of improved therapeutics. We therefore investigated the effects of riluzole in female transgenic mice that inducibly express nuclear localisation sequence (NLS)-deficient human TDP-43 in neurons (NEFH-tTA/tetO-hTDP-43ΔNLS, 'rNLS8', mice). Riluzole treatment from the first day of hTDP-43ΔNLS expression did not alter disease onset, weight loss or performance on multiple motor behavioural tasks. Riluzole treatment also did not alter TDP-43 protein levels, solubility or phosphorylation. Although we identified a significant decrease in GluA2 and GluA3 proteins in the cortex of rNLS8 mice, riluzole did not ameliorate this disease-associated molecular phenotype. Likewise, riluzole did not alter the disease-associated atrophy of hindlimb muscle in rNLS8 mice. Finally, riluzole treatment beginning after disease onset in rNLS8 mice similarly had no effect on progression of late-stage disease or animal survival. Together, we demonstrate specific glutamatergic receptor alterations and muscle fibre-type changes reminiscent of ALS in female rNLS8 mice, but riluzole had no effect on these or any other disease phenotypes. Future targeting of pathways related to accumulation of TDP-43 pathology may be needed to develop better treatments for ALS., (© 2021 Federation of European Neuroscience Societies and John Wiley & Sons Ltd.)

Pancreatic cancer is always diagnosed at an advanced stage. Hence, chemotherapy becomes the best choice for patients. Therefore, new anticancer drugs for pancreatic cancer are needed. Riluzole (RIL) is mainly used to treat amyotrophic lateral sclerosis clinically, but many previous studies have shown that RIL could inhibit tumors. However, no report has explored the association between RIL and pancreatic cancer. To validate this association, we performed this study. Our data showed that RIL could induce cytotoxicity, block the cell cycle, and inhibit clone formation, apoptosis, and migration in pancreatic cancer cells. Moreover, we demonstrated that RIL could suppress autophagy. However, more experiments will be needed to validate the reliability of our conclusions. In summary, our data suggest that RIL might provide clues for the development of a treatment for human pancreatic cancer in the future., (© 2019 Wiley Periodicals, Inc.)

Chronic sensitization to serotonin 1A and 7 receptors agonist 8-OH-DPAT induces compulsive checking and perseverative behavior. As such, it has been used to model obsessive-compulsive disorder (OCD)-like behavior in mice and rats. In this study, we tested spatial learning in the 8-OH-DPAT model of OCD and the effect of co-administration of memantine and riluzole-glutamate-modulating agents that have been shown to be effective in several clinical trials. Rats were tested in the active place avoidance task in the Carousel maze, where they learned to avoid the visually imperceptible shock sector. All rats were subcutaneously injected with 8-OH-DPAT (0.25 mg/kg) or saline (control group) during habituation. During acquisition, they were pretreated with riluzole (1 mg/kg), memantine (1 mg/kg), or saline solution 30 min before each session and injected with 8-OH-DPAT ("OH" groups) or saline ("saline" groups) right before the experiment. We found that repeated application of 8-OH-DPAT during both habituation and acquisition significantly increased locomotion, but it impaired the ability to avoid the shock sector. However, the application of 8-OH-DPAT in habituation had no impact on the learning process if discontinued in acquisition. Similarly, memantine and riluzole did not affect the measured parameters in the "saline" groups, but in the "OH" groups, they significantly increased locomotion. In addition, riluzole increased the number of entrances and decreased the maximum time avoided of the shock sector. We conclude that monotherapy with glutamate-modulating agents does not reduce but exacerbates cognitive symptoms in the animal model of OCD.

Dyskinesias are characterized by abnormal repetitive involuntary movements due to dysfunctional neuronal activity. Although levodopa-induced dyskinesia, characterized by tic-like abnormal involuntary movements, has no clinical treatment for Parkinson's disease patients, animal studies indicate that Riluzole, which interferes with glutamatergic neurotransmission, can improve the phenotype. The rat model of Levodopa-Induced Dyskinesia is a unilateral lesion with 6-hydroxydopamine in the medial forebrain bundle, followed by the repeated administration of levodopa. The molecular pathomechanism of Levodopa-Induced Dyskinesia is still not deciphered; however, the implication of epigenetic mechanisms was suggested. In this study, we investigated the striatum for DNA methylation alterations under chronic levodopa treatment with or without co-treatment with Riluzole. Our data show that the lesioned and contralateral striata have nearly identical DNA methylation profiles. Chronic levodopa and levodopa + Riluzole treatments led to DNA methylation loss, particularly outside of promoters, in gene bodies and CpG poor regions. We observed that several genes involved in the Levodopa-Induced Dyskinesia underwent methylation changes. Furthermore, the Riluzole co-treatment, which improved the phenotype, pinpointed specific methylation targets, with a more than 20% methylation difference relative to levodopa treatment alone. These findings indicate potential new druggable targets for Levodopa-Induced Dyskinesia.

Soluble amyloid beta (Aβ) is believed to contribute to cognitive deficits in the early stages of Alzheimer's disease (AD). Increased soluble Aβ 1-42 in the hippocampus is closely correlated with spatial learning and memory deficits in AD. Riluzole (RLZ), an FDA-approved drug for amyotrophic lateral sclerosis (ALS), has beneficial effects for AD. However, the mechanism underlying the effects remains unclear. In this study, its neuroprotective effect against soluble Aβ 1-42 -induced spatial cognitive deficits in rats was assessed. We found that intrahippocampal injection of soluble Aβ 1-42 impaired spatial cognitive function and suppressed long-term potentiation (LTP) of the DG region, which was relevant to soluble Aβ 1-42 -induced shift of the hippocampal excitation/inhibition balance toward excitation. Interestingly, RLZ ameliorated Aβ 1-42 -induced behavioral and LTP impairments through rescuing the soluble Aβ 1-42 -induced excitation/inhibition imbalance. RLZ attenuated Aβ 1-42 -mediated facilitation of excitatory synaptic transmission by facilitating glutamate reuptake and decreasing presynaptic glutamate release. Meanwhile, RLZ attenuated the suppression of inhibitory synaptic transmission caused by Aβ 1-42 by potentiating postsynaptic GABA receptor function. These results suggest that RLZ exerts a neuroprotective effect against soluble Aβ 1-42 -related spatial cognitive deficits through rescuing the excitation/inhibition imbalance, and it could be a potential therapy for AD., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Riluzole is an anticonvulsant drug also used to treat the amyotrophic lateral sclerosis and major depressive disorder. This compound has antiglutamatergic activity and is an important multichannel blocker. However, little is known about its actions on the Kv4.2 channels, the molecular correlate of the A-type K + current (I A ) and the fast transient outward current (I tof ). Here, we investigated the effects of riluzole on Kv4.2 channels transiently expressed in HEK-293 cells. Riluzole inhibited Kv4.2 channels with an IC 50 of 190 ± 14 μM and the effect was voltage- and frequency-independent. The activation rate of the current (at +50 mV) was not affected by the drug, nor the voltage dependence of channel activation, but the inactivation rate was accelerated by 100 and 300 μM riluzole. When Kv4.2 channels were maintained at the closed state, riluzole incubation induced a tonic current inhibition. In addition, riluzole significantly shifted the voltage dependence of inactivation to hyperpolarized potentials without affecting the recovery from inactivation. In the presence of the drug, the closed-state inactivation was significantly accelerated, and the percentage of inactivated channels was increased. Altogether, our findings indicate that riluzole inhibits Kv4.2 channels mainly affecting the closed and closed-inactivated states., (Copyright © 2021 Elsevier B.V. All rights reserved.)

This paper provides evidence to support that riluzole, an FDA-approved treatment for amyotrophic lateral sclerosis (ALS), like many neuroprotective agents, displays and exerts hormetic biphasic dose responses. These findings have important implications for the experimental study and clinical treatment of ALS., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Background and Purpose: Sodium channel inhibitors can be used to treat hyperexcitability-related diseases, including epilepsies, pain syndromes, neuromuscular disorders and cardiac arrhythmias. The applicability of these drugs is limited by their nonspecific effect on physiological function. They act mainly by sodium channel block and in addition by modulation of channel kinetics. While channel block inhibits healthy and pathological tissue equally, modulation can preferentially inhibit pathological activity. An ideal drug designed to target the sodium channels of pathological tissue would act predominantly by modulation. Thus far, no such drug has been described., Experimental Approach: Patch-clamp experiments with ultra-fast solution exchange and photolabeling-coupled electrophysiology were applied to describe the unique mechanism of riluzole on Nav1.4 sodium channels. In silico docking experiments were used to study the molecular details of binding., Key Results: We present evidence that riluzole acts predominantly by non-blocking modulation. We propose that, being a relatively small molecule, riluzole is able to stay bound to the binding site, but nonetheless stay off the conduction pathway, by residing in one of the fenestrations. We demonstrate how this mechanism can be recognized., Conclusions and Implications: Our results identify riluzole as the prototype of this new class of sodium channel inhibitors. Drugs of this class are expected to selectively prevent hyperexcitability, while having minimal effect on cells firing at a normal rate from a normal resting potential., (© 2021 The Authors. British Journal of Pharmacology published by John Wiley & Sons Ltd on behalf of British Pharmacological Society.)