Your browser doesn't support javascript.
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 37.984
Filtrar
1.
Life Sci ; 244: 117333, 2020 Mar 01.
Artículo en Inglés | MEDLINE | ID: mdl-31962132

RESUMEN

AIMS: Detect the antiarrhythmic effect of crotonoside (Cro). MAIN METHODS: We used whole-cell patch-clamp techniques to detect the effects of Cro on action potentials (APs) and transmembrane ion currents in isolated rabbit left ventricular myocytes. We also verified the effect of Cro on ventricular arrhythmias caused by aconitine in vivo. KEY FINDINGS: Cro reduced the maximum depolarization velocity (Vmax) of APs and shortened the action potential duration (APD) in a concentration-dependent manner, but it had no significant effect on the resting membrane potential (RMP) or action potential amplitude (APA). It also inhibited the peak sodium current (INa) and L-type calcium current (ICaL) in a concentration-dependent manner with half-maximal inhibitory concentrations (IC50) of 192 µmol/L and 159 µmol/L, respectively. However, Cro had no significant effects on the inward rectifier potassium current (IK1) or rapidly activating delayed rectifier potassium current (IKr). Sea anemone toxin II (ATX II) increased the late sodium current (INaL), but Cro abolished this effect. Moreover, Cro significantly abolished ATX II-induced early afterdepolarizations (EADs) and high extracellular Ca2+ concentration (3.6 mmol/L)-induced delayed afterdepolarizations (DADs). We also verified that Cro effectively delayed the onset time and reduced the incidence of ventricular arrhythmias caused by aconitine in vivo. SIGNIFICANCE: These results revealed that Cro effectively inhibits INa, INaL, and ICaL in ventricular myocytes. Cro has antiarrhythmic potential and thus deserves further study.


Asunto(s)
Guanina/farmacología , Miocitos Cardíacos/efectos de los fármacos , Potenciales de Acción/efectos de los fármacos , Animales , Antiarrítmicos/metabolismo , Antiarrítmicos/farmacología , Arritmias Cardíacas/fisiopatología , Calcio/metabolismo , Canales de Calcio/efectos de los fármacos , China , Femenino , Guanina/metabolismo , Ventrículos Cardíacos/metabolismo , Técnicas de Placa-Clamp/métodos , Conejos , Sodio/metabolismo , Canales de Sodio/efectos de los fármacos
2.
Life Sci ; 240: 117068, 2020 Jan 01.
Artículo en Inglés | MEDLINE | ID: mdl-31751583

RESUMEN

AIMS: Bradycardia contributes to tachy-brady arrhythmias or sinus arrest during heart failure (HF). Sinoatrial node (SAN) adenosine A1 receptors (ADO A1Rs) are upregulated in HF, and adenosine is known to exert negative chronotropic effects on the SAN. Here, we investigated the role of A1R signaling at physiologically relevant ADO concentrations on HF SAN pacemaker cells. MAIN METHODS: Dogs with tachypacing-induced chronic HF and normal controls (CTL) were studied. SAN tissue was collected for A1R and GIRK mRNA quantification. SAN cells were isolated for perforated patch clamp recordings and firing rate (bpm), slope of slow diastolic depolarization (SDD), and maximum diastolic potential (MDP) were measured. Action potentials (APs) and currents were recorded before and after addition of 1 and 10 µM ADO. To assess contributions of A1R and G protein-coupled Inward Rectifier Potassium Current (GIRK) to ADO effects, APs were measured after the addition of DPCPX (selective A1R antagonist) or TPQ (selective GIRK blocker). KEY FINDINGS: A1R and GIRK mRNA expression were significantly increased in HF. In addition, ADO induced greater rate slowing and membrane hyperpolarization in HF vs CTL (p < 0.05). DPCPX prevented ADO-induced rate slowing in CTL and HF cells. The ADO-induced inward rectifying current, IKado, was observed significantly more frequently in HF than in CTL. TPQ prevented ADO-induced rate slowing in HF. SIGNIFICANCE: An increase in A1R and GIRK expression enhances IKAdo, causing hyperpolarization, and subsequent negative chronotropic effects in canine chronic HF at relevant [ADO]. GIRK blockade may be a useful strategy to mitigate bradycardia in HF.


Asunto(s)
Agonistas del Receptor de Adenosina A1/farmacología , Adenosina/farmacología , Canales de Potasio Rectificados Internamente Asociados a la Proteína G/agonistas , Insuficiencia Cardíaca/fisiopatología , Frecuencia Cardíaca/efectos de los fármacos , Receptor de Adenosina A1/metabolismo , Nodo Sinoatrial/citología , Nodo Sinoatrial/efectos de los fármacos , Potenciales de Acción/efectos de los fármacos , Antagonistas del Receptor de Adenosina A1/farmacología , Animales , Venenos de Abeja/farmacología , Relojes Biológicos , Enfermedad Crónica , Perros , Femenino , Canales de Potasio Rectificados Internamente Asociados a la Proteína G/antagonistas & inhibidores , Canales de Potasio Rectificados Internamente Asociados a la Proteína G/efectos de los fármacos , Técnicas In Vitro , Masculino , Técnicas de Placa-Clamp , Bloqueadores de los Canales de Potasio/farmacología , Receptor de Adenosina A1/efectos de los fármacos , Xantinas/farmacología
3.
Life Sci ; 242: 117209, 2020 Feb 01.
Artículo en Inglés | MEDLINE | ID: mdl-31870776

RESUMEN

AIMS: Hypertension is an independent risk factor for atrial fibrillation (AF). However, the direct effect of hydrostatic pressure on atrial electrical remodeling is unclear. The present study investigated whether hydrostatic pressure is responsible for atrial electrical remodeling and addressed a potential role of inflammation in this pathology. MAIN METHODS: Whole-cell patch-clamp recordings and biochemical assays were used to study the regulation and expression of ion channels in left atrial appendages in patients with AF, spontaneously hypertensive rats (SHRs), and atrium-derived cells (HL-1 cells) exposed to standard (0 mmHg) and elevated (20, 40 mmHg) hydrostatic pressure. KEY FINDINGS: Both TNF-α and MIF were highly expressed in patients with AF and SHRs. AF inducibility in SHRs was higher after atrial burst pacing, accompanied by a decrease in the L-type calcium current (ICa,L), an increase in the transient outward K+ current (Ito) and ultra-rapid delayed rectifier K+ current (IKur), and a shortened action potential duration (APD), which could be inhibited by atorvastatin. Furthermore, exposure to elevated pressure was associated with electrical remodeling of the HL-1 cells. The peak current density of ICa,L was reduced, while Ito and IKur were increased. Moreover, the expression levels of Kv4.3, Kv1.5, TNF-α, and MIF were upregulated, while the expression of Cav1.2 was downregulated in HL-1 cells after treatment with high hydrostatic pressure (40 mmHg). Atorvastatin alleviated the electrical remodeling and increased inflammatory markers in HL-1 cells induced by high hydrostatic pressure. SIGNIFICANCE: Elevated hydrostatic pressure led to atrial electrical remodeling and increased AF susceptibility by upregulating inflammation.


Asunto(s)
Remodelación Atrial , Citocinas/metabolismo , Presión Hidrostática/efectos adversos , Adulto , Animales , Western Blotting , Femenino , Atrios Cardíacos/metabolismo , Humanos , Masculino , Persona de Mediana Edad , Miocitos Cardíacos/metabolismo , Técnicas de Placa-Clamp , Ratas Endogámicas SHR , Ratas Wistar , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Regulación hacia Arriba
4.
Chin J Physiol ; 62(5): 175-181, 2019.
Artículo en Inglés | MEDLINE | ID: mdl-31670280

RESUMEN

The substantia gelatinosa (SG) of the trigeminal subnucleus caudalis (Vc) is admitted as a pivotal site of integrating and regulating orofacial nociceptive inputs. Although citral (3,7-dimethyl-2,6-octadienal) is involved in antinociception, the action mechanism of citral on the SG neurons of the Vc has not been fully clarified yet. In this study, we examined the direct membrane effects of citral and how citral mediates responses on the SG neurons of the Vc in juvenile mice using a whole-cell patch-clamp technique. Under high chloride pipette solution, citral showed repeatable inward currents that persisted in the presence of tetrodotoxin, a voltage-gated Na+ channel blocker, and 6-cyano-7-nitro-quinoxaline-2,3-dione, a non-N-methyl-D-aspartate (NMDA) glutamate receptor antagonist, D-2-amino-5-phosphonopentanoic acid, an NMDA receptor antagonist. However, the citral-induced inward currents were partially blocked by picrotoxin, a gamma-aminobutyric acid (GABAA)-receptor antagonist, or by strychnine, a glycine receptor antagonist. Further, the citral-induced responses were almost blocked by picrotoxin with strychnine. We also found that citral exhibited additive effect with GABA-induced inward currents and glycine-induced inward currents were potentiated by citral. In addition, citral suppressed the firing activities by positive current injection on the SG neurons of the Vc. Taken together, these results demonstrate that citral has glycine- and/or GABA-mimetic actions and suggest that citral might be a potential target for orofacial pain modulation by the activation of inhibitory neurotransmission in the SG area of the Vc.


Asunto(s)
Sustancia Gelatinosa , Animales , Ratones , Monoterpenos , Neuronas , Técnicas de Placa-Clamp , Ratas Sprague-Dawley
5.
Nan Fang Yi Ke Da Xue Xue Bao ; 39(9): 1078-1082, 2019 Sep 30.
Artículo en Chino | MEDLINE | ID: mdl-31640967

RESUMEN

OBJECTIVE: To observe the effect of cinobufagin on transient outward potassium current (IA) in rat dorsal root ganglion cells of cancer-induced bone pain (CIBP) and explore the possible analgesic mechanism of cinobufagin. METHODS: Whole cell patch clamp technique was used to examine the effect of cionbufagin on IA in acutely isolated dorsal root ganglion (DRG) cells from normal SD rats and rats with bone cancer pain. RESULTS: The DRG cells from rats with CIBP showed obviously decreased IA current density, an activation curve shift to the right, and an inactivation curve shift to the left. Cinobufagin treatment significantly increased the IA current density and reversed the changes in the activation and inactivation curves in the DRG cells. CONCLUSIONS: IA current is decreased in DRG neurons from rats with CIBP. Cinobufagin can regulate the activation and inactivation of IA current in the DRG cells, which may be related to its analgesic mechanism.


Asunto(s)
Analgésicos/farmacología , Bufanólidos/farmacología , Dolor en Cáncer/tratamiento farmacológico , Ganglios Espinales/efectos de los fármacos , Canales de Potasio/metabolismo , Animales , Células Cultivadas , Técnicas de Placa-Clamp , Ratas , Ratas Sprague-Dawley
6.
Invest Ophthalmol Vis Sci ; 60(12): 4063-4073, 2019 09 03.
Artículo en Inglés | MEDLINE | ID: mdl-31560762

RESUMEN

Purpose: The balance of neuronal excitation and inhibition is important for proper retinal signaling. A previous report showed that diabetes selectively reduces light-evoked inhibition to the retinal dim light rod pathway, changing this balance. Here, changes in mechanisms of retinal inhibitory synaptic transmission after 6 weeks of diabetes are investigated. Methods: Diabetes was induced in C57BL/6J mice by three intraperitoneal injections of streptozotocin (STZ, 75 mg/kg), and confirmed by blood glucose levels more than 200 mg/dL. After 6 weeks, whole-cell voltage-clamp recordings of electrically evoked inhibitory postsynaptic currents from rod bipolar cells and light-evoked excitatory postsynaptic currents from A17-amacrine cells were made in dark-adapted retinal slices. Results: Diabetes shortened the timecourse of directly activated lateral GABAergic inhibitory amacrine cell inputs to rod bipolar cells. The timing of GABA release onto rod bipolar cells depends on a prolonged amacrine cell calcium signal that is reduced by slow calcium buffering. Therefore, the effects of calcium buffering with EGTA-acetoxymethyl ester (AM) on diabetic GABAergic signaling were tested. EGTA-AM reduced GABAergic signaling in diabetic retinas more strongly, suggesting that diabetic amacrine cells have reduced calcium signals. Additionally, the timing of release from reciprocal inhibitory inputs to diabetic rod bipolar cells was reduced, but the activation of the A17 amacrine cells responsible for this inhibition was not changed. Conclusions: These results suggest that reduced light-evoked inhibitory input to rod bipolar cells is due to reduced and shortened calcium signals in presynaptic GABAergic amacrine cells. A reduction in calcium signaling may be a common mechanism limiting inhibition in the retina.


Asunto(s)
Señalización del Calcio/fisiología , Retinopatía Diabética/metabolismo , Células Bipolares de la Retina/metabolismo , Células Amacrinas/metabolismo , Animales , Glucemia/metabolismo , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Tipo 1/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Técnicas de Placa-Clamp , Estimulación Luminosa , Estreptozocina , Transmisión Sináptica/fisiología , Ácido gamma-Aminobutírico/metabolismo
7.
Invest Ophthalmol Vis Sci ; 60(12): 3821-3829, 2019 09 03.
Artículo en Inglés | MEDLINE | ID: mdl-31529078

RESUMEN

Purpose: Gap junction channels exhibit connexin specific biophysical properties, including the selective intercellular passage of larger solutes, such as second messengers. Here, we have examined the cyclic nucleotide permeability of the lens connexins, which could influence events like epithelial cell division and differentiation. Methods: We compared the cAMP permeability through channels composed of Cx43, Cx46, or Cx50 using simultaneous measurements of junctional conductance and intercellular transfer. For cAMP detection, the recipient cells were transfected with a cAMP sensor gene, the cyclic nucleotide-modulated channel from sea urchin sperm (SpIH). cAMP was introduced via patch pipette into the cell of the pair that did not express SpIH. SpIH-derived currents were recorded from the other cell of a pair that expressed SpIH. cAMP permeability was also directly visualized in transfected cells using a chemically modified fluorescent form of the molecule. Results: cAMP transfer was observed for homotypic Cx43 channels over a wide range of junctional conductance. Homotypic Cx46 channels also transferred cAMP, but permeability was reduced compared with Cx43. In contrast, homotypic Cx50 channels exhibited extremely low permeability to cAMP, when compared with either Cx43, or Cx46. Conclusions: These data show that channels made from Cx43 and Cx46 result in the intercellular delivery of cAMP in sufficient quantity to activate cyclic nucleotide-modulated channels. The data also suggest that the greatly reduced cAMP permeability of Cx50 channels could play a role in the regulation of cell division in the lens.


Asunto(s)
Conexina 43/metabolismo , Conexinas/metabolismo , AMP Cíclico/metabolismo , Cristalino/metabolismo , Sistemas de Mensajero Secundario/fisiología , Colorantes Fluorescentes , Uniones Comunicantes/fisiología , Células HeLa , Humanos , Activación del Canal Iónico/fisiología , Técnicas de Placa-Clamp , Permeabilidad , Transfección
8.
Elife ; 82019 09 06.
Artículo en Inglés | MEDLINE | ID: mdl-31490124

RESUMEN

Voltage-gated potassium channels (Kvs) are gated by transmembrane voltage sensors (VS) that move in response to changes in membrane voltage. Kv10.1 or Eag1 also has three intracellular domains: PAS, C-linker, and CNBHD. We demonstrate that the Eag1 intracellular domains are not required for voltage-dependent gating but likely interact with the VS to modulate gating. We identified specific interactions between the PAS, CNBHD, and VS that modulate voltage-dependent gating and provide evidence that VS movement destabilizes these interactions to promote channel opening. Additionally, mutation of these interactions renders Eag1 insensitive to calmodulin inhibition. The structure of the calmodulin insensitive mutant in a pre-open conformation suggests that channel opening may occur through a rotation of the intracellular domains and calmodulin may prevent this rotation by stabilizing interactions between the VS and intracellular domains. Intracellular domains likely play a similar modulatory role in voltage-dependent gating of the related Kv11-12 channels.


Asunto(s)
Canales de Potasio Éter-A-Go-Go/química , Canales de Potasio Éter-A-Go-Go/metabolismo , Proteínas Mutantes/química , Proteínas Mutantes/metabolismo , Animales , Células CHO , Calmodulina/metabolismo , Cricetinae , Cricetulus , Canales de Potasio Éter-A-Go-Go/genética , Modelos Moleculares , Proteínas Mutantes/genética , Técnicas de Placa-Clamp , Conformación Proteica
9.
Int J Mol Sci ; 20(17)2019 Sep 03.
Artículo en Inglés | MEDLINE | ID: mdl-31484392

RESUMEN

Brain-derived neurotrophic factor (BDNF) has previously been shown to play an important role in glutamatergic synaptic plasticity in the amygdala, correlating with cued fear learning. While glutamatergic neurotransmission is facilitated by BDNF signaling in the amygdala, its mechanism of action at inhibitory synapses in this nucleus is far less understood. We therefore analyzed the impact of chronic BDNF depletion on GABAA-mediated synaptic transmission in BDNF heterozygous knockout mice (BDNF+/-). Analysis of miniature and evoked inhibitory postsynaptic currents (IPSCs) in the lateral amygdala (LA) revealed neither pre- nor postsynaptic differences in BDNF+/- mice compared to wild-type littermates. In addition, long-term potentiation (LTP) of IPSCs was similar in both genotypes. In contrast, facilitation of spontaneous IPSCs (sIPSCs) by norepinephrine (NE) was significantly reduced in BDNF+/- mice. These results argue against a generally impaired efficacy and plasticity at GABAergic synapses due to a chronic BDNF deficit. Importantly, the increase in GABAergic tone mediated by NE is reduced in BDNF+/- mice. As release of NE is elevated during aversive behavioral states in the amygdala, effects of a chronic BDNF deficit on GABAergic inhibition may become evident in response to states of high arousal, leading to amygdala hyper-excitability and impaired amygdala function.


Asunto(s)
Amígdala del Cerebelo/metabolismo , Factor Neurotrófico Derivado del Encéfalo/metabolismo , Potenciación a Largo Plazo/fisiología , Animales , Factor Neurotrófico Derivado del Encéfalo/genética , Femenino , Potenciación a Largo Plazo/genética , Ratones , Ratones Noqueados , Plasticidad Neuronal/genética , Plasticidad Neuronal/fisiología , Técnicas de Placa-Clamp , Transmisión Sináptica/genética , Transmisión Sináptica/fisiología , Ácido gamma-Aminobutírico/metabolismo
10.
BMC Biol ; 17(1): 63, 2019 08 15.
Artículo en Inglés | MEDLINE | ID: mdl-31412898

RESUMEN

BACKGROUND: Voltage-gated sodium (Nav) channels have traditionally been considered a trademark of excitable cells. However, recent studies have shown the presence of Nav channels in several non-excitable cells, such as astrocytes and macrophages, demonstrating that the roles of these channels are more diverse than was previously thought. Despite the earlier discoveries, the presence of Nav channel-mediated currents in the cells of retinal pigment epithelium (RPE) has been dismissed as a cell culture artifact. We challenge this notion by investigating the presence and possible role of Nav channels in RPE both ex vivo and in vitro. RESULTS: Our work demonstrates that several subtypes of Nav channels are found in human embryonic stem cell (hESC)-derived and mouse RPE, most prominently subtypes Nav1.4, Nav1.6, and Nav1.8. Whole cell patch clamp recordings from the hESC-derived RPE monolayers showed that the current was inhibited by TTX and QX-314 and was sensitive to the selective blockers of the main Nav subtypes. Importantly, we show that the Nav channels are involved in photoreceptor outer segment phagocytosis since blocking their activity significantly reduces the efficiency of particle internalization. Consistent with this role, our electron microscopy results and immunocytochemical analysis show that Nav1.4 and Nav1.8 accumulate on phagosomes and that pharmacological inhibition of Nav channels as well as silencing the expression of Nav1.4 with shRNA impairs the phagocytosis process. CONCLUSIONS: Taken together, our study shows that Nav channels are present in RPE, giving this tissue the capacity of fast electrical signaling. The channels are critical for the physiology of RPE with an important role in photoreceptor outer segment phagocytosis.


Asunto(s)
Fagocitosis/genética , Epitelio Pigmentado de la Retina/fisiología , Transducción de Señal/genética , Canales de Sodio/fisiología , Animales , Células Madre Embrionarias Humanas , Humanos , Ratones , Ratones Endogámicos C57BL , Técnicas de Placa-Clamp
11.
Br J Anaesth ; 123(4): 439-449, 2019 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-31383364

RESUMEN

BACKGROUND: Nerve growth factor (NGF) has been implicated in hyperalgesia by sensitising nociceptors. A role for NGF in modulating myocardial injury through ischaemic nociceptive signalling is plausible. We examined whether inhibition of spinal NGF attenuates myocardial ischaemia-reperfusion injury and explored the underlying mechanisms. METHODS: In adult rats, lentivirus-mediated short-hairpin RNA targeted at reducing NGF gene expression (NGF-shRNA) or a transient receptor potential vanilloid 1 (TRPV1) antagonist (capsazepine) was injected intrathecally before myocardial ischaemia-reperfusion. Infarct size (expressed as the ratio of area at risk) and risk of arrhythmias were quantified. Whole-cell clamp patch electrophysiology was used to record capsaicin currents in primary dorsal root ganglion neurones. The co-expression of substance P (SP) and calcitonin gene-related peptide (CGRP), plus activation of TRPV1, protein kinase B (Akt) and extracellular signal-regulated kinase (ERK) were also quantified. RESULTS: NGF levels increased by 2.95 (0.34)-fold in dorsal root ganglion and 2.12 (0.27)-fold in spinal cord after myocardial ischaemia-reperfusion injury. Intrathecal injection of NGF-shRNA reduced infarct area at risk from 0.58 (0.02) to 0.37 (0.02) (P<0.01) and reduced arrhythmia score from 3.67 (0.33) to 1.67 (0.33) (P<0.01). Intrathecal capsazepine was similarly cardioprotective. NGF-shRNA suppressed expression of SP/CGRP and activation of Akt/ERK and TRPV1 in spinal cord. NGF increased capsaicin current amplitude from 144 (42) to 840 (132) pA (P<0.05), which was blocked by the TRPV1 antagonist 5'-iodoresiniferatoxin. Exogenous NGF enhanced capsaicin-induced Akt/ERK and TRPV1 activation in PC12 neuroendocrine tumour cells in culture. CONCLUSIONS: Spinal NGF contributes to myocardial ischaemia-reperfusion injury by mediating nociceptive signal transmission.


Asunto(s)
Terapia Genética/métodos , Lentivirus/genética , Daño por Reperfusión Miocárdica/genética , Daño por Reperfusión Miocárdica/prevención & control , Factor de Crecimiento Nervioso/genética , ARN Interferente Pequeño/administración & dosificación , ARN Interferente Pequeño/uso terapéutico , Animales , Arritmias Cardíacas/prevención & control , Capsaicina/análogos & derivados , Capsaicina/farmacología , Cardiotónicos/administración & dosificación , Cardiotónicos/uso terapéutico , Ganglios Espinales/efectos de los fármacos , Ganglios Espinales/metabolismo , Inyecciones Espinales , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Infarto del Miocardio/diagnóstico por imagen , Infarto del Miocardio/prevención & control , Factor de Crecimiento Nervioso/biosíntesis , Células PC12 , Técnicas de Placa-Clamp , Ratas , Ratas Sprague-Dawley , Canales Catiónicos TRPV/antagonistas & inhibidores , Canales Catiónicos TRPV/metabolismo
12.
J Enzyme Inhib Med Chem ; 34(1): 1465-1473, 2019 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-31411081

RESUMEN

In this investigation, we studied a family of compounds with an oxathiazolidine-4-one-2,2-dioxide skeleton and their amide synthetic precursors as new anticonvulsant drugs. The cyclic structures were synthesized using a three-step protocol that include solvent-free reactions and microwave-assisted heating. The compounds were tested in vivo through maximal electroshock seizure test in mice. All the structures showed activity at the lower doses tested (30 mg/Kg) and no signs of neurotoxicity were detected. Compound encoded as 1g displayed strong anticonvulsant effects in comparison with known anticonvulsants (ED50 = 29 mg/Kg). First approximations about the mechanisms of action of the cyclic structures were proposed by docking simulations and in vitro assays against sodium channels (patch clamp methods).


Asunto(s)
Anticonvulsivantes/química , Anticonvulsivantes/farmacología , Diseño de Drogas , Imidas/química , Imidas/farmacología , Tiazoles/química , Animales , Anticonvulsivantes/administración & dosificación , Anticonvulsivantes/síntesis química , Espectroscopía de Resonancia Magnética con Carbono-13 , Relación Dosis-Respuesta a Droga , Células HEK293 , Humanos , Imidas/síntesis química , Masculino , Ratones , Canal de Sodio Activado por Voltaje NAV1.1/efectos de los fármacos , Óxidos/química , Técnicas de Placa-Clamp , Espectroscopía de Protones por Resonancia Magnética
13.
Gen Physiol Biophys ; 38(5): 399-406, 2019 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-31411570

RESUMEN

The substantia gelatinosa of the trigeminal subnucleus caudalis has been considered to be an essential location for the transference of orofacial sensory signals. The co-localization of inhibitory and excitatory neurotransmitters in the same substantia gelatinosa (SG) neurons has demonstrated their essential part in the modification of nociceptive transmission. Zn2+ is particularly numerous in the mammalian central nervous system. There are proofs demonstrating the role of Zn2+ in the modulation of voltage- and ligand-gated ion channels. However, little is known about what roles Zn2+ may play in the modulation of signal transmission in the SG neurons of the trigeminal subnucleus caudalis (Vc). Therefore, in this study, we used the whole-cell patch clamp technique to find out the effect of Zn2+ on the responses of three main neurotransmitters (glycine, GABA, and glutamate) on SG neurons of the Vc in mice. We have proved that Zn2+ induces a big potentiation of glycine receptor-mediated response but attenuates GABA- and glutamate-induced responses at micromolar concentrations, however, enhances glutamate-induced response at nanomolar concentration. Taken together, these data demonstrated that Zn2+ can modulate glycine, GABA and glutamate-mediated actions on the SG neurons of the Vc and support an important mechanism in spinal sensory information signaling.


Asunto(s)
Neuronas/efectos de los fármacos , Sustancia Gelatinosa/citología , Transmisión Sináptica/efectos de los fármacos , Zinc/farmacología , Animales , Ácido Glutámico/metabolismo , Glicina/metabolismo , Ratones , Neurotransmisores/metabolismo , Técnicas de Placa-Clamp , Ratas Sprague-Dawley , Receptores de Glicina/metabolismo , Ácido gamma-Aminobutírico/metabolismo
14.
Biophys Chem ; 254: 106246, 2019 11.
Artículo en Inglés | MEDLINE | ID: mdl-31426023

RESUMEN

The inhibitory effect of the flavonoid naringenin on plant and human Two-Pore Channels (TPCs) was assessed by means of electrophysiological measurements. By acting on human TPC2, naringenin, was able to dampen intracellular calcium responses to VEGF in cultured human endothelial cells and to impair angiogenic activity in VEGF-containing matrigel plugs implanted in mice. Molecular docking predicts selective binding sites for naringenin in the TPC structure, thus suggesting a specific interaction between the flavonoid and the channel.


Asunto(s)
Canales de Calcio/química , Flavanonas/química , Plantas/metabolismo , Animales , Sitios de Unión , Calcio/química , Calcio/metabolismo , Canales de Calcio/metabolismo , Colágeno/química , Combinación de Medicamentos , Células Endoteliales/citología , Células Endoteliales/metabolismo , Flavanonas/metabolismo , Humanos , Laminina/química , Ratones , Simulación del Acoplamiento Molecular , Técnicas de Placa-Clamp , Proteínas de Plantas/antagonistas & inhibidores , Proteínas de Plantas/metabolismo , Proteoglicanos/química
15.
Zhonghua Xin Xue Guan Bing Za Zhi ; 47(8): 608-613, 2019 Aug 24.
Artículo en Chino | MEDLINE | ID: mdl-31434431

RESUMEN

Objective: To investigate the effects and mechanism of digoxin on atrium electrical remodeling and susceptibility of atrial fibrillation (AF) in aged rabbits. Methods: Twenty aged male New Zealand rabbits were divided into aged group and aged plus digoxin group (n=10 each). Electrical parameters including heart rate (HR), RR and QT interval, ST segment and P wave dispersion from normal Ⅱ electrocardiogram, and the maximum upstroke velocity (Max(dv/dt)), plateau potential (plateau P), action potential duration of 10%, 20% and 90% (APD(10), APD(20), APD(90)) from recording of monophasic action potential (MAP), as well as atrial effective refractory period (AERP(200)) and dispersion (dERP(200)) with 200 ms of basic cycle length (BCL), and frequency self adaptation of AERP with 300 ms and 150 ms of BCLs (fERP) were recorded and compared between the 2 groups. BCLs and inducibility of AF post programmed electrical stimulation and Burst-pacing in left atrium tissue of rabbits in vivo were also analyzed. The L-type calcium current (I(Ca-L)) in 2 groups were recorded via whole-cell patch clamp technique, and the fluorescence intensity of intracellular free Ca(2+) was detected with Flup-3/AM loading by the laser scanning confocal microscope in enzymatically dissociated single rabbit atrial myocytes. Results: Compared with aged group, the heart rate was faster, RR and QT interval were obvious shorter, ST segment was raised and P wave dispersion was significantly increased in aged plus digoxin group (all P<0.05). Moreover, compared with aged group, the Max(dv/dt) and plateau P were obviously increased, APD(10) and APD(20) were significantly prolongated, and APD(9)0 was significantly shorter in aged plus digoxin group (all P<0.01). Otherwise, the fERP was markedly increased (0.81±0.15 vs. 0.67±0.05), and the induced rate of AF was obviously higher in aged plus digoxin group than in aged group (6/8 vs. 4/9) (all P<0.01). With voltage clamp model, digoxin significantly increased I(Ca-L) of atrial myocytes of aged rabbits, When command potential was 10 mV, the current densities of I(Ca-L) were significantly higher in digoxin group than that in aged group ((15.45±2.38) pA/pF vs. (7.03±1.69) pA/pF, P<0.01). Otherwise, the I-V curve of I(Ca-L) was downward shifted of all I-V curves in digoxin perfused aged atrial cells of rabbits. Moreover, the fluorescence intensities of intracellular free Ca(2+) was significantly higher in aged plus digoxin group than in aged group ((1 748±173) µmol/L vs. (478.13±87.63) µmol/L, P<0.01). Conclusion: Digoxin could aggravate the atrial electrical remodeling in atrium of aged rabbits, facilitate susceptibility of atrial fibrillation in aged rabbit, increased current density of I(Ca-L) and concentration of intracellular free Ca(2+), followed Ca(2+) overload and oscillations might be part of the underlying mechanisms.


Asunto(s)
Fibrilación Atrial , Remodelación Atrial , Potenciales de Acción , Animales , Digoxina , Atrios Cardíacos , Masculino , Técnicas de Placa-Clamp , Conejos
16.
Nat Commun ; 10(1): 3740, 2019 08 20.
Artículo en Inglés | MEDLINE | ID: mdl-31431622

RESUMEN

The transient receptor potential melastatin 2 (TRPM2) channel plays a key role in redox sensation in many cell types. Channel activation requires binding of both ADP-ribose (ADPR) and Ca2+. The recently published TRPM2 structures from Danio rerio in the ligand-free and the ADPR/Ca2+-bound conditions represent the channel in closed and open states, which uncovered substantial tertiary and quaternary conformational rearrangements. However, it is unclear how these rearrangements are achieved within the tetrameric channel during channel gating. Here we report the cryo-electron microscopy structures of Danio rerio TRPM2 in the absence of ligands, in complex with Ca2+ alone, and with both ADPR and Ca2+, resolved to ~4.3 Å, ~3.8 Å, and ~4.2 Å, respectively. In contrast to the published results, our studies capture ligand-bound TRPM2 structures in two-fold symmetric intermediate states, offering a glimpse of the structural transitions that bridge the closed and open conformations.


Asunto(s)
Adenosina Difosfato Ribosa/metabolismo , Calcio/metabolismo , Estructura Cuaternaria de Proteína , Canales Catiónicos TRPM/metabolismo , Proteínas de Pez Cebra/metabolismo , Animales , Línea Celular , Microscopía por Crioelectrón , Células HEK293 , Humanos , Activación del Canal Iónico , Técnicas de Placa-Clamp , Células Sf9 , Spodoptera , Canales Catiónicos TRPM/química , Pez Cebra , Proteínas de Pez Cebra/química
17.
Nat Commun ; 10(1): 3830, 2019 08 23.
Artículo en Inglés | MEDLINE | ID: mdl-31444362

RESUMEN

Brain activity and connectivity alter drastically during epileptic seizures. The brain networks shift from a balanced resting state to a hyperactive and hypersynchronous state. It is, however, less clear which mechanisms underlie the state transitions. By studying neural and glial activity in zebrafish models of epileptic seizures, we observe striking differences between these networks. During the preictal period, neurons display a small increase in synchronous activity only locally, while the gap-junction-coupled glial network was highly active and strongly synchronized across large distances. The transition from a preictal state to a generalized seizure leads to an abrupt increase in neural activity and connectivity, which is accompanied by a strong alteration in glia-neuron interactions and a massive increase in extracellular glutamate. Optogenetic activation of glia excites nearby neurons through the action of glutamate and gap junctions, emphasizing a potential role for glia-glia and glia-neuron connections in the generation of epileptic seizures.


Asunto(s)
Encéfalo/fisiopatología , Comunicación Celular , Excitabilidad Cortical/fisiología , Epilepsia/fisiopatología , Convulsiones/fisiopatología , Animales , Animales Modificados Genéticamente , Encéfalo/citología , Encéfalo/diagnóstico por imagen , Modelos Animales de Enfermedad , Uniones Comunicantes/fisiología , Ácido Glutámico/metabolismo , Humanos , Microscopía Confocal , Red Nerviosa/citología , Red Nerviosa/fisiopatología , Neuroglía/fisiología , Neuronas/fisiología , Imagen Óptica , Optogenética , Técnicas de Placa-Clamp , Pez Cebra
18.
Invest Ophthalmol Vis Sci ; 60(10): 3297-3309, 2019 08 01.
Artículo en Inglés | MEDLINE | ID: mdl-31369032

RESUMEN

Purpose: We investigate the contribution of TRPV1 and TRPV4 channels to retinal angiogenesis. Methods: Primary retinal microvascular endothelial cells (RMECs) were used for RT-PCR, Western blotting, immunolabeling, Ca2+ signaling, and whole-cell patch-clamp studies while localization of TRPV1 also was assessed in retinal endothelial cells using whole mount preparations. The effects of pharmacologic blockers of TRPV1 and TRPV4 on retinal angiogenic activity was evaluated in vitro using sprout formation, cell migration, proliferation, and tubulogenesis assays, and in vivo using the mouse model of oxygen-induced retinopathy (OIR). Heteromultimerization of TRPV1 and TRPV4 channels in RMECs was assessed using proximity ligation assays (PLA) and electrophysiologic recording. Results: TRPV1 mRNA and protein expression were identified in RMECs. TRPV1 labelling was found to be mainly localized to the cytoplasm with some areas of staining colocalizing with the plasma membrane. Staining patterns for TRPV1 were broadly similar in endothelial cells of intact vessels within retinal flat mounts. Functional expression of TRPV1 and TRPV4 in RMECs was confirmed by patch-clamp recording. Pharmacologic inhibition of TRPV1 or TRPV4 channels suppressed in vitro retinal angiogenesis through a mechanism involving the modulation of tubulogenesis. Blockade of these channels had no effect on VEGF-stimulated angiogenesis or Ca2+ signals in vitro. PLA and patch-clamp studies revealed that TRPV1 and TRPV4 form functional heteromeric channel complexes in RMECs. Inhibition of either channel reduced retinal neovascularization and promoted physiologic revascularization of the ischemic retina in the OIR mouse model. Conclusions: TRPV1 and TRPV4 channels represent promising targets for therapeutic intervention in vasoproliferative diseases of the retina.


Asunto(s)
Células Endoteliales/metabolismo , Neovascularización Retiniana/metabolismo , Vasos Retinianos/citología , Canales Catiónicos TRPV/fisiología , Animales , Animales Recién Nacidos , Western Blotting , Calcio/metabolismo , Señalización del Calcio/fisiología , Movimiento Celular/fisiología , Proliferación Celular/fisiología , Células Cultivadas , Ratones , Ratones Endogámicos C57BL , Oxígeno/toxicidad , Técnicas de Placa-Clamp , Piridinas/farmacología , ARN Mensajero/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Neovascularización Retiniana/patología , Sulfonamidas/farmacología , Sulfonas/farmacología , Canales Catiónicos TRPV/antagonistas & inhibidores , Factor A de Crecimiento Endotelial Vascular/farmacología
19.
PLoS Comput Biol ; 15(8): e1007226, 2019 08.
Artículo en Inglés | MEDLINE | ID: mdl-31381555

RESUMEN

We have previously shown that the physiological size of postsynaptic currents maximises energy efficiency rather than information transfer across the retinothalamic relay synapse. Here, we investigate information transmission and postsynaptic energy use at the next synapse along the visual pathway: from relay neurons in the thalamus to spiny stellate cells in layer 4 of the primary visual cortex (L4SS). Using both multicompartment Hodgkin-Huxley-type simulations and electrophysiological recordings in rodent brain slices, we find that increasing or decreasing the postsynaptic conductance of the set of thalamocortical inputs to one L4SS cell decreases the energy efficiency of information transmission from a single thalamocortical input. This result is obtained in the presence of random background input to the L4SS cell from excitatory and inhibitory corticocortical connections, which were simulated (both excitatory and inhibitory) or injected experimentally using dynamic-clamp (excitatory only). Thus, energy efficiency is not a unique property of strong relay synapses: even at the relatively weak thalamocortical synapse, each of which contributes minimally to the output firing of the L4SS cell, evolutionarily-selected postsynaptic properties appear to maximise the information transmitted per energy used.


Asunto(s)
Modelos Neurológicos , Transmisión Sináptica/fisiología , Tálamo/fisiología , Corteza Visual/fisiología , Potenciales de Acción/fisiología , Animales , Biología Computacional , Simulación por Computador , Metabolismo Energético/fisiología , Potenciales Postsinápticos Excitadores/fisiología , Técnicas In Vitro , Neuronas/fisiología , Técnicas de Placa-Clamp , Ratas , Ratas Sprague-Dawley , Tálamo/citología , Corteza Visual/citología , Vías Visuales/citología , Vías Visuales/fisiología
20.
Fitoterapia ; 137: 104272, 2019 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-31326417

RESUMEN

In the current study effects of fungal extracts on the G-protein-activated inwardly rectifying potassium channel (GIRK1/4) were screened using the automated patch-clamp method. 40 organic (n-hexane, chloroform, and 50% methanol) and aqueous extracts were prepared from 10 mushroom species native to Hungary. Among the examined fungal fractions of different polarities some n-hexane and chloroform extracts exerted considerable ion channel activity. One of the most active fungal species, Hypholoma lateritium was selected for further detailed examination to determine the compounds responsible for the observed pharmacological property. Evaluation of the ion channel activity of mushroom metabolites 1-10 revealed that lanosta-7,9(11)-diene-12ß,21α-epoxy-2α,3ß,24ß,25-tetraol (5) demonstrates remarkable blocking activity on GIRK current (IC50 395.1 ±â€¯31.8 nM). Investigation of the selectivity of the GIRK inhibitory effect proved that lanosta-7,9(11)-diene-12ß,21α-epoxy-2α,3ß,24ß,25-tetraol (5) has only weak inhibitory activity on hERG channel (7.9 ±â€¯2.8% at 100 µM), exerting more than three orders of magnitude lower blocking activity on hERG channel than on GIRK channel.


Asunto(s)
Agaricales/química , Canal de Potasio ERG1/antagonistas & inhibidores , Canales de Potasio Rectificados Internamente Asociados a la Proteína G/antagonistas & inhibidores , Células HEK293 , Humanos , Hungría , Estructura Molecular , Técnicas de Placa-Clamp
SELECCIÓN DE REFERENCIAS
DETALLE DE LA BÚSQUEDA