RESUMO
The objective of this project was to find a bronchodilatory compound from herbs and clarify the mechanism. We found that the ethanol extract of Folium Sennae (EEFS) can relax airway smooth muscle (ASM). EEFS inhibited ASM contraction, induced by acetylcholine, in mouse tracheal rings and lung slices. High-performance liquid chromatography assay showed that EEFS contained emodin. Emodin had a similar reversal action. Acetylcholine-evoked contraction was also partially reduced by nifedipine (a selective inhibitor of L-type voltage-dependent Ca2+ channels, LVDCCs), YM-58483 (a selective inhibitor of store-operated Ca2+ entry, SOCE), as well as Y-27632 (an inhibitor of Rho-associated protein kinase). In addition, LVDCC- and SOCE-mediated currents and cytosolic Ca2+ elevations were inhibited by emodin. Emodin reversed acetylcholine-caused increases in phosphorylation of myosin phosphatase target subunit 1. Furthermore, emodin, in vivo, inhibited acetylcholine-induced respiratory system resistance in mice. These results indicate that EEFS-induced relaxation results from emodin inhibiting LVDCC, SOCE, and Ca2+ sensitization. These findings suggest that Folium Sennae and emodin may be new sources of bronchodilators.
Assuntos
Emodina/farmacologia , Contração Muscular/efeitos dos fármacos , Músculo Liso/efeitos dos fármacos , Acetilcolina/efeitos adversos , Acetilcolina/farmacologia , Animais , Broncodilatadores/metabolismo , Broncodilatadores/farmacologia , Pulmão/efeitos dos fármacos , Pulmão/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Contração Muscular/fisiologia , Músculo Liso/metabolismo , Fosfatase de Miosina-de-Cadeia-Leve/metabolismo , Fosfatase de Miosina-de-Cadeia-Leve/fisiologia , Extratos Vegetais/farmacologia , Senna/metabolismoRESUMO
The purpose of this study was to screen a bronchodilator from old drugs and elucidate the underlying mechanism. Paracetamol (acetaminophen) is a widely used analgesic and antipyretic drug. It has been reported that it inhibits the generation of prostaglandin and histamine, which play roles in asthma. These findings led us to explore whether paracetamol could be a potential bronchodilator. Paracetamol inhibited high K+- and acetylcholine (ACH)-induced precontraction of mouse tracheal and bronchial smooth muscles. Moreover, the ACH-induced contraction was partially inhibited by nifedipine (selective blocker of LVDCCs), YM-58483 (selective inhibitor of store-operated Ca2+ entry (SOCE), canonical transient receptor potential 3 (TRPC3) and TRPC5 channels) and Y-27632 (selective blocker of ROCK, a linker of the Ca2+ sensitization pathway). In single airway smooth muscle cells, paracetamol blocked the currents sensitive to nifedipine and YM-58483, and inhibited intracellular Ca2+ increases. In addition, paracetamol inhibited ACH-induced phosphorylation of myosin phosphatase target subunit 1 (MYPT1, another linker of the Ca2+ sensitization pathway). Finally, in vivo paracetamol inhibited ACH-induced increases of mouse respirator system resistance. Collectively, we conclude that paracetamol inhibits ASM contraction through blocking LVDCCs, SOCE and/or TRPC3 and/or TRPC5 channels, and Ca2+ sensitization. These results suggest that paracetamol might be a new bronchodilator.
Assuntos
Acetaminofen/farmacologia , Antipiréticos/farmacologia , Asma/metabolismo , Canais de Cálcio/metabolismo , Sinalização do Cálcio/efeitos dos fármacos , Miócitos de Músculo Liso/metabolismo , Acetilcolina/química , Acetilcolina/farmacologia , Animais , Asma/tratamento farmacológico , Brônquios/efeitos dos fármacos , Bloqueadores dos Canais de Cálcio/farmacologia , Permeabilidade da Membrana Celular/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos BALB C , Contração Muscular/efeitos dos fármacos , Músculo Liso/efeitos dos fármacos , Músculo Liso/metabolismo , Nifedipino/farmacologia , Potássio/metabolismoRESUMO
Azithromycin (AZM) has been used for the treatment of asthma and chronic obstructive pulmonary disease (COPD); however, the effects and underlying mechanisms of AZM remain largely unknown. The effects of AZM on airway smooth muscles (ASMs) and the underlying mechanisms were studied using isometric muscle force measurements, the examination of lung slices, imaging, and patch-clamp techniques. AZM completely inhibited acetylcholine (ACH)-induced precontraction of ASMs in animals (mice, guinea pigs, and rabbits) and humans. Two other macrolide antibiotics, roxithromycin and Klaricid, displayed a decreased inhibitory activity, and the aminoglycoside antibiotics penicillin and streptomycin did not have an inhibitory effect. Precontractions were partially inhibited by nifedipine (selective inhibitor of L-type voltage-dependent Ca2+ channels (LVDCCs)), Pyr3 (selective inhibitor of TRPC3 and/or STIM/Orai channels, which are nonselective cation channels (NSCCs)), and Y-27632 (selective inhibitor of Rho-associated kinase (ROCK)). Moreover, LVDCC- and NSCC-mediated currents were inhibited by AZM, and the latter were suppressed by the muscarinic (M) 2 receptor inhibitor methoctramine. AZM inhibited LVDCC Ca2+ permeant ion channels, M2 receptors, and TRPC3 and/or STIM/Orai, which decreased cytosolic Ca2+ concentrations and led to muscle relaxation. This relaxation was also enhanced by the inhibition of Ca2+ sensitization. Therefore, AZM has potential as a novel and potent bronchodilator. The findings of this study improve the understanding of the effects of AZM on asthma and COPD.
RESUMO
BACKGROUND/AIMS: Tetraethylammonium chloride (TEA) induces oscillatory contractions in mouse airway smooth muscle (ASM); however, the generation and maintenance of oscillatory contractions and their role in ASM are unclear. METHODS: In this study, oscillations of ASM contraction and intracellular Ca2+ were measured using force measuring and Ca2+ imaging technique, respectively. TEA, nifedipine, niflumic acid, acetylcholine chloride, lithium chloride, KB-R7943, ouabain, 2-Aminoethoxydiphenyl borate, thapsigargin, tetrodotoxin, and ryanodine were used to assess the mechanism of oscillatory contractions. RESULTS: TEA induced depolarization, resulting in activation of L-type voltage-dependent Ca2+ channels (LVDCCs) and voltage-dependent Na+ (VNa) channels. The former mediated Ca2+ influx to trigger a contraction and the latter mediated Na+ entry to enhance the contraction via activating LVDCCs. Meanwhile, increased Ca2+-activated Cl- channels, inducing depolarization that resulted in contraction through LVDCCs. In addition, the contraction was enhanced by intracellular Ca2+ release from Ca2+ stores mediated by inositol (1,4,5)-trisphosphate receptors (IP3Rs). These pathways together produce the contractile phase of the oscillatory contractions. Furthermore, the increased Ca2+ activated the Na+-Ca2+ exchanger (NCX), which transferred Ca2+ out of and Na+ into the cells. The former induced relaxation and the latter activated Na+/K+-ATPase that induced hypopolarization to inactivate LVDCCs causing further relaxation. This can also explain the relaxant phase of the oscillatory contractions. Moreover, the depolarization induced by VNa channels and NCX might be greater than the hypopolarization caused by Na+/K+-ATPase alone, inducing LVDCC activation and resulting in further contraction. CONCLUSIONS: These data indicate that the TEA-induced oscillatory contractions were cooperatively produced by LVDCCs, VNa channels, Ca2+-activated Cl- channels, NCX, Na+/K+ ATPase, IP3Rs-mediated Ca2+ release, and extracellular Ca2+.
Assuntos
Relógios Biológicos/efeitos dos fármacos , Canais de Cálcio Tipo L/metabolismo , Sinalização do Cálcio/efeitos dos fármacos , Contração Muscular/efeitos dos fármacos , Músculo Liso/metabolismo , Tetraetilamônio/farmacologia , Traqueia/metabolismo , Animais , Masculino , Potenciais da Membrana/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos BALB CRESUMO
This study aimed to elucidate the mechanisms of nuciferine (a main aporphine alkaloid of lotus leaf extract), which can induce relaxation in contracted tracheal rings. Under Ca2+-free and 2 mM Ca2+ conditions, we found that nuciferine had no effect on the resting muscle tone of tracheal rings. In contrast, nuciferine relaxed high K+-contracted mouse tracheal rings in a dose-dependent manner and inhibited both Ca2+ influx and voltage-dependent L-type Ca2+ channel currents induced by high K+. Similarly, nuciferine also inhibited acetylcholine-induced contractions in mouse tracheal rings in a dose-dependent manner. Meanwhile, both acetylcholine-induced intracellular Ca2+ influx and whole-cell currents of nonselective cation channels were blocked by nuciferine. Together, the results indicate that nuciferine-induced relaxation in tracheal rings mainly occurred due to the inhibition of extracellular Ca2+ influx through the blockade of voltage-dependent L-type Ca2+ channels and/or nonselective cation channels. These results suggest that nuciferine has a therapeutic effect on respiratory diseases associated with the aberrant contraction of airway smooth muscles and/or bronchospasm.
Assuntos
Aporfinas/farmacologia , Medicamentos de Ervas Chinesas/farmacologia , Relaxamento Muscular/efeitos dos fármacos , Músculo Liso/efeitos dos fármacos , Nelumbo/química , Parassimpatolíticos/farmacologia , Traqueia/efeitos dos fármacos , Acetilcolina/metabolismo , Animais , Aporfinas/química , Aporfinas/metabolismo , Cálcio/metabolismo , Canais de Cálcio/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Estrutura Molecular , Músculo Liso/metabolismoRESUMO
The effects of hypertonic solution on airway smooth muscle (ASM) contraction and the underlying mechanisms are largely unknown. We found that hypertonic saline (HS) inhibited acetylcholine (ACh)-induced contraction of ASM from the mouse trachea and human bronchi. In single mouse ASM cells (ASMCs), ACh induced an increase in intracellular Ca2+ that was further enhanced by 5% NaCl, indicating that the HS-induced inhibition of ASM contraction was not mediated by a decrease in cytosolic Ca2+ . The Rho-associated kinase (ROCK) inhibitor Y-27632 relaxed ACh-induced precontraction of mouse tracheal rings. However, such inhibition was not observed after the relaxation induced by 5% NaCl. Moreover, the incubation of mouse tracheal rings with 5% NaCl decreased ACh-induced phosphorylation of myosin light chain 20 and myosin phosphatase target subunit 1. These data indicate that HS inhibits the contraction of ASM by inhibiting Ca2+ sensitization, not by decreasing intracellular Ca2+ .
Assuntos
Cálcio/metabolismo , Pulmão/fisiologia , Contração Muscular/efeitos dos fármacos , Músculo Liso/efeitos dos fármacos , Músculo Liso/fisiologia , Solução Salina Hipertônica/farmacologia , Acetilcolina/farmacologia , Animais , Asma/metabolismo , Asma/patologia , Asma/fisiopatologia , Humanos , Espaço Intracelular/efeitos dos fármacos , Espaço Intracelular/metabolismo , Pulmão/efeitos dos fármacos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Músculo Liso/citologia , Músculo Liso/metabolismoRESUMO
AIM: Ghrelin can act as a signal for meal initiation and play a role in the regulation of gastrointestinal (GI) motility via hypothalamic circuit. This study investigated the correlation between changes of hypothalamic ghrelin system and GI motility dysfunction and anorexia in rats with chronic renal failure (CRF). METHODS: Sprague-Dawley (SD) rats (male/female 1:1, 180 ± 20 g) were randomly classified into a CRF group and control group (n = 8 per group). 5/6 nephrectomy was used to construct the CRF model. When plasma creatinine concentration (PCr) and blood urea nitrogen (BUN) in the CRF group were twice higher than the normal, food intake (g/24 h) and gastrointestinal interdigestive myoelectric complex (IMC) were detected. Then all rats were killed for assessment of the mRNA expression of ghrelin and growth hormone secretagogue receptor (GHS-R) in hypothalamus using reverse transcription-polymerase chain reaction. Analysis of variance, Student-Newman-Keuls-q-test and Correlation Analysis were used to do statistical analysis. P < 0.05 was considered as statistically significant. RESULTS: Compared to the control group, the CRF group was obviously decreased in the food intake (g/24 h), the phase III duration and amplitude and the ghrelin and GHS-R expression in the hypothalamus (P < 0.05). There was a positive correlation between them (P < 0.05). CONCLUSION: Changes of ghrelin and GHS-R in the hypothalamus correlate with gastrointestinal motility dysfunction and anorexia in rats with CRF.
Assuntos
Anorexia/etiologia , Gastroenteropatias/etiologia , Motilidade Gastrointestinal , Grelina/metabolismo , Hipotálamo/metabolismo , Falência Renal Crônica/complicações , Uremia/etiologia , Animais , Anorexia/genética , Anorexia/metabolismo , Anorexia/fisiopatologia , Biomarcadores/sangue , Nitrogênio da Ureia Sanguínea , Creatinina/sangue , Modelos Animais de Doenças , Ingestão de Alimentos , Feminino , Gastroenteropatias/genética , Gastroenteropatias/metabolismo , Gastroenteropatias/fisiopatologia , Grelina/genética , Hipotálamo/fisiopatologia , Falência Renal Crônica/genética , Falência Renal Crônica/metabolismo , Falência Renal Crônica/fisiopatologia , Masculino , Complexo Mioelétrico Migratório , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Receptores de Grelina/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fatores de Tempo , Uremia/genética , Uremia/metabolismo , Uremia/fisiopatologiaRESUMO
BACKGROUND: Transforming growth factor-ß1 (TGF-ß1) is a polypeptide member of the transforming growth factor ß superfamily of cytokines and performs many cellular functions. Its overexpression may lead to renal fibrosis. AIM: This study planed to investigate the effects of TGF-ß1 on the cell cycle and phenotype of mesangial cells. METHODS: Rat mesangial cells were cultured together with different concentrations (0, 1, 2, 5, and 10 ng/mL) of TGF-ß1 for specified times from 0 min to 72 h. 0 ng/mL TGF-ß1 and 0 min served as controls. Cell cycles were assessed by flow cytometry and α-smooth muscle actin expression (α-SMA) protein expression by western blot analysis. All data were presented as Mean ± SD. Statistical analysis was performed by using one-way analysis of variance and correlation analysis. Results were considered significant at p < 0.05. RESULTS: After 15 min of co-culture with different concentrations of TGF-ß1, the percentage of mesangial cells in G0/G1 phase was significantly elevated compared to the control (p < 0.05). 12 h co-culture induced cell hyperplasia, 24 h co-culture obvious up-regulation of α-SMA (p < 0.01) and one or two cells' myofibroblast phenotype transition, and 36 h co-culture several cells' phenotype transition. Correlation analysis prompted that the TGF-ß1-induced premature aging was time-dependent (p < 0.01). CONCLUSION: TGF-ß1 may induce mesangial cells' premature senescence and myofibroblast-like phenotype transformation time-dependently, which may contribute to the development of early stage of glomerulosclerosis.
Assuntos
Ciclo Celular/efeitos dos fármacos , Senescência Celular/efeitos dos fármacos , Células Mesangiais/efeitos dos fármacos , Fator de Crescimento Transformador beta1/farmacologia , Actinas/metabolismo , Animais , Técnicas de Cultura de Células , Técnicas de Cocultura , Células Mesangiais/citologia , Células Mesangiais/fisiologia , Ratos , Fatores de TempoRESUMO
BACKGROUND/AIMS: Ghrelin plays a central role in the regulation of gastrointestinal (GI) motility. This study aimed to investigate the expression of ghrelin and growth hormone secretagogue receptor (GHSR) in the central nervous system of rats with chronic renal failure (CRF). METHODS: Sprague-Dawley rats (male, 180 ± 20 g, n = 24) were treated by 5/6 nephrectomy to construct CRF model. As their plasma creatinine concentration and blood urea nitrogen were maintained more than double the normal level for 2 weeks, they were killed for assessing the expression of ghrelin and GHSR in hypothalamus and hippocampus using immunohistochemistry and real-time polymerase chain reaction (RT-PCR). The rats (male, 180 ± 20 g, n = 24) treated by Sham operation served as a control. One-way analysis of variance and Student-Newman-Keuls q test were used to analyze group difference and a p-value of <0.05 was considered as statistically significant. RESULTS: Compared with the controls, the ghrelin and GHSR expression was obviously increased in the hippocampus (p < 0.05) but decreased in the hypothalamus of rats with CRF (p < 0.05). CONCLUSIONS: CRF was found to impact the expression of ghrelin and GHSR in hypothalamus and hippocampus. This might be associated with the CRF-induced GI motility dysfunction.
Assuntos
Grelina/metabolismo , Hipocampo/metabolismo , Hipotálamo/metabolismo , Falência Renal Crônica/metabolismo , Receptores de Grelina/metabolismo , Animais , Expressão Gênica , Grelina/genética , Imuno-Histoquímica , Masculino , RNA Mensageiro/isolamento & purificação , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
ETHNOPHARMACOLOGICAL RELEVANCE: Areca Thirteen Pill, also called Gao You-13 (GY-13), is a traditional Mongolian herbal formula and has been extensively used to treat depression in Mongolian areas, which belongs to Heyi disease in Mongolian medicine. Major depressive disorder is a serious psychiatric disease, only one-third of individuals with depression are responsive to current antidepressants in clinic. Growing attention has been attracted by traditional herbal medicines in fighting depression because they are considered safer alternatives to pharmacotherapy. AIM OF THE STUDY: To reveal the mechanism of GY-13 in the treatment of depression. MATERIALS AND METHODS: The rat depression model was established by chronic unpredictable mild stress (CUMS), and primary hippocampal neurons were used to construct a glutamate-induced excitotoxicity model. The antidepressant effect of GY-13 was then assessed by performing sucrose preference tests, open field tests, and body weight measurements on rats. The expression of cAMP and PKA, mRNA levels of brain-derived neurotrophic factor (BDNF) and cAMP response element binding protein (CREB), and hippocampal neuronal apoptosis were measured. RESULTS: The results indicate that GY-13 significantly improves depression-like behavior, rescues decreased cAMPï¼ PKA, recovers the mRNA levels of CREB and BDNF, and increases the proliferative activity of hippocampus. In addition, blockade of PKA reverses the effects of GY-13 treatment on CREB mRNA, BDNF mRNA levels. In vitro, GY-13 treatment increased hippocampal proliferative activity and attenuated Glu-induced apoptosis of hippocampal neurons as well as reduced CREB mRNA and BDNF mRNA expression levels. CONCLUSIONS: Our research demonstrated that GY-13 treatment exerted a potent antidepressant action via activation of cAMP/CREB/BDNF signaling pathway, promoting proliferation, and suppressing apoptosis. This research provides molecular biological ground for developing GY-13 into a potent alternative for the intervention of depression.
Assuntos
Fator Neurotrófico Derivado do Encéfalo , Transtorno Depressivo Maior , Animais , Antidepressivos/metabolismo , Antidepressivos/farmacologia , Antidepressivos/uso terapêutico , Areca , Fator Neurotrófico Derivado do Encéfalo/genética , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Depressão/metabolismo , Transtorno Depressivo Maior/tratamento farmacológico , Hipocampo , Medicina Tradicional da Mongólia , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Transdução de Sinais , Estresse Psicológico/tratamento farmacológicoRESUMO
Temporomandibular disorders (TMDs) predominantly affect reproductive female patients, with pain the most frequent complaint. Although estrogens are believed to play important roles in TMD pain, the mechanism underlying modulation of TMD pain by estrogens remains largely unknown. Accumulating evidence implies that the hippocampus is involved in sexual dimorphism of pain sensitivity. In this study, we investigated the hippocampal TRPV1 (transient receptor potential vanilloid 1) expression in ovariectomized rats that received 17-beta-estradiol substitution and found that 17-beta-estradiol enhanced the mechanical allodynia of inflamed temporomandibular joint (TMJ) induced by complete Freund's adjuvant. Real-time PCR and immunoblotting demonstrated that TMJ inflammation significantly induced hippocampal TRPV1 expression compared with the control group but failed to induce it in the ovariectomized rats that received no estradiol replacement. In addition, estradiol potentiated TMJ inflammation-induced hippocampal TRPV1 expression in a dose-dependent manner in the ovariectomized rats. In contrast, TRPV1 transcription in amygdala, prefrontal cortex, and thalamus was not affected by TMJ inflammation and estradiol. Immunostaining showed TRPV1 localized in the processes and cytoplasm of pyramidal neurons in CA1-CA3 regions of the hippocampus. Moreover, intrahippocampal injection of TRPV1 antagonists capsazepine and 5'-iodo-resiniferatoxin into the CA1 region of the hippocampus significantly attenuated allodynia of inflamed TMJ in both nonovariectomized and ovariectomized rats that received estradiol replacement. Our results suggested that hippocampal TRPV1 can modulate central pain processing and estradiol may contribute to the sexual dimorphism of TMD pain sensitivity through upregulation of TRPV1 expression in the hippocampus.
Assuntos
Estradiol/metabolismo , Hipocampo/metabolismo , Dor/metabolismo , Canais de Cátion TRPV/metabolismo , Transtornos da Articulação Temporomandibular/metabolismo , Articulação Temporomandibular/metabolismo , Animais , Transtorno Autístico , Encéfalo/efeitos dos fármacos , Encéfalo/metabolismo , Feminino , Adjuvante de Freund , Hipocampo/efeitos dos fármacos , Ovariectomia , Dor/induzido quimicamente , Dor/tratamento farmacológico , Estimulação Física , Células Piramidais/efeitos dos fármacos , Células Piramidais/metabolismo , Distribuição Aleatória , Ratos , Ratos Sprague-Dawley , Canais de Cátion TRPV/antagonistas & inibidores , Transtornos da Articulação Temporomandibular/induzido quimicamente , Transtornos da Articulação Temporomandibular/tratamento farmacológico , Regulação para CimaRESUMO
OBJECTIVE: To investigate the role of dietary capsaicin in activating transient receptor potential vanilloid 1 (TRPV1) and thus influencing the vascular dysfunction mediated by high-fat diet and the potential mechanisms. METHODS: A total of 80 male C57BL/6J mice aged 10 weeks were equally divided into four groups, in which the mice were fed with normal diet (ND), normal diet plus capsaicin (NC), high-fat diet (HD), or high-fat diet plus capsaicin (HC) for 20 weeks. Tail-cuff blood pressure (BP), vascular function of mice aortic rings, expressions of voltage-gated potassium-channel Kv1.4, RhoA and Rho kinase in aorta were examined. RESULTS: Compared with ND group, both nitroglycerin [(18.9 +/- 13)% vs. 100%, P < 0.01] and acetylcholine [(26 +/- 12)% vs. 100%, P < 0.01] induced vasorelaxation of aortic rings were significantly reduced in HD group. Both endothelium dependent and independent aortic rings vasorelaxation in HC group were significantly improved compared with that in HD group [acetylcholine: (69 +/- 15)%; nitroglycerin: (46.5 +/- 6)%, P < 0.05], but still reduced compared with that in ND group (P < 0.05, P < 0.01). High fat diet induced the expression of RhoA and Rho kinase. Dietary capsaicin down-regulated the expression of RhoA and Rho kinase but up-regulated the expression of Kv1.4 in aorta in mice fed with normal or high fat diet (all P < 0.05). CONCLUSION: Dietary capsaicin can ameliorate vasorelaxation dysfunction mediated by high-fat diet. The potential mechanisms may be related with TRPV1 activation, which in turn stimulates potassium channel and inhibits RhoA and Rho kinase in the vasculature.
Assuntos
Capsaicina/farmacologia , Dieta Hiperlipídica/efeitos adversos , Canais de Cátion TRPV/efeitos dos fármacos , Vasodilatação/efeitos dos fármacos , Animais , Aorta/efeitos dos fármacos , Aorta/metabolismo , Aorta/fisiologia , Endotélio Vascular/metabolismo , Endotélio Vascular/fisiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Vasodilatação/fisiologia , Quinases Associadas a rho/metabolismo , Proteína rhoA de Ligação ao GTP/metabolismoRESUMO
Recent evidences indicate the existence of an atypical D(1) dopamine receptor other than traditional D(1) dopamine receptor in the brain that mediates PI hydrolysis via activation of phospholipase C(beta) (PLC(beta)). To further understand the basic physiological function of this receptor in brain, the effects of a selective phosphoinositide (PI)-linked D(1) dopamine receptor agonist SKF83959 on cytosolic free calcium concentration ([Ca(2+)](i)) in cultured rat prefrontal cortical astrocytes were investigated by calcium imaging. The results indicated that SKF83959 caused a transient dose-dependent increase in [Ca(2+)](i). Application of D(1) receptor, but not D(2), alpha(1) adrenergic, 5-HT receptor, or cholinergic antagonist prevented SKF83959-induced [Ca(2+)](i) rise, indicating that activation of the D(1) dopamine receptor was essential for this response. Increase in [Ca(2+)](i) was a two-step process characterized by an initial increase in [Ca(2+)](i) mediated by release from intracellular stores, supplemented by influx through voltage-gated calcium channels, receptor-operated calcium channels, and capacitative Ca(2+) entry. Furthermore, SKF83959-stimulated increase in [Ca(2+)](i) was abolished following treatment with a PLC inhibitor. Overall, these results suggested that activation of D(1) receptor by SKF83959 mediates a dose-dependent mobilization of [Ca(2+)](i) via the PLC signaling pathway in cultured rat prefrontal cortical astrocytes.
Assuntos
Astrócitos/metabolismo , Sinalização do Cálcio , Fosfatidilinositóis/metabolismo , Córtex Pré-Frontal/citologia , Receptores de Dopamina D1/metabolismo , 2,3,4,5-Tetra-Hidro-7,8-Di-Hidroxi-1-Fenil-1H-3-Benzazepina/análogos & derivados , 2,3,4,5-Tetra-Hidro-7,8-Di-Hidroxi-1-Fenil-1H-3-Benzazepina/farmacologia , Animais , Astrócitos/efeitos dos fármacos , Astrócitos/enzimologia , Canais de Cálcio/metabolismo , Sinalização do Cálcio/efeitos dos fármacos , Células Cultivadas , Ativação Enzimática/efeitos dos fármacos , Espaço Extracelular/efeitos dos fármacos , Espaço Extracelular/metabolismo , Feminino , Inositol 1,4,5-Trifosfato/metabolismo , Espaço Intracelular/efeitos dos fármacos , Espaço Intracelular/metabolismo , Masculino , Ratos , Ratos Sprague-Dawley , Transdução de Sinais/efeitos dos fármacos , Fosfolipases Tipo C/metabolismoRESUMO
We tested the hypothesis that activation of transient receptor potential vanilloid type-1 (TRPV1) by capsaicin prevents adipogenesis. TRPV1 channels in 3T3-L1-preadipocytes and visceral adipose tissue from mice and humans were detected by immunoblotting and quantitative real-time RT-PCR. The effect of TRPV1 on cytosolic calcium was determined fluorometrically in 3T3-L1-preadipocytes and in human visceral fat tissue. Adipogenesis in stimulated 3T3-L1-preadipocytes was determined by oil red O-staining of intracellular lipid droplets, triglyceride levels, expression of peroxisome proliferator-activated receptor-gamma, and expression of fatty acid synthase. Long-term feeding experiments were undertaken in wild-type mice and TRPV1 knockout mice. We detected TRPV1 channels in 3T3-L1-preadipocytes and visceral adipose tissue from mice and humans. In vitro, the TRPV1 agonist capsaicin dose-dependently induced calcium influx and prevented the adipogenesis in stimulated 3T3-L1-preadipocytes. RNA interference knockdown of TRPV1 in 3T3-L1-preadipocytes attenuated capsaicin-induced calcium influx, and adipogenesis in stimulated 3T3-L1-preadipocytes was no longer prevented. During regular adipogenesis TRPV1 channels were downregulated which was accompanied by a significant and time-dependent reduction of calcium influx. Compared with lean counterparts in visceral adipose tissue from obese db/db and ob/ob mice, and from obese human male subjects we observed a reduced TRVP1 expression. The reduced TRPV1 expression in visceral adipose tissue from obese humans was accompanied by reduced capsaicin-induced calcium influx. The oral administration of capsaicin for 120 days prevented obesity in male wild type mice but not in TRPV1 knockout mice assigned to high fat diet. We conclude that the activation of TRPV1 channels by capsaicin prevented adipogenesis and obesity.
Assuntos
Capsaicina/farmacologia , Obesidade/prevenção & controle , Canais de Cátion TRPV/metabolismo , Células 3T3-L1 , Adipócitos/metabolismo , Adipogenia , Tecido Adiposo/metabolismo , Animais , Cálcio/metabolismo , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos NOD , Camundongos Knockout , Camundongos Obesos , RNA Interferente Pequeno/farmacologia , Células-Tronco/metabolismo , Canais de Cátion TRPV/agonistas , Canais de Cátion TRPV/deficiência , Canais de Cátion TRPV/efeitos dos fármacos , Canais de Cátion TRPV/genética , VíscerasRESUMO
Using the static chamber-GC technique, greenhouse gas (CO2, CH4, N2O) fluxes of sand dunes and meadow wetlands were measured in a typical sand dune-meadow cascade ecological zone of Horqin. The dynamics of the greenhouse gas fluxes and driving factors were analyzed. The results showed that soil CH4 flux underwent absorption during the growing season, with average CH4 fluxes of semi-mobile dunes and meadow wetlands were -52.7 µg·m-2·h-1 and -34.7 µg·m-2·h-1, respectively, ranging from -176.1 to 49.8 µg·m-2·h-1. The peak of CH4 absorption in the growing season occurred at August 22nd, 2017. In August and September, the months with heavy rainfall, the CH4 flux in meadow wetlands showed continuous emission, being significantly different from that in semi-mobile dunes. The peak of N2O flux during the growing season was at July 21st. The monthly average N2O flux in semi-mobile dunes was following the order of July > August > September > June > May. Soil temperature and moisture were the key factors affecting CO2 and CH4 fluxes, whereas the N2O flux was mainly affected by soil temperature. The soil temperature sensitivity (Q10) showed the sequence of semi-mobile dune (1.009) < meadow wetland (1.474). The water stress rendered the greenhouse gas fluxes in semi-mobile dunes being less sensitive to soil temperature change than that in meadow wetlands.
Assuntos
Ecossistema , Monitoramento Ambiental , Gases de Efeito Estufa/análise , Dióxido de Carbono , Metano , Óxido Nitroso , Solo/química , Áreas AlagadasRESUMO
AIMS: This study focused on investigating whether NS8593 reverses airway smooth muscle (ASM) contraction and the underlying mechanism. MAIN METHODS: ASM contraction in mouse tracheal rings and lung slices was measured. Currents mediated by voltage dependent Ca2+ channels (VDCCs) and ACH-activated channels were measured using the whole-cell patch-clamp technique in single tracheal smooth muscle cells (TSMCs). Intracellular Ca2+ level and cell length were measured using an LSM 700 laser confocal microscope and a Zen 2010 software. Mouse respiratory system resistance (Rrs) was assessed using a FlexiVent FX system. KEY FINDINGS: High K+ (80â¯mMâ¯K+) and ACH induced ASM contraction in mouse tracheal rings and lung slices, which was partially relaxed by nifedipine (blocker of L-type VDCCs, LVDCCs), YM-58483 (blocker of store-operated Ca2+ entry (SOCE), transient receptor potential C3 (TRPC3) and TRPC5 channels), respectively. However, the contraction was completely reversed by NS8593, whereas, slightly relaxed by formoterol. ACH activated inward currents, which displayed linear and reversed around 0â¯mV, indicating the currents were mediated by non-selective cation channels (NSCCs). Moreover, these currents were blocked by YM-58483. In addition, such currents were abolished by NS8593, implicating that NS8593 inhibits the same channels. Besides, NS8593 inhibited increases of intracellular Ca2+ and the associated cell shortening. Finally, NS8593 inhibited ACH-induced increases of mouse respirator system resistance (Rrs). SIGNIFICANCE: Our results indicate that NS8593 inhibits LVDCCs and NSCCs, resulting in decreases of intracellular Ca2+ and then leading to ASM relaxation. These data suggest that NS8593 might be a new bronchodilator.
Assuntos
1-Naftilamina/análogos & derivados , Asma/tratamento farmacológico , Canais de Cálcio Tipo L/química , Cálcio/metabolismo , Contração Muscular/efeitos dos fármacos , Relaxamento Muscular/efeitos dos fármacos , Músculo Liso/efeitos dos fármacos , 1-Naftilamina/farmacologia , Animais , Antialérgicos/farmacologia , Asma/induzido quimicamente , Asma/patologia , Canais de Cálcio Tipo L/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Músculo Liso/metabolismo , Músculo Liso/patologia , Ovalbumina/toxicidadeRESUMO
Alzheimer disease (AD) is an age-related neurodegenerative disorder. Many observations indicate that impaired redox regulation is implicated in AD with synaptic failure. The aim of the current investigation was to characterize the role of redox-active agents on long-term potentiation (LTP) in the CA1 region of rat hippocampal slices and to elucidate the molecular sequence of events leading to these changes. The results presented here indicate that the membrane-permeable oxidizing agent chloramine-T (CH-T) inhibits the induction of LTP, whereas the membrane-permeable reducing agent dithiothreitol (DTT) enhances the induction of LTP. In contrast, neither the membrane-impermeable oxidizing agent 5,5'-dithio-bis-(2-nitrobenzoic acid) (DTNB) nor the membrane-impermeable reducing agent tris-(2-carboxyethyl) phosphine (TCEP) can affect the induction of LTP. The inhibition of LTP by CH-T can be restored by pretreatment with DTT but not with TCEP, whereas the enhancement of LTP by DTT can be reversed by pretreatment with CH-T but not with DTNB. We also provide evidence that the CH-T-evoked inhibition of LTP is mediated via activation of glycogen synthase kinase-3beta (GSK-3beta), whereas the DTT-evoked enhancement of LTP is mediated via inactivation of GSK-3beta. These findings will benefit the understanding of the redox contribution to the mechanisms underlying synaptic plasticity and AD pathogenesis.
Assuntos
Quinase 3 da Glicogênio Sintase/metabolismo , Hipocampo/metabolismo , Potenciação de Longa Duração/fisiologia , Oxidantes/farmacologia , Substâncias Redutoras/farmacologia , Animais , Western Blotting , Potenciais Pós-Sinápticos Excitadores/efeitos dos fármacos , Potenciais Pós-Sinápticos Excitadores/fisiologia , Hipocampo/efeitos dos fármacos , Potenciação de Longa Duração/efeitos dos fármacos , Microeletrodos , Plasticidade Neuronal/efeitos dos fármacos , Plasticidade Neuronal/fisiologia , Técnicas de Cultura de Órgãos , Oxirredução/efeitos dos fármacos , Ratos , Ratos Sprague-DawleyRESUMO
Aconitine is the main bioactive ingredient of Aconitum plants, which are well-known botanical herbs in China. Aconitine is also notorious for its high cardiotoxicity, as it can induce life-threatening ventricular arrhythmias. Unfortunately, there are few effective antidotes to aconitine toxicity. This study aimed to evaluate the potent protective effects of the ingredients from V. baillonii on aconitine toxicity on H9c2 cell line. Cell viability was assessed by methylthiazoltetrazolium bromide (MTT). Intracellular Ca2+ concentration alteration and reactive oxygen species (ROS) generation were observed by confocal microscopy and flow cytometry, respectively. Cellular oxidative stress was analyzed by measuring malondialdehyde (MDA) and superoxide dismutase (SOD) levels. Mitochondrial membrane potential (ΔΨ) was determined using JC-1 kit. RT-PCR and Hoechst staining techniques were conducted to determine the levels of autophagy/apoptosis. The mRNA levels of dihydropyridine receptor (DHPR), ryanodine receptors (RyR2) and sarcoplasmic reticulum Ca2+-ATPase (SERCA) were measured by RT-PCR. We screened six components from V. baillonii, among which, sweroside exhibited the strongest protective effects on aconitine-induced cardiac toxicity. Sweroside suppressed the aconitine-induced mRNA expressions of NaV1.5 (encoded by SCN5A), RyR2 and DHPR, and reversed the aconitine-induced decrease in mRNA level of SERCA, thus preventing the aconitine-induced persistent intracellular Ca2+ accumulation and avoiding intracellular Ca2+ overload. We further found that sweroside restabilized the aconitine-disrupted mitochondrial membrane potential (ΔΨ) and reversed the aconitine-induced increase in the mRNA levels of cell autophagy-related factors (Beclin-1, Caspase-3, and LC3- II) in H9c2 cells. In the whole-animal experiments, we observed that sweroside (50 mg/kg) alleviated effectively aconitine-induced arrhythmias by analysis of electrocardiogram (ECG) recording in rats. Our results demonstrate that sweroside may protect cardiomyocytes from aconitine toxicity by maintaining intracellular Ca2+ homeostasis, restabilizing mitochondrial membrane potential (ΔΨ) and avoiding cell autophagy/apoptosis.
RESUMO
ß2-adrenoceptor agonists are commonly used as bronchodilators to treat obstructive lung diseases such as asthma and chronic obstructive pulmonary disease (COPD), however, they induce severe side effects. Therefore, developing new bronchodilators is essential. Herbal plants were extracted and the extracts' effect on airway smooth muscle (ASM) precontraction was assessed. The ethyl alcohol extract of semen cassiae (EESC) was extracted from Semen cassia. The effects of EESC on the ACh- and 80 mM K+-induced sustained precontraction in mouse and human ASM were evaluated. Ca2+ permeant ion channel currents and intracellular Ca2+ concentration were measured. HPLC analysis was employed to determine which compound was responsible for the EESC-induced relaxation. The EESC reversibly inhibited the ACh- and 80 mM K+-induced precontraction. The sustained precontraction depends on Ca2+ influx, and it was mediated by voltage-dependent L-type Ca2+ channels (LVDCCs), store-operated channels (SOCs), TRPC3/STIM/Orai channels. These channels were inhibited by aurantio-obtusin, one component of EESC. When aurantio-obtusin removed, EESC's action disappeared. In addition, aurantio-obtusin inhibited the precontraction of mouse and human ASM and intracellular Ca2+ increases. These results indicate that Semen cassia-contained aurantio-obtusin inhibits sustained precontraction of ASM via inhibiting Ca2+-permeant ion channels, thereby, which could be used to develop new bronchodilators.
RESUMO
Because of the serious side effects of the currently used bronchodilators, new compounds with similar functions must be developed. We screened several herbs and found that Polygonum aviculare L. contains ingredients that inhibit the precontraction of mouse and human airway smooth muscle (ASM). High K+-induced precontraction in ASM was completely inhibited by nifedipine, a selective blocker of L-type voltage-dependent Ca2+ channels (LVDCCs). However, nifedipine only partially reduced the precontraction induced by acetylcholine chloride (ACH). Additionally, the ACH-induced precontraction was partly reduced by pyrazole-3 (Pyr3), a selective blocker of TRPC3 and stromal interaction molecule (STIM)/Orai channels. These channel-mediated currents were inhibited by the compounds present in P. aviculare extracts, suggesting that this inhibition was mediated by LVDCCs, TRPC3 and/or STIM/Orai channels. Moreover, these channel-mediated currents were inhibited by quercetin, which is present in P. aviculare extracts. Furthermore, quercetin inhibited ACH-induced precontraction in ASM. Overall, our data indicate that the ethyl acetate fraction of P. aviculare and quercetin can inhibit Ca2+-permeant LVDCCs, TRPC3 and STIM/Orai channels, which inhibits the precontraction of ASM. These findings suggest that P. aviculare could be used to develop new bronchodilators to treat obstructive lung diseases such as asthma and chronic obstructive pulmonary disease.