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Nanomedicine : Nanotechnology, Biology,... Jul 2024Fisetin has displayed potential as an anticonvulsant in preclinical studies yet lacks clinical validation. Challenges like low solubility and rapid metabolism may limit...
Fisetin-loaded chitosan nanoparticles ameliorate pilocarpine-induced temporal lobe epilepsy and associated neurobehavioral alterations in mice: Role of ROS/TNF-α-NLRP3 inflammasomes pathway.
Fisetin has displayed potential as an anticonvulsant in preclinical studies yet lacks clinical validation. Challenges like low solubility and rapid metabolism may limit its efficacy. This study explores fisetin-loaded chitosan nanoparticles (NP) to address these issues. Using a murine model of pilocarpine-induced temporal lobe epilepsy, we evaluated the anticonvulsant and neuroprotective effects of fisetin NP. Pilocarpine-induced seizures and associated neurobehavioral deficits were assessed after administering subtherapeutic doses of free fisetin and fisetin NP. Changes in ROS, inflammatory cytokines, and NLRP3/IL-18 expression in different brain regions were estimated. The results demonstrate that the fisetin NP exerts protection against seizures and associated depression-like behavior and memory impairment. Furthermore, biochemical, and histological examinations supported behavioral findings suggesting attenuation of ROS/TNF-α-NLRP3 inflammasome pathway as a neuroprotective mechanism of fisetin NP. These findings highlight the improved pharmacodynamics of fisetin using fisetin NP against epilepsy, suggesting a promising therapeutic approach against epilepsy and associated behavioral deficits.
Topics: Animals; Epilepsy, Temporal Lobe; Chitosan; Flavonols; Mice; NLR Family, Pyrin Domain-Containing 3 Protein; Inflammasomes; Pilocarpine; Nanoparticles; Male; Reactive Oxygen Species; Tumor Necrosis Factor-alpha; Flavonoids; Behavior, Animal; Anticonvulsants; Neuroprotective Agents
PubMed: 38740358
DOI: 10.1016/j.nano.2024.102752 -
Molecular and Cellular Neurosciences Jun 2024Muscarinic neurotransmission is fundamentally involved in supporting several brain functions by modulating flow of information in brain neural circuits including the...
Muscarinic neurotransmission is fundamentally involved in supporting several brain functions by modulating flow of information in brain neural circuits including the hippocampus which displays a remarkable functional segregation along its longitudinal axis. However, how muscarinic neuromodulation contributes to the functional segregation along the hippocampus remains unclear. In this study we show that the nonselective muscarinic receptor agonist carbachol similarly suppresses basal synaptic transmission in the dorsal and ventral CA1 hippocampal field, in a concentration-depended manner. Furthermore, using a ten-pulse stimulation train of varying frequency we found that carbachol changes the frequency filtering properties more in ventral than dorsal hippocampus by facilitating synaptic inputs at a wide range of input frequencies in the ventral compared with dorsal hippocampus. Using the M2 receptor antagonist gallamine and the M4 receptor antagonist tropicamide, we found that M2 receptors are involved in controlling basal synaptic transmission and short-term synaptic plasticity (STSP) in the ventral but not the dorsal hippocampus, while M4 receptors participate in modulating basal synaptic transmission and STSP in both segments of the hippocampus. These results were corroborated by the higher protein expression levels of M2 receptors in the ventral compared with dorsal hippocampus. We conclude that muscarinic transmission modulates excitatory synaptic transmission and short-term synaptic plasticity along the entire rat hippocampus by acting through M4 receptors and recruiting M2 receptors only in the ventral hippocampus. Furthermore, M4 receptors appear to exert a permissive role on the actions of M2 receptors on STSP in the ventral hippocampus. This dorsoventral differentiation of muscarinic modulation is expected to have important implications in information processing along the endogenous hippocampal circuitry.
Topics: Animals; Neuronal Plasticity; Synaptic Transmission; Rats; Hippocampus; Male; Carbachol; Receptor, Muscarinic M2; Receptors, Muscarinic; Rats, Wistar; Muscarinic Antagonists; Receptor, Muscarinic M4; Muscarinic Agonists; Excitatory Postsynaptic Potentials
PubMed: 38703973
DOI: 10.1016/j.mcn.2024.103935 -
BMJ Open Respiratory Research May 2024Methods used to assess ventilation heterogeneity through inert gas washout have been standardised and showed high sensitivity in diagnosing many respiratory diseases. We... (Observational Study)
Observational Study Comparative Study
BACKGROUND
Methods used to assess ventilation heterogeneity through inert gas washout have been standardised and showed high sensitivity in diagnosing many respiratory diseases. We hypothesised that nitrogen single or multiple breath washout tests, respectively nitrogen single breath washout (NSBW) and nitrogen multiple breath washout (NMBW), may be pathological in patients with clinical suspicion of asthma but normal spirometry. Our aim was to assess whether NSBW and NMBW are associated with methacholine challenge test (MCT) results in this population. We also postulated that an alteration in S at NSBW could be detected before the 20% fall of forced expiratory volume in the first second (FEV) in MCT.
STUDY DESIGN AND METHODS
This prospective, observational, single-centre study included patients with suspicion of asthma with normal spirometry. Patients completed questionnaires on symptoms and health-related quality-of-life and underwent the following lung function tests: NSBW (S), NMBW (Lung clearance index (LCI), S, S), MCT (FEV and sGeff) as well as NSBW between each methacholine dose.
RESULTS
182 patients were screened and 106 were included in the study, with mean age of 41.8±14 years. The majority were never-smokers (58%) and women (61%). MCT was abnormal in 48% of participants, NSBW was pathological in 10.6% at baseline and NMBW abnormality ranged widely (LCI 81%, S 18%, S 43%). The dose response rate of the MCT showed weak to moderate correlation with the subsequent NSBW measurements during the provocation phases (ρ 0.34-0.50) but no correlation with NMBW.
CONCLUSIONS
Both MCT and N washout tests are frequently pathological in patients with suspicion of asthma with normal spirometry. The weak association and lack of concordance across the tests highlight that they reflect different but not interchangeable pathological pathways of the disease.
Topics: Humans; Asthma; Methacholine Chloride; Female; Male; Prospective Studies; Adult; Spirometry; Breath Tests; Middle Aged; Nitrogen; Bronchial Provocation Tests; Forced Expiratory Volume; Respiratory Function Tests; Lung; Bronchoconstrictor Agents
PubMed: 38697675
DOI: 10.1136/bmjresp-2023-001919 -
Scientific Reports Apr 2024Selective activation of individual subtypes of muscarinic receptors is a promising way to safely alleviate a wide range of pathological conditions in the central nervous...
Selective activation of individual subtypes of muscarinic receptors is a promising way to safely alleviate a wide range of pathological conditions in the central nervous system and the periphery as well. The flexible G-protein interface of muscarinic receptors allows them to interact with several G-proteins with various efficacy, potency, and kinetics. Agonists biased to the particular G-protein mediated pathway may result in selectivity among muscarinic subtypes and, due to the non-uniform expression of individual G-protein alpha subunits, possibly achieve tissue specificity. Here, we demonstrate that novel tetrahydropyridine-based agonists exert specific signalling profiles in coupling with individual G-protein α subunits. These signalling profiles profoundly differ from the reference agonist carbachol. Moreover, coupling with individual Gα induced by these novel agonists varies among subtypes of muscarinic receptors which may lead to subtype selectivity. Thus, the novel tetrahydropyridine-based agonist can contribute to the elucidation of the mechanism of pathway-specific activation of muscarinic receptors and serve as a starting point for the development of desired selective muscarinic agonists.
Topics: Muscarinic Agonists; Receptors, Muscarinic; Animals; Signal Transduction; Humans; Pyridines; Carbachol; CHO Cells; Cricetulus; GTP-Binding Proteins; GTP-Binding Protein alpha Subunits
PubMed: 38671143
DOI: 10.1038/s41598-024-60259-4 -
Cell Reports May 2024The molecular mechanisms underlying seizure generation remain elusive, yet they are crucial for developing effective treatments for epilepsy. The current study shows...
The molecular mechanisms underlying seizure generation remain elusive, yet they are crucial for developing effective treatments for epilepsy. The current study shows that inhibiting c-Abl tyrosine kinase prevents apoptosis, reduces dendritic spine loss, and maintains N-methyl-d-aspartate (NMDA) receptor subunit 2B (NR2B) phosphorylated in in vitro models of excitotoxicity. Pilocarpine-induced status epilepticus (SE) in mice promotes c-Abl phosphorylation, and disrupting c-Abl activity leads to fewer seizures, increases latency toward SE, and improved animal survival. Currently, clinically used c-Abl inhibitors are non-selective and have poor brain penetration. The allosteric c-Abl inhibitor, neurotinib, used here has favorable potency, selectivity, pharmacokinetics, and vastly improved brain penetration. Neurotinib-administered mice have fewer seizures and improved survival following pilocarpine-SE induction. Our findings reveal c-Abl kinase activation as a key factor in ictogenesis and highlight the impact of its inhibition in preventing the insurgence of epileptic-like seizures in rodents and humans.
Topics: Animals; Male; Mice; Apoptosis; Mice, Inbred C57BL; Neurons; Phosphorylation; Pilocarpine; Protein Kinase Inhibitors; Proto-Oncogene Proteins c-abl; Pyrimidines; Seizures; Status Epilepticus
PubMed: 38656874
DOI: 10.1016/j.celrep.2024.114144 -
Proceedings of the National Academy of... Apr 2024The development of seizures in epilepsy syndromes associated with malformations of cortical development (MCDs) has traditionally been attributed to intrinsic cortical...
The development of seizures in epilepsy syndromes associated with malformations of cortical development (MCDs) has traditionally been attributed to intrinsic cortical alterations resulting from abnormal network excitability. However, recent analyses at single-cell resolution of human brain samples from MCD patients have indicated the possible involvement of adaptive immunity in the pathogenesis of these disorders. By exploiting the MethylAzoxyMethanol (MAM)/pilocarpine (MP) rat model of drug-resistant epilepsy associated with MCD, we show here that the occurrence of status epilepticus and subsequent spontaneous recurrent seizures in the malformed, but not in the normal brain, are associated with the outbreak of a destructive autoimmune response with encephalitis-like features, involving components of both cell-mediated and humoral immune responses. The MP brain is characterized by blood-brain barrier dysfunction, marked and persisting CD8+ T cell invasion of the brain parenchyma, meningeal B cell accumulation, and complement-dependent cytotoxicity mediated by antineuronal antibodies. Furthermore, the therapeutic treatment of MP rats with the immunomodulatory drug fingolimod promotes both antiepileptogenic and neuroprotective effects. Collectively, these data show that the MP rat could serve as a translational model of epileptogenic cortical malformations associated with a central nervous system autoimmune response. This work indicates that a preexisting brain maldevelopment predisposes to a secondary autoimmune response, which acts as a precipitating factor for epilepsy and suggests immune intervention as a therapeutic option to be further explored in epileptic syndromes associated with MCDs.
Topics: Rats; Humans; Animals; Pilocarpine; Autoimmunity; Epilepsy; Seizures; Brain; Disease Models, Animal; Methylazoxymethanol Acetate
PubMed: 38635635
DOI: 10.1073/pnas.2319607121 -
International Journal of Developmental... Jun 2024According to experimental and clinical studies, status epilepticus (SE) causes neurodegenerative morphological changes not only in the hippocampus and other limbic... (Comparative Study)
Comparative Study
According to experimental and clinical studies, status epilepticus (SE) causes neurodegenerative morphological changes not only in the hippocampus and other limbic structures, it also affects the thalamus and the neocortex. In addition, several studies reported atrophy, metabolic changes, and neuronal degeneration in the dorsal striatum. The literature lacks studies investigating potential neuronal damage in the ventral component of the striatopallidal complex (ventral striatum [VS] and ventral pallidum) in SE experimentations. To better understand the development of neuronal damage in the striatopallidal complex associated with SE, the detected neuronal degeneration in the compartments of the VS, namely, the nucleus accumbens (NAc) and the olfactory tubercle (OT), was analyzed. The experiments were performed on Wistar rats at age of 25-day-old pups and 3-month-old adult animals. Lithium-pilocarpine model of SE was used. Lithium chloride (3 mmol/kg, ip) was injected 24 h before administering pilocarpine (40 mg/kg, ip). This presented study demonstrates the variability of post SE neuronal damage in 25-day-old pups in comparison with 3-month-old adult rats. The NAc exhibited small to moderate number of Fluoro-Jade B (FJB)-positive neurons detected 4 and 8 h post SE intervals. The number of degenerated neurons in the shell subdivision of the NAc significantly increased at survival interval of 12 h after the SE. FJB-positive neurons were evidently more prominent occupying the whole anteroposterior and mediolateral extent of the nucleus at longer survival intervals of 24 and 48 h after the SE. This was also the case in the bordering vicinity between the shell and the core compartments but with clusters of degenerating cells. The severity of damage of the shell subdivision of the NAc reached its peak at an interval of 24 h post SE. Isolated FJB-positive neurons were detected in the ventral peripheral part of the core compartment. Degenerated neurons persisted in the shell subdivision of the NAc 1 week after SE. However, the quantity of cell damage had significantly reduced in comparison with the aforementioned shorter intervals. The third layer of the OT exhibited more degenerated neurons than the second layer. The FJB-positive cells in the young animals were higher than in the adult animals. The morphology of those cells was identical in the two age groups except in the OT.
Topics: Animals; Status Epilepticus; Rats; Rats, Wistar; Male; Nerve Degeneration; Ventral Striatum; Neurons; Animals, Newborn; Pilocarpine; Disease Models, Animal; Lithium Chloride; Age Factors; Fluoresceins
PubMed: 38631684
DOI: 10.1002/jdn.10331 -
ACS Chemical Neuroscience May 2024The development of antiepileptic drugs is still a long process. In this study, heparin-modified superparamagnetic iron oxide nanoparticles (UFH-SPIONs) were prepared,...
Heparin-Modified Superparamagnetic Iron Oxide Nanoparticles Suppress Lithium Chloride/Pilocarpine-Induced Temporal Lobe Epilepsy in Rats through Attenuation of Inflammation and Oxidative Stress.
The development of antiepileptic drugs is still a long process. In this study, heparin-modified superparamagnetic iron oxide nanoparticles (UFH-SPIONs) were prepared, and their antiepileptic effect and underlying mechanism were investigated. UFH-SPIONs are stable, homogeneous nanosystems with antioxidant enzyme activity that are able to cross the blood-brain barrier (BBB) and enriched in hippocampal epileptogenic foci. The pretreatment with UFH-SPIONs effectively prolonged the onset of seizures and reduced seizure severity after lithium/pilocarpine (LP)-induced seizures in rats. The pretreatment with UFH-SPIONs significantly decreased the expression of inflammatory factors in hippocampal tissues, including IL-6, IL-1β, and TNF-α. LP-induced oxidative stress in hippocampal tissues was in turn reduced upon pretreatment with UFH-SPIONs, as evidenced by an increase in the levels of the antioxidant enzymes superoxide dismutase (SOD) and catalase (CAT) and a decrease in the level of lipid peroxidation (MDA). Moreover, the LP-induced upregulation of apoptotic cells was decreased upon pretreatment with UFH-SPIONs. Together, these observations suggest that the pretreatment with UFH-SPIONs ameliorates LP-induced seizures and downregulates the inflammatory response and oxidative stress, which exerts neuronal protection during epilepsy.
Topics: Animals; Oxidative Stress; Pilocarpine; Rats; Male; Magnetic Iron Oxide Nanoparticles; Epilepsy, Temporal Lobe; Lithium Chloride; Heparin; Inflammation; Rats, Sprague-Dawley; Hippocampus; Anticonvulsants
PubMed: 38630556
DOI: 10.1021/acschemneuro.4c00188 -
Journal of Oral Biosciences Jun 2024Local anesthetics act on G protein-coupled receptors (GPCRs); thus, their potential as allosteric modulators of GPCRs has attracted attention. Intracellular signaling...
OBJECTIVES
Local anesthetics act on G protein-coupled receptors (GPCRs); thus, their potential as allosteric modulators of GPCRs has attracted attention. Intracellular signaling via GPCRs involves both G-protein- and β-arrestin-mediated pathways. To determine the effects of local anesthetics on muscarinic acetylcholine receptors (mAChR), a family of GPCRs, we analyzed the effects of local anesthetics on mAChR-mediated Ca responses and formation of receptor-β-arrestin complexes in the HSY human parotid cell line.
METHODS
Ca responses were monitored by fura-2 spectrofluorimetry. Ligand-induced interactions between mAChR and β-arrestin were examined using a β-arrestin GPCR assay kit.
RESULTS
Lidocaine reduced mAChR-mediated Ca responses but did not change the intracellular Ca concentration in non-stimulated cells. The membrane-impermeant lidocaine analog QX314 and procaine inhibited mAChR-mediated Ca responses, with EC values of 48.0 and 20.4 μM, respectively, for 50 μM carbachol-stimulated Ca responses. In the absence of extracellular Ca, the pretreatment of cells with QX314 reduced carbachol-induced Ca release, indicating that QX314 reduced Ca release from intracellular stores. Lidocaine and QX314 did not affect store-operated Ca entry as they did not alter the thapsigargin-induced Ca response. QX314 and procaine reduced the carbachol-mediated recruitment of β-arrestin, and administration of procaine suppressed pilocarpine-induced salivary secretion in mice.
CONCLUSION
Local anesthetics, including QX314, act on mAChR to reduce carbachol-induced Ca release from intracellular stores and the recruitment of β-arrestin. These findings support the notion that local anesthetics and their derivatives are starting points for the development of functional allosteric modulators of mAChR.
Topics: Humans; Anesthetics, Local; beta-Arrestins; Calcium; Receptors, Muscarinic; Animals; Mice; Parotid Gland; Lidocaine; Cell Line; Carbachol; Calcium Signaling; Procaine
PubMed: 38614428
DOI: 10.1016/j.job.2024.04.002 -
Clinical Science (London, England :... May 2024Epilepsy, a chronic neurological disorder characterized by recurrent seizures, affects millions of individuals worldwide. Despite extensive research, the underlying...
Epilepsy, a chronic neurological disorder characterized by recurrent seizures, affects millions of individuals worldwide. Despite extensive research, the underlying mechanisms leading to epileptogenesis, the process by which a normal brain develops epilepsy, remain elusive. We, here, explored the immune system and spleen responses triggered by pilocarpine-induced status epilepticus (SE) focusing on their role in the epileptogenesis that follows SE. Initial examination of spleen histopathology revealed transient disorganization of white pulp, in animals subjected to SE. This disorganization, attributed to immune activation, peaked at 1-day post-SE (1DPSE) but returned to control levels at 3DPSE. Alterations in peripheral blood lymphocyte populations, demonstrated a decrease following SE, accompanied by a reduction in CD3+ T-lymphocytes. Further investigations uncovered an increased abundance of T-lymphocytes in the piriform cortex and choroid plexus at 3DPSE, suggesting a specific mobilization toward the Central Nervous System. Notably, splenectomy mitigated brain reactive astrogliosis, neuroinflammation, and macrophage infiltration post-SE, particularly in the hippocampus and piriform cortex. Additionally, splenectomized animals exhibited reduced lymphatic follicle size in the deep cervical lymph nodes. Most significantly, splenectomy correlated with improved neuronal survival, substantiated by decreased neuronal loss and reduced degenerating neurons in the piriform cortex and hippocampal CA2-3 post-SE. Overall, these findings underscore the pivotal role of the spleen in orchestrating immune responses and neuroinflammation following pilocarpine-induced SE, implicating the peripheral immune system as a potential therapeutic target for mitigating neuronal degeneration in epilepsy.
Topics: Pilocarpine; Animals; Status Epilepticus; Spleen; Male; Neuroinflammatory Diseases; Splenectomy; Rats, Sprague-Dawley; Hippocampus; Disease Models, Animal; T-Lymphocytes; Piriform Cortex; Neurons
PubMed: 38602323
DOI: 10.1042/CS20231621