Regulation Of Neurotransmission Research Articles

Detergent-free purification and reconstitution of functional human serotonin transporter (SERT) using diisobutylene maleic acid (DIBMA) copolymer

Structure and function analysis of human membrane proteins in lipid bilayer environments is acutely lacking despite the fundame1ntal cellular importance of these proteins and their dominance of drug targets. An underlying reason is that detailed study usually requires a potentially destabilising detergent purification of the proteins from their host membranes prior to subsequent reconstitution in a membrane mimic; a situation that is exacerbated for human membrane proteins due to the inherent difficulties in overexpressing suitable quantities of the proteins. We advance the promising styrene maleic acid polymer (SMA) extraction approach to introduce a detergent-free method of obtaining stable, functional human membrane transporters in bilayer nanodiscs directly from yeast cells. We purify the human serotonin transporter (hSERT) following overexpression in Pichia pastoris using diisobutylene maleic acid (DIBMA) as a superior method to traditional detergents or the more established styrene maleic acid polymer. hSERT plays a pivotal role in neurotransmitter regulation being responsible for the transport of the neurotransmitter 5-hydroxytryptamine (5-HT or serotonin). It is representative of the neurotransmitter sodium symporter (NSS) family, whose importance is underscored by the numerous diseases attributed to their malfunction. We gain insight into hSERT activity through an in vitro transport assay and find that DIBMA extraction improves the thermostability and activity of hSERT over the conventional detergent method.

Onecut Regulates Core Components of the Molecular Machinery for Neurotransmission in Photoreceptor Differentiation.

Photoreceptor cells (PRC) are neurons highly specialized for sensing light stimuli and have considerably diversified during evolution. The genetic mechanisms that underlie photoreceptor differentiation and accompanied the progressive increase in complexity and diversification of this sensory cell type are a matter of great interest in the field. A role of the homeodomain transcription factor Onecut (Oc) in photoreceptor cell formation is proposed throughout multicellular organisms. However, knowledge of the identity of the Oc downstream-acting factors that mediate specific tasks in the differentiation of the PRC remains limited. Here, we used transgenic perturbation of the Ciona robusta Oc protein to show its requirement for ciliary PRC differentiation. Then, transcriptome profiling between the trans-activation and trans-repression Oc phenotypes identified differentially expressed genes that are enriched in exocytosis, calcium homeostasis, and neurotransmission. Finally, comparison of RNA-Seq datasets in Ciona and mouse identifies a set of Oc downstream genes conserved between tunicates and vertebrates. The transcription factor Oc emerges as a key regulator of neurotransmission in retinal cell types.

Proteomic Analysis of Emodin Treatment in Neuropathic Pain Reveals Dysfunction of the Calcium Signaling Pathway.

BackgroundNeuropathic pain (NP) is a syndrome of pain mediated by distinct pathophysiological processes, and current treatments are not fully satisfactory. Emodin is an effective component of Chinese traditional medicine and has an alleviating effect on NP, but the pharmacological mechanism is not clear.MethodsWe used isobaric tags for relative and absolute quantitation (iTRAQ) technique integrated with liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis to identify potential targets of emodin in a rat peripheral nerve chronic constriction injury (CCI) model.ResultsA total of 177 differentially expressed proteins were identified among the sham group, CCI group, and emodin group, with a threshold of 1.2-fold change and a P value ≤ 0.05. Among them, 100 differentially expressed proteins (51 up-regulated and 49 down-regulated) were identified in the CCI group compared with sham group. Moreover, 108 differentially expressed proteins (65 up-regulated and 43 down-regulated) were identified in the emodin group with the CCI group as reference. The enrichment analysis of Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) revealed an important role of calcium signaling pathway, neurotransmitter regulation, and long-term potentiation (LTP) in emodin-treated CCI model. Real-time quantitative fluorescence PCR (qRT-PCR) and Western blot analysis revealed that emodin decreased expression of calcium signaling related proteins, including calmodulin (CaM) dependent protein kinase II (CaMK II), phospholipase Cβ1 (PLCβ1), protein kinase C (PKC), protein kinase C (PKA), and tropomyosin-related kinase B (TrkB), compared with the CCI group.ConclusionOverall, these findings indicated that emodin might alleviate NP by regulating the calcium signaling pathway.

A mathematical model of circadian rhythms and dopamine

BackgroundThe superchiasmatic nucleus (SCN) serves as the primary circadian (24hr) clock in mammals and is known to control important physiological functions such as the sleep-wake cycle, hormonal rhythms, and neurotransmitter regulation. Experimental results suggest that some of these functions reciprocally influence circadian rhythms, creating a highly complex network. Among the clock’s downstream products, orphan nuclear receptors REV-ERB and ROR are particularly interesting because they coordinately modulate the core clock circuitry. Recent experimental evidence shows that REV-ERB and ROR are not only crucial for lipid metabolism but are also involved in dopamine (DA) synthesis and degradation, which could have meaningful clinical implications for conditions such as Parkinson’s disease and mood disorders.MethodsWe create a mathematical model consisting of differential equations that express how the circadian variables are influenced by light, how REV-ERB and ROR feedback to the clock, and how REV-ERB, ROR, and BMAL1-CLOCK affect the dopaminergic system. The structure of the model is based on the findings of experimentalists.ResultsWe compare our model predictions to experimental data on clock components in different light-dark conditions and in the presence of genetic perturbations. Our model results are consistent with experimental results on REV-ERB and ROR and allow us to predict the circadian variations in tyrosine hydroxylase and monoamine oxidase seen in experiments. By connecting our model to an extant model of dopamine synthesis, release, and reuptake, we are able to predict circadian oscillations in extracellular DA and homovanillic acid that correspond well with experimental observations.ConclusionsThe predictions of the mathematical model are consistent with a wide variety of experimental observations. Our calculations show that the mechanisms proposed by experimentalists by which REV-ERB, ROR, and BMAL1-CLOCK influence the DA system are sufficient to explain the circadian oscillations observed in dopaminergic variables. Our mathematical model can be used for further investigations of the effects of the mammalian circadian clock on the dopaminergic system. The model can also be used to predict how perturbations in the circadian clock disrupt the dopaminergic system and could potentially be used to find drug targets that ameliorate these disruptions.

MiR-128 regulates epilepsy sensitivity in mice by suppressing SNAP-25 and SYT1 expression in the hippocampus

Epilepsy is accompanied by abnormal neurotransmission, and microRNAs, as versatile players in the modulation of gene expression, are important in epilepsy pathology. Here, we found that miR-128 expression was elevated in the acute seizure phase and decreased during the recurrent seizure phase after status epilepticus in mice. Both SNAP-25 and SYT1 are regulated by miR-128 in vitro and in vivo. Overexpressing miR-128 in cultured neurons decreased neurotransmitter released by suppressing SNAP-25 and SYT1 expression. Anti-miR-128 injection before kainic acid (KA) injection increased the sensitivity of mice to KA-induced seizures, while overexpressing miR-128 at the latent and recurrent phases had a neuroprotective effect in KA-induced seizures. Our study shows for the first time that miR-128, a key regulator of neurotransmission, plays an important role in epilepsy pathology and that miR-128 might be a potential candidate molecular target for epilepsy therapy.

Astroglia in the Vulnerability and Maintenance of Alcohol Use Disorders.

Changes induced in the morphology and the multiplicity of functional roles played by astrocytes in brain regions critical to the establishment and maintenance of alcohol abuse suggest that they make an important contribution to the vulnerability to alcohol use disorders. The understanding of the relevant mechanisms accounting for that contribution is complicated by the fact that alcohol itself acts directly on astrocytes altering their metabolism, gene expression, and plasticity, so that the ultimate result is a complex interaction of various cellular pathways, including intracellular calcium regulation, neuroimmune responses, and regulation of neurotransmitter and gliotransmitter release and uptake. The recent years have seen a steady increase in the characterization of several of the relevant mechanisms, but much remains to be done for a full understanding of the astrocytes' contribution to the vulnerability to alcohol dependence and abuse and for using that knowledge in designing effective therapies for AUDs.

IL-33 Changes Our “Gut Feelings” about Serotonin

Many studies highlight direct interactions between immune cells and enteric neurons, but whether immune signals can indirectly modulate enteric function through neurotransmitter regulation is poorly understood. In this issue of Immunity, Chen etal. reveal how IL-33 induces intestinal serotonin to promote gut motility.

A dual role for α-synuclein in facilitation and depression of dopamine release from substantia nigra neurons in vivo

α-Synuclein is expressed at high levels at presynaptic terminals, but defining its role in the regulation of neurotransmission under physiologically relevant conditions has proven elusive. We report that, in vivo, α-synuclein is responsible for the facilitation of dopamine release triggered by action potential bursts separated by short intervals (seconds) and a depression of release with longer intervals between bursts (minutes). These forms of presynaptic plasticity appear to be independent of the presence of β- and γ-synucleins or effects on presynaptic calcium and are consistent with a role for synucleins in the enhancement of synaptic vesicle fusion and turnover. These results indicate that the presynaptic effects of α-synuclein depend on specific patterns of neuronal activity.

Wireless nanotechnologies light up the next frontier in cell Calcium signalling

AbstractCalcium ions impact nearly every aspect of cellular life, playing crucial roles as secondary messengers in regulation of neurotransmission, cell proliferation, migration and differentiation processes, intracellular homeostasis, long-distance signal propagation and stimuli physiological response. Despite its key-role, available techniques to study and selectively regulate Ca2+ signalling largely rely on chemical and electrical approaches, which often cannot ensure the necessary spatial and temporal resolution, specificity, modulation and reversal capability. In this context, Ca2+ modulation based on physical stimuli, such as magnetic, mechanical and optical tools, are emerging ass promising innovative solutions. Here, we focus our attention on a subclass of these approaches, namely wireless-activated techniques, and on functional materials able to act as non-invasive transduction elements. We present an overview of most recent outcomes in the field, and we critically evaluate their advantages and drawbacks. This work is mainly directed to the material science community, but hopefully it will provide a useful perspective also to the broader readership of biotechnologists, physiologists and clinicians.

Mustard Leaf Extract Suppresses Psychological Stress in Chronic Restraint Stress-Subjected Mice by Regulation of Stress Hormone, Neurotransmitters, and Apoptosis.

Mustard leaf (Brassica juncea var. crispifolia L. H. Bailey) has been reported to have psychological properties such as anti-depressant activities. However, studies on chronic stress and depression caused by restraint have not been conducted. Therefore, this study aimed to evaluate the effects of a mustard leaf (ML) extract on chronic restraint stress (CRS) in mice. Male mice were subjected to a CRS protocol for a period of four weeks to induce stress. The results showed that the ML extract (100 and 500 mg/kg/perorally administered for four weeks) significantly decreased corticosterone levels and increased neurotransmitters levels in stressed mice. Apoptosis by CRS exposure was induced by Bcl-2 and Bax expression regulation and was suppressed by reducing caspase-3 and poly (ADP-ribose) polymerase expression after treatment with the ML extract. Our results confirmed that apoptosis was regulated by increased expression of brain-derived neurotrophic factor (BDNF). Additionally, cytokine levels were regulated by the ML extract. In conclusion, our results showed that the ML extract relieved stress effects by regulating hormones and neurotransmitters in CRS mice, BDNF expression, and apoptosis in the brain. Thus, it can be suggested that the studied ML extract is an agonist that can help relieve stress and depression.

Investigation of the antidepressant effects of Shu-Gan-Jie- Yu granule and its mechanism of action

Purpose: To study the antidepressant effects of Shu-Gan-Jie-Yu granule (SJG) and its possible mechanisms in mice.Methods: The anti-depressive effects of SJG were evaluated by three techniques, viz, forced swimming test (FST), tail suspension test (TST) and open field test (OFT). The levels of the neurotransmitters norepinephrine (NE), DA, and 5-HT in the brains of depressive mice were determined using commercially available kits. In addition, the effects of SJG on the BDNF expression in the mice brain were determined by western blot.Results: Administration of SJG significantly reduced the duration time of immobility in the experiments of FST and TST. In addition, relative to the control mice, SJG (800 mg/kg) administration significantly affected the mobility performance (p < 0.05) of mice. The levels of the three neurotransmitters (DA, NE and 5-HT) and BDNF in the brains of depressive mice were increased by treatment with SJG at the doses of 200, 400 and 800 mg/kg (p < 0.05). The results suggested that SJG exerted a significant antidepressant effect, which could be attributed to increases in the levels of neurotransmitters, and the up-regulation of BDNF expression.Conclusion: The results suggested that SJG exerted a significant antidepressant effect, most probably via regulation of related neurotransmitters (including DA, NE, and 5-HT) and BDNF in the brain.
 Keywords: Shu-Gan-Jie-Yu granule, Antidepressant, dopamine, norepinephrine, 5-hydroxytryptamine, brain-derived neurotrophic factor

The Underlying Mechanism of Paeonia lactiflora Pall. in Parkinson's Disease Based on a Network Pharmacology Approach.

Background: Parkinson’s disease (PD) is the second most common neurodegenerative disease worldwide, yet as of currently, there is no disease-modifying therapy that could delay its progression. Paeonia lactiflora Pall. is the most frequently used herb in formulas for PD in Traditional Chinese Medicine and also a potential neuroprotective agent for neurodegenerative diseases, while its mechanisms remain poorly understood. In this study, we aim to explore the underlying mechanism of P. lactiflora in treating PD utilizing a network pharmacology approach. Methods: The protein targets of P. lactiflora ingredients and PD were first obtained from several databases. To clarify the key targets, a Protein-Protein-Interaction (PPI) network was constructed and analyzed on the String database, and then enrichment analysis was performed by the Metascape platform to determine the main Gene Ontology biological processes and Kyoto Encyclopedia of Genes and Genomes pathways. Finally, the Ingredient-Target-Pathway (I-T-P) network was constructed and analyzed by Cytoscape software. Results: Six active ingredients of P. lactiflora (kaempferol, ß-sitosterol, betulinic acid, palbinone, paeoniflorin and (+)-catechin) as well as six core targets strongly related to PD treatment [AKT1, interleukin-6, CAT, Tumor necrosis factor (TNF), CASP3, and PTGS2] were identified. The main pathways were shown to involve neuroactive ligand-receptor interaction, Calcium signaling pathway, PI3-Akt signaling pathway, TNF signaling pathway, and apoptosis signaling pathway. The main biological process included the regulation of neurotransmitter levels. Conclusion: P. lactiflora may retard neurodegeneration by reducing neuroinflammation, inhibiting intrinsic and extrinsic apoptosis, and may improve motor and non-motor symptoms by regulating the levels of neurotransmitters. Our study has revealed the mechanism of P. lactiflora in the treatment of PD and may contribute to novel drug development for PD.

Transcranial Direct Current Stimulation as an Add-on Treatment to Cognitive-Behavior Therapy in First Episode Drug-Naïve Major Depression Patients: The ESAP Study Protocol.

Background: Major Depressive Disorder (MDD) affects more than 264 million people worldwide. Current treatments include the use of psychotherapy and/or drugs, however ~30% of patients either do not respond to these treatments, or do not tolerate the side effects associated to the use of pharmacological interventions. Thus, it is important to study non-pharmacological interventions targeting mechanisms not directly involved with the regulation of neurotransmitters. Several studies demonstrated that transcranial Direct Current Stimulation (tDCS) can be effective for symptoms relief in MDD. However, tDCS seems to have a better effect when used as an add-on treatment to other interventions.Methods/Design: This is a study protocol for a parallel, randomized, triple-blind, sham-controlled clinical trial in which a total of 90 drug-naïve, first-episode MDD patients (45 per arm) will be randomized to one of two groups to receive a 6-weeks of CBT combined with either active or sham tDCS to the dorsolateral prefrontal cortex (DLPFC). The primary outcome will depressive symptoms improvement as assessed by the Montgomery-Asberg Depression Rating Scale (MADRS) at 6-weeks. The secondary aim is to test whether CBT combined with tDCS can engage the proposed mechanistic target of restoring the prefrontal imbalance and connectivity through the bilateral modulation of the DLPFC, as assessed by changes over resting-state and emotional task eliciting EEG.Discussion: This study evaluates the synergetic clinical effects of CBT and tDCS in the first episode, drug-naïve, patients with MDD. First episode MDD patients provide an interesting opportunity, as their brains were not changed by the pharmacological treatments, by the time course, or by the recurrence of MDD episodes (and other comorbidities).Trial Registration: This study is registered with the United States National Library of Medicine Clinical Trials Registry (NCT03548545). Registered June 7, 2018, clinicaltrials.gov/ct2/show/NCT03548545. Protocol Version 1.

CAMP-regulated phosphoproteins DARPP-32, ARPP16/19, and RCS modulate striatal signal transduction through protein kinases and phosphatases.

Decades of research led by Paul Greengard identified protein phosphorylation as a ubiquitous and vital post-translational modification involved in many neuronal signaling pathways. In particular, his discovery that second messenger-regulated protein phosphorylation plays a central role in the propagation and transduction of signals in the nervous system has been essential in understanding the molecular mechanisms of neuronal communication. The establishment of dopamine (DA) as an essential neurotransmitter in the central nervous system, combined with observations that DA activates G-protein-coupled receptors to control the production of cyclic adenosine monophosphate (cAMP) in postsynaptic neurons, has provided fundamental insight into the regulation of neurotransmission. Notably, DA signaling in the striatum is involved in many neurological functions such as control of locomotion, reward, addiction, and learning, among others. This review focuses on the history, characterization, and function of cAMP-mediated regulation of serine/threonine protein phosphatases and their role in DA-mediated signaling in striatal neurons. Several small, heat- and acid-stable proteins, including DARPP-32, RCS, and ARPP-16/19, were discovered by the Greengard laboratory to be regulated by DA- and cAMP signaling, and found to undergo a complex but coordinated sequence of phosphorylation and dephosphorylation events. These studies have contributed significantly to the establishment of protein phosphorylation as a ubiquitous and vital process in signal propagation in neurons, paradigm shifting discoveries at the time. Understanding DA-mediated signaling in the context of signal propagation has led to numerous insights into human conditions and the development of treatments and therapies.

Esophagus: serotoninerdgic regulation

Introduction: Serotonin (5-hydroxytryptamine, 5-HT) is a regulatory neurotransmitter and a hormone in the CNS and hole organs, the esophagus including. It is known that serotonin, activating its own receptors, stimulates contractile activity of the esophageal muscles. However, role of different type receptors in the 5-HT induced contractile activity of the esophagus is insufficiently known.The aim: — to determine which type of 5-HT receptors mediate serotonin dependent contractile activity of the esophagus.Material and methods: This is a electromyography study of rat esophagus contractile activity under serotonin stimulation of 5-HT3,4 and 5-HT2,1 receptors separately modulated. The role of different serotonin receptors in the 5-HT induced contractile activity of the esophagus was evaluated by measuring the amplitude and frequency of the slow wave electromyogram (EMG) by the noninvasive microelectrodes imposed on the adventitial layer of the esophagus.Results: Administration of the 5-HT3,4 receptors inhibitors excluded caused by serotonin the increment of EMG activity of the contractile activity of the esophagus. Administration of the 5-HT1,2 receptors inhibitors blocked the serotonin enhanced EMG activity of the esophagus.Conclusion: Our results indicate that serotonin is the important player in the regulation of the rat's esophagus contractility; 5-HT enhancing effect on contraction of the esophageal smooth muscles is mediated through the activation of 5-HT1,2 receptors expressed on the smooth muscle cells, and by activation of 5-HT3,4 receptors expressed on the ganglion neurons.

Perceptions of violence based on genetics: the influence of monoamine oxidase A in the origin of violent behavior

Objective: To gather scientific literature regarding the influence of the low activity of the enzyme Monoaminoxidase A, responsible for the regulation of brain neurotransmitters, on criminal and / or violent behavior. Methods: Searches were performed in the PubMed and SCOPUS databases using “genes”, “crime” and “monoaminoxidase”, selecting articles published between 2009 and 2019. Results: The selected sample, after applying the inclusion criteria and exclusion, was composed of 14 articles. Discussion: A relationship was found between low levels of activity of Monoaminoxidase A, when associated with socio-environmental factors, with a greater tendency to commit criminal and / or violent acts. Among the socio-environmental factors, the most cited is the experience of trauma in childhood. However, there were copyright disagreements, which demonstrates the need for further investigation and development of studies on the subject. Final considerations: It was found that individuals with low Monoaminoxidase A activity and who experienced stressful socioenvironmental factors are more prone to aggressive and / or violent behavior.

Alleviation of Depression by Glucagon-Like Peptide 1 Through the Regulation of Neuroinflammation, Neurotransmitters, Neurogenesis, and Synaptic Function.

Depression has emerged as a major cause of mortality globally. Many studies have reported risk factors and mechanisms associated with depression, but it is as yet unclear how these findings can be applied to the treatment and prevention of this disorder. The onset and recurrence of depression have been linked to diverse metabolic factors, including hyperglycemia, dyslipidemia, and insulin resistance. Recent studies have suggested that depression is accompanied by memory loss as well as depressive mood. Thus, many researchers have highlighted the relationship between depressive behavior and metabolic alterations from various perspectives. Glucagon-like peptide-1 (GLP-1), which is secreted from gut cells and hindbrain areas, has been studied in metabolic diseases such as obesity and diabetes, and was shown to control glucose metabolism and insulin resistance. Recently, GLP-1 was highlighted as a regulator of diverse pathways, but its potential as the therapeutic target of depressive disorder was not described comprehensively. Therefore, in this review, we focused on the potential of GLP-1 modulation in depression.

Plasma metabolites associated with biomarker evidence of neurodegeneration in cognitively normal older adults.

Alzheimer's disease (AD) is a progressive neurodegenerative disorder that currently has no cure. Identifying biochemical changes associated with neurodegeneration prior to symptom onset, will provide insight into the biological mechanisms associated with neurodegenerative processes, that may also aid in identifying potential drug targets. The current study therefore investigated associations between plasma neurofilament light chain (NF-L), a marker of neurodegeneration, with plasma metabolites that are products of various cellular processes. Plasma NF-L, measured by ultrasensitive Single molecule array (Simoa) technology (Quanterix) and plasma metabolites, measured by mass-spectrometry (AbsoluteIDQ® p400HR kit, BIOCRATES), were assessed in the Kerr Anglican Retirement Village Initiative in Ageing Health (KARVIAH) cohort comprising 100 cognitively normal older adults. Metabolites belonging to biogenic amine (creatinine, symmetric dimethylarginine, asymmetric dimethylarginine; ADMA, kynurenine, trans-4-hydroxyproline), amino acid (citrulline, proline, arginine, asparagine, phenylalanine, threonine) and acylcarnitine classes were observed to have positive correlations with plasma NF-L, suggesting a link between neurodegeneration and biological pathways associated with neurotransmitter regulation, nitric oxide homoeostasis, inflammation and mitochondrial function. Additionally, after stratifying participants based on low/high brain amyloid-β load (Aβ ±) assessed by positron emission tomography, while creatinine, SDMA and citrulline correlated with NF-L in both Aβ- and Aβ+ groups, ADMA, proline, arginine, asparagine, phenylalanine and acylcarnitine species correlated with NF-L only in the Aβ+ group after adjusting for confounding variables, suggesting that the association of these metabolites with neurodegeneration may be relevant to AD-related neuropathology. Metabolites identified to be associated with plasma NF-L may have the potential to serve as prognostic markers for neurodegenerative diseases, however, further studies are required to validate the current findings in an independent cohort, both cross-sectionally and longitudinally.

Non-invasive vagus nerve stimulation for primary headache: A clinical update.

Non-invasive vagus nerve stimulation (nVNS) is a proven treatment for cluster headache and migraine. Several possible mechanisms of action by which nVNS mitigates headache have been identified. We conducted a narrative review of recent scientific and clinical research into nVNS for headache, including findings from mechanistic studies and their possible relationships to the clinical effects of nVNS. Findings from animal and human studies have provided possible mechanistic explanations for nVNS efficacy in headache involving four core areas: Autonomic nervous system functions; cortical spreading depression inhibition; neurotransmitter regulation; and nociceptive modulation. We discuss how overlap and interplay among these areas may underlie the utility of nVNS in the context of clinical evidence supporting its safety and efficacy as acute and preventive therapy for both cluster headache and migraine. Possible future nVNS applications are also discussed. Significant progress over the past several years has yielded valuable mechanistic and clinical evidence that, combined with the excellent safety and tolerability profile of nVNS, suggests that it should be considered a first-line treatment for both acute and preventive treatment of cluster headache, an effective option for acute treatment of migraine, and a highly relevant, practical option for migraine prevention.

Virodhamine, an endocannabinoid, induces megakaryocyte differentiation by regulating MAPK activity and function of mitochondria.

Endocannabinoids are well-known regulators of neurotransmission by activating the cannabinoid (CB) receptors. Endocannabinoids are being used extensively for the treatment of various neurological disorders such as Alzheimer's and Parkinson's diseases. Although endocannabinoids are well studied in cell survival, proliferation, and differentiation in various neurological disorders and several cancers, the functional role in the regulation of blood cell development is less examined. In the present study, virodhamine, which is an agonist of CB receptor-2, was used to examine its effect on megakaryocytic development from a megakaryoblastic cell. We observed that virodhamine increases cell adherence, cell size, and cytoplasmic protrusions. Interestingly, we have also observed large nucleus and increased expression of megakaryocytic marker (CD61), which are the typical hallmarks of megakaryocytic differentiation. Furthermore, the increased expression of CB2 receptor was noticed in virodhamine-induced megakaryocytic cells. The effect of virodhamine on megakaryocytic differentiation could be mediated through CB2 receptor. Therefore, we have studied virodhamine induced molecular regulation of megakaryocytic differentiation; mitogen-activated protein kinase (MAPK) activity, mitochondrial function, and reactive oxygen species (ROS) production were majorly affected. The altered mitochondrial functions and ROS production is the crucial event associated with megakaryocytic differentiation and maturation. In the present study, we report that virodhamine induces megakaryocytic differentiation by triggering MAPK signaling and ROS production either through MAPK effects on ROS-generating enzymes or by the target vanilloid receptor 1-mediated regulation of mitochondrial function.