Benzylideneacetophenone Derivative Alleviates Arthritic Symptoms via Modulation of the MAPK Signaling Pathway.

The benzylideneacetophenone derivative 3-(4-hydroxy-3-methoxy-phenyl)-1-{3-[1]-phenyl}-propenone (JC3 dimer) was synthesized through the dimerization of JC3. To investigate the inhibitory effects of JC3 dimer, the carrageenan/kaolin (C/K)-induced knee arthritis rat model was used in vivo and rheumatoid arthritis (RA) patient-derived fibroblast-like synoviocytes (FLS) were used in vitro. In the C/K rat model, JC3 dimer was given after arthritis induction for 6 days at the concentrations of 1, 5, or 10 mg/kg/day. Manifestation of arthritis was evaluated using knee thickness, weight distribution ratio (WDR), and squeaking test. The levels of prostaglandin E2 (PGE2), interleukin (IL)-6, and tumor necrosis factor (TNF)-α in the serum of JC3 dimer-treated arthritic rats were also analyzed. Histological examination of the knee joints was also done. For the FLS, the cells were stimulated using IL-1ß and concentrations of 1, 5, and 10 µg/mL JC3 dimer were used. The levels of IL-8, IL-6, and PGE2 were measured in stimulated FLS treated with JC3 dimer. At days 5 to 6 after arthritis induction, JC3 dimer treatment significantly decreased arthritic symptoms and reduced the inflammation in the knee joints in the histology of knee tissues in C/K-arthritic rats. In stimulated FLS, JC3 dimer suppressed the increase of IL-8, IL-6, and PGE2. These findings suggest that JC3 dimer has suppressive effects on arthritis, and that JC3 dimer can be a potential agent for arthritis therapy.

Morphine Dependence is Attenuated by Treatment of 3,4,5-Trimethoxy Cinnamic Acid in Mice and Rats.

The effect of 3, 4, 5-trimethoxy cinnamic acid (TMCA) against morphine-induced dependence in mice and rats was investigated. Mice were pretreated with TMCA and then morphine was injected intraperitoneally; whereas rats were treated with TMCA (i.p.) and infused with morphine into the lateral ventricle of brain. Naloxone-induced morphine withdrawal syndrome and conditioned place preference test were performed. Moreover, western blotting and immunohistochemistry were used to measure protein expressions. Number of naloxone-precipitated jumps and conditioned place preference score in mice were attenuated by TMCA. Likewise, TMCA attenuated morphine dependent behavioral patterns such as diarrhea, grooming, penis licking, rearing, teeth chattering, and vocalization in rats. Moreover, the expression levels of pNR1and pERK in the frontal cortex of mice and cultured cortical neurons were diminished by TMCA. In the striatum, pERK expression was attenuated despite unaltered expression of pNR1 and NR1. Interestingly, morphine-induced elevations of FosB/ΔFosB+ cells were suppressed by TMCA (50, 100 mg/kg) in the nucleus accumbens sub-shell region of mice. In conclusion, TMCA could be considered as potential therapeutic agent against morphine-induced dependence.

Chronic stress-induced dendritic reorganization and abundance of synaptosomal PKA-dependent CP-AMPA receptor in the basolateral amygdala in a mouse model of depression.

Chronic stress is a precipitating factor for disorders including depression. The basolateral amygdala (BLA) is a critical substrate that interconnects with stress-modulated neural networks to generate emotion- and mood-related behaviors. The current study shows that 3 h per day of restraint stress for 14 days caused mice to exhibit long-term depressive behaviors, manifested by disrupted sociality and despair levels, which were rescued by fluoxetine. These behavioral changes corresponded with morphological and molecular changes in BLA neurons, including chronic stress-elicited increases in arborization, dendritic length, and spine density of BLA principal neurons. At the molecular level, calcium-permeable α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (CP-AMPARs) within the synaptosome exhibited an increased GluR1:GluR2 subunit ratio. We also observed increased GluR1 phosphorylation at Ser 845 and enhanced cyclic AMP-dependent protein kinase (PKA) activity in the BLA. These molecular changes reverted to the basal state post-treatment with fluoxetine. The expression of synaptophysin (SYP) and postsynaptic density protein 95 (PSD-95) at BLA neuronal synapses was also enhanced by chronic stress, which was reversed post-treatment. Finally, chronic stress-provoked depressive behavior was overcome by local blockage of CP-AMPARs in the BLA via stereotaxic injection (IEM-1460). Chronic stress-elicited depressive behavior may be due to hypertrophy of BLA neuronal dendrites and increased of PKA-dependent CP-AMPAR levels in BLA neurons. Furthermore, fluoxetine can reverse chronic stress-triggered cytoarchitectural and functional changes of BLA neurons. These findings provide insights into depression-linked structural and functional changes in BLA neurons.

Concanavalin A Induces Cortical Neuron Apoptosis by Causing ROS Accumulation and Tyrosine Kinase Activation.

The lectin, concanavalin A (Con A), is the most extensively investigated member of the lectin family of plant proteins, but its effects on cortical neurons and astrocytes are poorly understood. In cultured cortical neurons and astrocytes, Con A exhibited dose-dependent neurotoxicity, but this was not observed in astrocytes. Similarly, in the cortical areas of rat brains, intracranial administration of Con A caused neuronal but no astrocyte damage. Methyl-α-D-mannopyranoside, a competitor of Con A, blocked Con A-induced cell death, whereas AMPA/KA receptor antagonists showed partial blocking effects. Furthermore, the mRNA levels of TNF-α, IL-1ß, and IL-6 were elevated in astrocytes and cortical neurons treated with Con A. Intracellular reactive oxygen species (ROS) levels were increased in Con A-treated cortical neurons, and N-acetyl-cysteine (NAC, an antioxidant) and diphenyleneiodonium (DPI, a NADPH oxidase inhibitor) reduced intracellular ROS accumulation. Likewise, AG556 (a TNF-α inhibitor) and AG82 (a tyrosine kinase inhibitor) both reduced Con A-induced intracellular ROS accumulation. Furthermore, Con A-induced tyrosine phosphorylation was decreased by NAC and by AG556. Taken together, Con A-induced apoptosis in cortical neurons occurred as a sequel to Con A binding to neuronal glycoproteins and intracellular ROS accumulation. Interestingly, Con A-induced cellular damage was observed in cortical neurons but not in astrocytes or microglia.

Inhibition of Ceramide Decreased the Expression of ATP-Binding Cassette Transporter G5/8 mRNA in an Animal Model of Cholesterol Gallstone.

BACKGROUND: The increased risk of gallstone has been reported in patients with ATP-binding cassette (ABC) transporter polymorphism. The half-transporters ABCG5 and ABCG8 mediate the efflux of cholesterol in hepatocytes and the intestine. We investigated whether ceramide plays a role in cholesterol efflux through the ABC transporters. METHODS: Six-week-old C57BL/6J mice were assigned to 3 groups. The normal group (n = 5) was fed a normal chow diet, the cholesterol group (n = 10) was fed a lithogenic diet, and the myriocin group (n = 15) was fed the lithogenic diet and myriocin, a specific inhibitor of serine-palmitoyl transferase. After 6 weeks, the ABCG5 and ABCG8 transporters were analyzed. RESULTS: The rate of cholesterol gallstone formation in cholesterol group was also higher than that in normal and myriocin groups (0, 70, and 40%, respectively). ABCG5 and ABCG8 mRNA levels were significantly increased in cholesterol group and less increased in myriocin group, relative to that in normal group (p < 0.05). CONCLUSIONS: The inhibition of ceramide biosynthesis by myriocin suppressed gallstone formation and ABCG5/8 mRNA expression. We expect that ceramide's role as a regulator of the ABCG5/8 transporter might be linked to cholesterol gallstone formation.

BPA-toxicity via superoxide anion overload and a deficit in ß-catenin signaling in human bone mesenchymal stem cells.

Bisphenol A (BPA), used in the manufacture of products based on polycarbonate plastics and epoxy resins, is well known as an endocrine-disrupting monomer. In the current study, BPA increased cytotoxicity in hBMSCs in a dose- and time-dependent manner, concomitantly with increased lipid peroxidation. Increased cell death in BPA-treated cells was markedly blocked by pretreatment with the superoxide dismutase mimetic MnTBAP and MnTMPyP, but not by catalase, glutathione, the glutathione peroxidase mimetic ebselen, the NOS inhibitor NAME, or the xanthine oxidase inhibitor allopurinol. Furthermore, the decline in nuclear ß-catenin and cyclin D1 levels in hBMSCs exposed to BPA was reversed by MnTBAP treatment. Finally, treatment of hBMSCs with the GSK3ß inhibitor LiCl2 increased nuclear ß-catenin levels and significantly attenuated cytotoxicity compared with BPA treatment. Our current results in hBMSCs exposed to BPA suggest that BPA causes a disturbance in ß-catenin signaling via a superoxide anion overload. © 2016 The Authors Environmental Toxicology Published by Wiley Periodicals, Inc. Environ Toxicol 32: 344-352, 2017.

Practical Synthesis of Chalcone Derivatives and Their Biological Activities.

Practical synthesis and biological activities of 4-hydroxy-3-methoxy-2-propene derivatives are described. The novel chalcone derivatives were prepared by acid catalysed one-step condensation of 1,3- or 1,4-diacetylbenzene and 1,3,5-triacetylbenzene with 4-hydroxy-3-methoxybenzaldehyde. They were then evaluated for free radical scavenging activity, suppression of lipopolysaccharides (LPS)-induced NO generation, and anti-excitotoxicity in vitro. It was found that all compounds showed good effects for 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radical scavenging, LPS-induced NO generation, and anti-neurotoxicity. Compounds 6 and 7 were potent suppressor of NO generation with the concentration range 10 µM and especially compound 8 showed very potent anti-inflammatory activity with 1 µM. In addition, the di- and tri-acetylbenzyl derivatives 6, 7, and 8 showed enhanced anti-neurotoxicity activity in cultured cortical neurons. Molecular modelling studies to investigate the chemical structural characteristics required for the enhanced biological activities interestingly revealed that compound 8 has the smallest highest occupied molecular orbital-lowest energy unoccupied molecular orbital (HOMO-LUMO) gap, which signifies easy electron and radical transfer between HOMO and LUMO in model studies.

Inhibition of p38 pathway-dependent MPTP-induced dopaminergic neurodegeneration in estrogen receptor alpha knockout mice.

Approximately, 7-10 million people in the world suffer from Parkinson's disease (PD). Recently, increasing evidence has suggested the protective effect of estrogens against nigrostriatal dopaminergic damage in PD. In this study, we investigated whether estrogen affects 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced behavioral impairment in estrogen receptor alpha (ERα)-deficient mice. MPTP (15mg/kg, four times with 1.5-h interval)-induced dopaminergic neurodegeneration was evaluated in ERα wild-type (WT) and knockout (KO) mice. Larger dopamine depletion, behavioral impairments (Rotarod test, Pole test, and Gait test), activation of microglia and astrocytes, and neuroinflammation after MPTP injection were observed in ERα KO mice compared to those in WT mice. Immunostaining for tyrosine hydroxylase (TH) after MPTP injection showed fewer TH-positive neurons in ERα KO mice than WT mice. Levels of dopamine and 3,4-dihydroxyphenylacetic acid (DOPAC, metabolite of dopamine) were also lowered in ERα KO mice after MPTP injection. Interestingly, a higher immunoreactivity for monoamine oxidase (MAO) B was found in the substantia nigra and striatum of ERα KO mice after MPTP injection. We also found an increased activation of p38 kinase (which positively regulates MAO B expression) in ERα KO mice. In vitro estrogen treatment inhibited neuroinflammation in 1-methyl-4-phenyl pyridium (MPP+)-treated cultured astrocyte cells; however, these inhibitory effects were removed by p38 inhibitor. These results indicate that ERα might be important for dopaminergic neuronal survival through inhibition of p38 pathway.

Modulation of NMDA Receptor Subunits Expression by Concanavalin A.

The processes of N-methyl-D-aspartate (NMDA) receptor subunits expression were examined in cortical neurons and rat brain in order to investigate how the concanavalin A (Con A) modulates neuronal cells. Con A modulated the expression of NMDA receptor subunits in cultured cortical cells. Con A augmented the level of intracellular Ca(2+) by α-amino-3-hydroxy-5-methyl-isoxazole-4-propionate (AMPA). We determined whether activation of AMPA receptors was involved in the regulation of NMDA receptor expression with Con A by blocking the desensitization of AMPA receptors. The results showed that AMPA receptor antagonists suppressed NMDA receptor subunits expression in Con A-treated cortical neuronal cells. PMA elevated the expression of NMDA receptor subunits, while PKC inhibitor and tyrosine kinases inhibitor suppressed the expression of NMDA receptor subunits. Furthermore, it was shown that NMDA receptor subunits expression was modulated in a region-specific manner after the sustained microinfusion of Con A into the cerebroventricle of the rat brain. Collectively, it could be presumed that the AMPA receptor activation was involved in Con A-induced modulation of NMDA receptor subunits expression.

Hepatoprotective effect of licorice, the root of Glycyrrhiza uralensis Fischer, in alcohol-induced fatty liver disease.

BACKGROUND: Our previous study suggested that licorice has anti-inflammatory activity in lipopolysaccharide-stimulated microglial cells and anti-oxidative activity in tert-butyl hydroperoxide-induced oxidative liver damage. In this study, we evaluated the effect of licorice on chronic alcohol-induced fatty liver injury mediated by inflammation and oxidative stress. METHODS: Raw licorice was extracted, and quantitative and qualitative analysis of its components was performed by using LC-MS/MS. Mice were fed a liquid alcohol diet with or without licorice for 4 weeks. RESULTS: We have standardized 70% fermented ethanol extracted licorice and confirmed by LC-MS/MS as glycyrrhizic acid (GA), 15.77 ± 0.34 µg/mg; liquiritin (LQ), 14.55 ± 0.42 µg/mg; and liquiritigenin (LG), 1.34 ± 0.02 µg/mg, respectively. Alcohol consumption increased serum alanine aminotransferase and aspartate aminotransferase activities and the levels of triglycerides and tumor necrosis factor (TNF)-α. Lipid accumulation in the liver was also markedly induced, whereas the glutathione level was reduced. All these alcohol-induced changes were effectively inhibited by licorice treatment. In particular, the hepatic glutathione level was restored and alcohol-induced TNF-α production was significantly inhibited by licorice. CONCLUSION: Taken together, our data suggests that protective effect of licorice against alcohol-induced liver injury may be attributed to its anti-inflammatory activity and enhancement of antioxidant defense.

Anti-tumor activity of benzylideneacetophenone derivatives via proteasomal inhibition in prostate cancer cells.

A number of some chalcone derivatives possess promising biological properties including anti-inflammation, anti-oxidant, and anti-tumor activity. Although it has been shown that some derivatives of chalcone induce apoptosis in different kinds of cancer cells, the involved mechanism of action is not well defined. The purpose of this study is to investigate the primary target of a benzylideneacetophenone derivative (JC3), which is a synthetic compound derived from the chalcone family, in human cancer, using prostate cancer cells as a working model. Herein, we show that JC3 inhibits proteasomal activity as indicated by both in vitro and in cell-based assays. Especially, the JC3-dimer was more potent than monomer in the aspect of proteasome inhibition, which induced apoptosis significantly in the prostate cancer cells. Owing to the critical roles of the proteasome in the biology of human tumor progression, invasion, and metastasis, these findings give an important clue for the development of novel anti-tumor agents.

Anti-Inflammatory activities of licorice extract and its active compounds, glycyrrhizic acid, liquiritin and liquiritigenin, in BV2 cells and mice liver.

This study provides the scientific basis for the anti-inflammatory effects of licorice extract in a t-BHP (tert-butyl hydrogen peroxide)-induced liver damage model and the effects of its ingredients, glycyrrhizic acid (GA), liquiritin (LQ) and liquiritigenin (LG), in a lipopolysaccharide (LPS)-stimulated microglial cell model. The GA, LQ and LG inhibited the LPS-stimulated elevation of pro-inflammatory mediators, such as inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), tumor necrosis factor (TNF)-alpha, interleukin (IL)-1beta and interleukin (IL)-6 in BV2 (mouse brain microglia) cells. Furthermore, licorice extract inhibited the expression levels of pro-inflammatory cytokines (TNF-α, IL-1ß and IL-6) in the livers of t-BHP-treated mice models. This result suggested that mechanistic-based evidence substantiating the traditional claims of licorice extract and its three bioactive components can be applied for the treatment of inflammation-related disorders, such as oxidative liver damage and inflammation diseases.

Corrigendum to "Gintonin regulates inflammation in human IL-1ß-stimulated fibroblast-like synoviocytes and carrageenan/kaolin-induced arthritis in rats through LPAR2" [J. Ginseng Res. 47 (1) (January 2023) 168].

[This corrects the article DOI: 10.1016/j.jgr.2021.02.001.][This corrects the article DOI: 10.1016/j.jgr.2022.12.004.].

Atypical formations of gintonin lysophosphatidic acids as new materials and their beneficial effects on degenerative diseases.

Fresh ginseng is prone to spoilage due to its high moisture content. For long-term storage, most fresh ginsengs are dried to white ginseng (WG) or steamed for hours at high temperature/pressure and dried to form Korean Red ginseng (KRG). They are further processed for ginseng products when subjected to hot water extraction/concentration under pressure. These WG or KRG preparation processes affect ginsenoside compositions and also other ginseng components, probably during treatments like steaming and drying, to form diverse bioactive phospholipids. It is known that ginseng contains high amounts of gintonin lysophosphatidic acids (LPAs). LPAs are simple lipid-derived growth factors in animals and humans and act as exogenous ligands of six GTP-binding-protein coupled LPA receptor subtypes. LPAs play diverse roles ranging from brain development to hair growth in animals and humans. LPA-mediated signaling pathways involve various GTP-binding proteins to regulate downstream pathways like [Ca2+]i transient induction. Recent studies have shown that gintonin exhibits anti-Alzheimer's disease and anti-arthritis effects in vitro and in vivo mediated by gintonin LPAs, the active ingredients of gintonin, a ginseng-derived neurotrophin. However, little is known about how gintonin LPAs are formed in high amounts in ginseng compared to other herbs. This review introduces atypical or non-enzymatic pathways under the conversion of ginseng phospholipids into gintonin LPAs during steaming and extraction/concentration processes, which exert beneficial effects against degenerative diseases, including Alzheimer's disease and arthritis in animals and humans via LPA receptors.

Phytoceramide Alleviates the Carrageenan/Kaolin-Induced Arthritic Symptoms by Modulation of Inflammation.

Phytoceramide (Pcer) is found mainly in plants and yeast. It can be neuroprotective and immunostimulatory on various cell types. In this study, the therapeutic effect of Pcer was explored using the carrageenan/kaolin (C/K)-induced arthritis rat model and fibroblast-like synoviocytes (FLS). Pcer treatment (1, 10, and 30 mg/kg/day) were given to the arthritic rats for 6 days after disease induction. Weight distribution ration (WDR), knee thickness, squeaking score, serum levels of proinflammatory mediators, and histological analysis were measured and performed to evaluate arthritic symptoms in the rat model. In interleukin (IL)­1ß­stimulated FLS, proinflammatory mediators were measured after Pcer (1-30 µM) treatment. Arthritic symptoms in rats with Pcer treatment were significantly decreased at days 4 to 6 after C/K arthritis induction. Inflammation in the knee joints were also significantly decreased in rats with Pcer treatment. Furthermore, in IL-1ß­stimulated FLS, the expressions of proinflammatory mediators were also inhibited by Pcer. As shown by the results, Pcer has anti-arthritic effects in the C/K rat model and in synovial cells, suggesting that Pcer has the potential to be a useful agent in arthritis treatment.

Corrigendum to' Morphine dependence is attenuated by red ginseng extract and ginsenosides Rh2, Rg3, and compound K' [Journal of Ginseng Research Volume 40, Issue 4, October 2016, Pages 445-452].

[This corrects the article DOI: 10.1016/j.jgr.2016.08.006.].

Chrysanthemum morifolium and Its Bioactive Substance Enhanced the Sleep Quality in Rodent Models via Cl- Channel Activation.

Dried Chrysanthemum morifolium (Chry) flowers have been used in Korea as a traditional insomnia treatment. In this study, the sleep-promoting activity and improving sleep quality of Chry extract (ext) and its active substance linarin were analyzed by pentobarbital-induced sleep experiment in mice and electroencephalography (EEG), electromyogram (EMG) analysis in rats. In a dose-dependent manner, Chry ext and linarin promoted longer sleep duration in the pentobarbital-induced sleep test compared to pentobarbital-only groups at both hypnotic and subhypnotic doses. Chry ext administration also significantly improved sleep quality, as seen in the relative power of low-frequency (delta) waves when compared with the control group. Linarin increased Cl- uptake in the SH-SY5Y human cell line and chloride influx was reduced by bicuculline. After administration of Chry ext, the hippocampus, frontal cortex, and hypothalamus from rodents were collected and blotted for glutamic acid decarboxylase (GAD)65/67 and gamma-aminobutyric acid (GABA)A receptors subunit expression levels. The expression of α1-subunits, ß2-subunits, and GAD65/67 of the GABAA receptor was modulated in the rodent brain. In conclusion, Chry ext augments pentobarbital-induced sleep duration and enhances sleep quality in EEG waves. These effects might be due to the activation of the Cl- channel.

Corrigendum to "Gintonin regulates inflammation in human IL-1ß-stimulated fibroblast-like synoviocytes and carrageenan/kaolin-induced arthritis in rats through LPAR2"[J Ginseng Res 45(5) September 2021, 575-582].

[This corrects the article DOI: 10.1016/j.jgr.2021.02.001.].

Derivatives of 3, 4, 5-Trimethoxycinnamic Acid Ameliorate Stress-Induced Anxiety in Mice and Rats.

Stress is an overwhelming problem associated with neuronal damage leading to anxiety and depression. The compound 3, 4, 5-trimethoxycinnamic acid (TMCA) has shown anti-stress effects; however, its derivatives remained unknown for their anxiolytic properties. Here, therefore, we investigated derivatives of TMCA (dTMCA) for their anxiolytic effects using immobilization and electric shock-induced stress in rats. Derivatives of TMCA ameliorated anxiety in mice and rats revealed by extended period of time spent in the open arms of elevated plus maze. Stress-mediated repression of tyrosine hydroxylase (TH) protein expression in the amygdala regions of rat brain and dopamine levels in the PC12 cells was restored by two selected derivatives (TMCA#5 and TMCA#9). Unlike TH expression, stress-induced protein expression of phospho-extracellular signal-regulated kinase (pERK) was unaffected by both derivatives in rats. Given the preferential inhibitory activity of dTMCA on dopamine and serotonin receptors, serotonergic road map of cellular signaling could be their target for anxiolytic effects. Thus, dTMCA would be promising agents to prevent neuronal damage associated with rampant stressful conditions.

Modafinil inhibits K(Ca)3.1 currents and muscle contraction via a cAMP-dependent mechanism.

Modafinil has been used as a psychostimulant for the treatment of narcolepsy. However, its primary mechanism of action remains elusive. Therefore, we examined the effects of modafinil on K(Ca)3.1 channels and vascular smooth muscle contraction. K(Ca)3.1 currents and channel activity were measured using a voltage-clamp technique and inside-out patches in mouse embryonic fibroblast cell line, NIH-3T3 fibroblasts. Intracellular adenosine 3',5'-cyclic monophosphate (cAMP) concentration was measured, and the phosphorylation of K(Ca)3.1 channel protein was examined using western blotting in NIH-3T3 fibroblasts and/or primary cultured mouse aortic smooth muscle cells (SMCs). Muscle contractions were recorded from mouse aorta and rat pulmonary artery by using a myograph developed in-house. Modafinil was found to inhibit K(Ca)3.1 currents in a concentration-dependent manner, and the half-maximal inhibition (IC(50)) of modafinil for the current inhibition was 6.8 ± 0.7 nM. The protein kinase A (PKA) activator forskolin also inhibited K(Ca)3.1 currents. The inhibitory effects of modafinil and forskolin on K(Ca)3.1 currents were blocked by the PKA inhibitors PKI(14-22) or H-89. In addition, modafinil relaxed blood vessels (mouse aorta and rat pulmonary artery) in a concentration-dependent manner. Modafinil increased cAMP concentrations in NIH-3T3 fibroblasts or primary cultured mouse aortic SMCs and phosphorylated K(Ca)3.1 channel protein in NIH-3T3 fibroblasts. However, open probability and single-channel current amplitudes of K(Ca)3.1 channels were not changed by modafinil. From these results, we conclude that modafinil inhibits K(Ca)3.1 channels and vascular smooth muscle contraction by cAMP-dependent phosphorylation, suggesting that modafinil can be used as a cAMP-dependent K(Ca)3.1 channel blocker and vasodilator.