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OBJECTIVE@#To investigate the effect of ferulic acid, a natural compound, on pancreatic beta cell viability, Ca2+ channels, and insulin secretion.@*METHODS@#We studied the effects of ferulic acid on rat insulinoma cell line viability using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide viability assay. The whole-cell patch-clamp technique and enzyme-linked immunosorbent assay were also used to examine the action of ferulic acid on Ca2+ channels and insulin secretion, respectively.@*RESULTS@#Ferulic acid did not affect cell viability during exposures up to 72 h. The electrophysiological study demonstrated that ferulic acid rapidly and concentration-dependently increased L-type Ca2+ channel current, shifting its activation curve in the hyperpolarizing direction with a decreased slope factor, while the voltage dependence of inactivation was not affected. On the other hand, ferulic acid have no effect on T-type Ca2+ channels. Furthermore, ferulic acid significantly increased insulin secretion, an effect inhibited by nifedipine and Ca2+-free extracellular fluid, confirming that ferulic acid-induced insulin secretion in these cells was mediated by augmenting Ca2+ influx through L-type Ca2+ channel. Our data also suggest that this may be a direct, nongenomic action.@*CONCLUSION@#This is the first electrophysiological demonstration that acute ferulic acid treatment could increase L-type Ca2+ channel current in pancreatic β cells by enhancing its voltage dependence of activation, leading to insulin secretion.
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Rats , Animals , Insulin Secretion , Insulin/pharmacology , Insulin-Secreting Cells/metabolism , Coumaric Acids/metabolism , Calcium/metabolismABSTRACT
The lamina II, also called the substantia gelatinosa (SG), of the trigeminal subnucleus caudalis (Vc), is thought to play an essential role in the control of orofacial nociception. Glycine and serotonin (5-hydroxytryptamine, 5-HT) are the important neurotransmitters that have the individual parts on the modulation of nociceptive transmission. However, the electrophysiological effects of 5-HT on the glycine receptors on SG neurons of the Vc have not been well studied yet. For this reason, we applied the whole-cell patch clamp technique to explore the interaction of intracellular signal transduction between 5-HT and the glycine receptors on SG neurons of the Vc in mice. In nine of 13 neurons tested (69.2%), pretreatment with 5-HT potentiated glycine-induced current (I(Gly)). Firstly, we examined with a 5-HT₁ receptor agonist (8-OH-DPAT, 5-HT(1/7) agonist, co-applied with SB-269970, 5-HT₇ antagonist) and antagonist (WAY-100635), but 5-HT₁ receptor agonist did not increase IGly and in the presence of 5-HT₁ antagonist, the potentiation of 5-HT on I(Gly) still happened. However, an agonist (α-methyl-5-HT) and antagonist (ketanserin) of the 5-HT₂ receptor mimicked and inhibited the enhancing effect of 5-HT on I(Gly) in the SG neurons, respectively. We also verified the role of the 5-HT₇ receptor by using a 5-HT₇ antagonist (SB-269970) but it also did not block the enhancement of 5-HT on I(Gly). Our study demonstrated that 5-HT facilitated I(Gly) in the SG neurons of the Vc through the 5-HT₂ receptor. The interaction between 5-HT and glycine appears to have a significant role in modulating the transmission of the nociceptive pathway.
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Animals , Mice , Glycine , Neurons , Neurotransmitter Agents , Nociception , Patch-Clamp Techniques , Receptors, Glycine , Serotonin , Signal Transduction , Substantia GelatinosaABSTRACT
AIM: To investigate whether oxytocin has neuroprotective effects on hippocampal CA 1 pyramidal neurons from neonatal rats exposed to hypoxic-ischemic brain injury and the underlying mechanisms.METHODS:An in vitro model of hypoxic-ischemic injury was used by exposing the brain slices to oxygen-glucose deprivation(OGD)solution. Acute dissociated brain slices(6~8 slices per rat)from 8 Sprague-Dawely rats of 7~10 d old were used.The slices were randomly divided into 4 groups: control group, OGD 20 min group, OGD 40 min group and OGD +oxytocin group.The effect of oxytocin on neuronal death was evaluated by TO-PRO-3 staining.Fresh brain slices from other 20 neonatal rats were divided into OGD group,OGD+oxytocin group,OGD+dVOT(oxytocin receptor antagonist)+oxytocin group,and OGD+bicucuclline(GABAAreceptor antagonist)+oxytocin group.The onset of anoxic depolarization in the hippocampal neurons treated with different drugs was recorded by whole-cell patch-clamp techniques.RESULTS: The results of TO-PRO-3 staining showed that neuronal deaths in hippocampal CA 1 area were increased over the prolonged OGD time.Oxyto-cin significantly reduced the hypoxic-ischemic deaths.Oxytocin dramatically prolonged the onset time of anoxic depolariza-tion after the application of OGD solution.Both dVOT and bicuculline blocked this effect.CONCLUSION: Oxytocin plays a neuroprotective role in neonatal rat hippocampal CA 1 pyramidal neurons by enhancing the inhibitory synaptic trans-mission via oxytocin receptors.Therefore,oxytocin is useful as a candidate for neuroprotective treatment after neonatal hy -poxic-ischemic brain injury.
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It has been reported that Korean red ginseng (KRG), a valuable and important traditional medicine, has varied effects on the central nervous system, suggesting its activities are complicated. The paraventricular nucleus (PVN) neurons of the hypothalamus has a critical role in stress responses and hormone secretions. Although the action mechanisms of KRG on various cells and systems have been reported, the direct membrane effects of KRG on PVN neurons have not been fully described. In this study, the direct membrane effects of KRG on PVN neuronal activity were investigated by using a perforated patch-clamp in ICR mice. In gramicidin perforated patch-clamp mode, KRG extract (KRGE) induced repeatable depolarization followed by hyperpolarization of PVN neurons. The KRGE-induced responses were concentration-dependent and persisted in the presence of tetrodotoxin, a voltage sensitive Na+ channel blocker. The KRGE-induced responses were suppressed by 6-cyano-7-nitroquinoxaline-2,3-dione (10 µM), a non-N-methyl-D-aspartate (NMDA) glutamate receptor antagonist, but not by picrotoxin, a type A gamma-aminobutyric acid receptor antagonist. The results indicate that KRG activates non-NMDA glutamate receptors of PVN neurons in mice, suggesting that KRG may be a candidate for use in regulation of stress responses by controlling autonomic nervous system and hormone secretion.
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Animals , Mice , 6-Cyano-7-nitroquinoxaline-2,3-dione , Autonomic Nervous System , Central Nervous System , Glutamic Acid , Gramicidin , Hypothalamus , Medicine, Traditional , Membranes , Mice, Inbred ICR , Neurons , Panax , Paraventricular Hypothalamic Nucleus , Patch-Clamp Techniques , Picrotoxin , Receptors, GABA , Receptors, Glutamate , TetrodotoxinABSTRACT
The hypothalamic paraventricular nucleus (PVN) contains two types of neurons projecting to either the rostral ventrolateral medulla (PVN(RVLM)) or the intermediolateral horn (IML) of the spinal cord (PVN(IML)). These two neuron groups are intermingled in the same subdivisions of the PVN and differentially regulate sympathetic outflow. However, electrophysiological evidence supporting such functional differences is largely lacking. Herein, we compared the electrophysiological properties of these neurons by using patch-clamp and retrograde-tracing techniques. Most neurons (>70%) in both groups spontaneously fired in the cell-attached mode. When compared to the PVN(IML) neurons, the PVN(RVLM) neurons had a lower firing rate and a more irregular firing pattern (p < 0.05). The PVN(RVLM) neurons showed smaller resting membrane potential, slower rise and decay times, and greater duration of spontaneous action potentials (p < 0.05). The PVN(RVLM) neurons received greater inhibitory synaptic inputs (frequency, p < 0.05) with a shorter rise time (p < 0.05). Taken together, the results indicate that the two pre-sympathetic neurons differ in their intrinsic and extrinsic electrophysiological properties, which may explain the lower firing activity of the PVN(RVLM) neurons. The greater inhibitory synaptic inputs to the PVN(RVLM) neurons also imply that these neurons have more integrative roles in regulation of sympathetic activity.
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Animals , Action Potentials , Fires , Horns , Inhibitory Postsynaptic Potentials , Membrane Potentials , Neurons , Paraventricular Hypothalamic Nucleus , Patch-Clamp Techniques , Spinal Cord , Spinal Cord Lateral HornABSTRACT
Objective To study the characteristic of ultra-rapid delayed rectifier-potassium channel K current (IKur) in rat isolated atrial myocytes using whole-cell patch clamp technique, and the effect of Ibulitide on rat isolated atrial myocytes. Methods Atrial myocytes of rats on an in vitro Langendorf perfusion system were obtained by double digestion (collagenase typeⅡ) method. Whole-cell patch-clamp recording technique was used to record IKur currents. The current-voltage (I-V) curves of K+currents of atrial myocytes were fitted. With filled Ibulitide (2μg/L, filled 3 minutes), IKur was recorded. Results The Results showed that IKur was activated rapidly, almost no lag,and deactivation slowly. This current was activated at about-20 mV, the peak current was present apparent voltage dependence. The peak current density was (2.01 ± 0.27) pA/pF. After filled with Ibulite, this current was no any changed activation. Ibulite decreased the peak current density from+30 mV to+50 mV,but no statistical significance. Conclusion The IKur of rats can activate rapidly with no delays and inactivate slowly. This current is ultra-rapid delayed rectify outside current,and obvious displays the characteristic of outward rectification of whole action potential duration. The 2 μg/L Ibulitide shows no statistical significance for this current.
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Objective To investigate the effects ofdocosahexenoic acid (DHA) on large conductance Ca2+-activated K+ (BK) channels in normal retinal artery smooth muscle cells (RASMCs).Methods Cultured human RASMCs (6 th-8 th generations) were used to patch clamp experiment.The open probabihties (NP0) in BK channels with different concentrations (0.0,1.0,3.0,5.0,7.5,10.0 μmol/L) of DHA were recorded by patch clamp technique in single channel configuration.RASMCs were intervened by different concentrations (0.0,1.0,5.0 μmol/L) of DHA as control group,low and high doses of DHA groups,respectively.The protein expressions of β subunit of BK channels in RASMCs from three groups were measured by Western blot.Results The NP0 of BK channels were 0.044 4±0.001 2,0.081 2±0.004 2,0.209 0±0.006 1,0.310 5±0.005 3,0.465 0±0.007 8 and 0.497 7±0.014 5 with perfusate of 0.0,1.0,3.0,5.0,7.5,10.0 μtmol/L DHA.DHA activated BK channels in a dose-dependent manner (F=2.621,P<0.05).There was no significant difference in the protein expression of control group,low and high doses of DHA groups (F=1 1.657,P>0.05).Conclusion DHA can directly activate BK channels,no increasing in subunit expression of BK channels.
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Background The visual development is completed during the critical period in human and mammals.However,the critical period is not the initial of receiving visual experience.It is known that before the onset of critical period in mammals,such as mouse,there is an earlier stage for visual development,the pre-critical period.The research of response characteristics of the visual cortical neurons and the synaptic plasticity in the pre-critical period is still in the exploratory stage.Objective The study aimed to preliminarily investigate the response properties of neurons and synaptic plasticity in mouse visual cortex during the pre-critical period.Methods Fortyeight postnatal day 13-17 C57BL/6J mice were used for in vivo whole-cell recordings and in vitro brain slice wholecell recordings.In vivo whole-cell recordings were done in anesthetized mice.Moving bars in different directions were produced and controlled by a Matlab program.Cell recordings were obtained at the depth of layer Ⅳ of visual cortex.Step current stimuli under current clamp were given to measure the membrane response properties of neurons.Optimal visual stimuli were given to measure the in vivo largest responses of membrane potentials.In vitro experiments were performed after in vivo experiments.All cells were given current step stimuli to measure the membrane response properties of neurons.Different intensities of white-matter-to-layer-Ⅳpathway stimulation were given to measure the evoked response properties.All cells from 48 mice were randomized into 4 groups according to different stimulus training modes,including low frequency stimulation (LFS),high frequency theta-burst stimulation (TBS),pre-post synaptic timing stimulation (pre-post TS) and post-pre synaptic timing stimulation (post-pre TS).Under the voltage clamp of-70 mV,excitatory postsynaptic currents (EPSCs) before and after training were recorded to measure the plastic changes of excitatory synaptic connections.pClamp 10 was used for the pre-analysis of data and Matlab 2008a was used for statistical analysis.The use and care of the animals followed the Statement for the Use of Animals in Ophthalmic and Vision Research.Results Thirty-nine cells and 48 cells were successfully recorded in the in vivo and in vitro experiments,respectively.The steady-state average number of action potentials (APs) were (1.01 ± 0.03)/sweep and (1.01 ±0.05)/sweep,the AP thresholds were (-40.2 ± 3.2) mV and (-39.6 ±2.0) mV,and the threshold step current levels were (126.7 ± 17.4) pA and (129.6 ± 17.5) pA in the in vivo and in vitro recordings,respectively,with no significant differences between them (APs:t =0.512,P =0.610;AP thresholds:t =-1.074,P =0.286;current levels:t =-0.776,P =0.440).Under the optimal visual or pathway stimulation,the average peak response of membrane potentials was (7.3 ±4.3)mV and (6.4±2.8)mV with rarely evoked APs in the in vivo and in vitro experiments,respectively,with no significant difference between them (t =1.234,P =0.221).Under the in vitro recording,the EPSCs before LFS were [(138.1 ±51.9)pA],which was significantly higher than that after LFS [(76.1 ± 34.8)pA] (t=4.437,P=0.001),but no significant differences were seen in EPSCs before and after TBS (t=-0.756,P=0.466).The EPSCs before and after pre-post TS were (122.4±62.2)pA and (78.5±46.7)pA,and those before and after post-pre TS were (131.9 ±48.0) pA and (74.3 ± 30.7) pA,showing significant differences between them (pre-post TS:t =3.558,P =0.004;post-pre TS:t =4.283,P =0.001).Conclusions The construction of fundamental neural circuits in layer Ⅳ of mouse visual cortex is completed during pre-critical period.However,the membrane responsive capability of neurons and the synaptic connections are in an immature state,and the evoked responses to visual pathway inputs are basically subthreshold.The strength of synaptic connections is depressed with low frequency stimulation or pre-post/post-pre synaptic timing stimulation,and kept unchanged with high frequency stimulation.The development of visual neural system of PSP in mouse presents different characteristics from CP.
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Aim To investigate the effect of gelsemium alkaloids on chloride channels and cell volume in he-patic carcinoma cells. Methods The time-lapse live cell imaging and whole-cell patch clamp techniques were used respectively to detect the volume changes and currents induced by gelsemium alkaloids in HepG2 cells. Results It was found that the cell volume was decreased by (12. 48 ± 2. 2) % (P<0. 01) when ex-posed to gelsemium alkaloids for 50 min and this phe-nomenon could be inhibited by the chloride channel blocker tamoxifen. It was shown by whole-cell patch clamping that a chloride current could be evoked by extracellular application of gelsemium alkaloids ( 2μmol·L-1 ) . The current was outward-rectified with-out obvious voltage- and time-dependent inactivation. The reversal potential of the current was ( -3. 21 ± 0. 67) mV ,which was close to the equilibrium poten-tial of chloride. The extracellular application of the chloride blockers, tamoxifen and 5-notro-2-(3-phenyl-propylamino)benzoic acid (NPPB), and 47% hyper-tonic solution inhibited the current significantly ( P <0. 01 ) . Conclusion Gelsemium alkaloids could acti-vate chloride channels and induce a volume decrease ( named apoptotic volume decrease, AVD) , and these effect could be inhibited by chloride channel blockers. The results suggest that the chloride channel can be one of the targets of gelsemium alkaloids in their anti-cancer action.
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Objective To observe the impact of alone or combined use of ezetimibe and simvastatin on transient outward potassium current (Ito) in ventricular myocytes of rat model of ischemia and reperfusion (IR). Methods Seventy-five male SD rats were randomly divided into five groups, control group (CON), control-IR group (CIR), ezetimibe treatment group (EIR), simvastatin treatment group (SIR) and combined ezetimibe and simvastatin treatment group (ESIR). After two weeks of treat?ment with intragastic normal saline or drugs (ezetimibe or simvastatin), myocytes were isolated from right ventricular with colla?genaseⅡ, and Ito was recorded by whole-cell patch clamp technique. Results (1) The Ito current density at+60 mV was sig?nificantly decreased in CIR group than that of CON group (P0.05). The Ito current densities were higher in SIR group and ESIR group compared to those of CIR group. There was no significant difference in Ito current density between SIR group and ESIR group (P>0.05). (2) There was a significant increase in the half-inactivation (V1/2) in CIR group than that of CON group, but no significant differ?ence between EIR group and CIR group (P >0.05). There was a significant difference in the half-inactivation (V1/2) in SIR group and ESIR group compared to that of CIR group (P0.05). There was no significant difference in the slope factor (K) between five groups (P>0.05). (3) The time-con?stant (τ) of Ito recovery curves from inactivation was significantly higher in CIR group than that of CON group (P0.05). There was a significant difference in the time-con?stant (τ) of Ito recovery curves from inactivation in SIR group and ESIR group compared to that of CIR group (P0.05). Conclusion Simvastatin pre-treatment or ezetimibe+simvastatin pre-treatment can reverse the effect of IR on Ito of ventricular myocytes, but ezetimibe shows no such effects.
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Objective To study the effect of spontaneous subarachnoid hemorrhage (SAH)on transient receptor potential melastatin 4 (TRPM4)channel activity. Methods Seventeen SD rats of clean grade were selected. They were randomly divided into either a SAH (n = 10)or a sham operation group (n = 7) according to the random number table. At day 5 after SAH modeling,the cerebral arteries were harvested and the cerebral arterial smooth muscle cells were isolated using enzymatic digestion method. Western blot was used to detect TRPM4 expression and translocation rate. Patch-clamp techniques were used to study the maximum current intensity of the TRPM4 single channel in cerebral arterial smooth muscle cells. Results The fluorescent-stained TRPM4 were observed in cerebral arterial smooth muscle cells in the 2 groups of rats. The relative quantities of TRPM4 in the total protein of the sham operation group and the SAH group were 24 ± 3% and 32 ± 4% respectively. There was significant difference between the 2 groups (t = 4. 47,P 0. 05). Conclusion SAH has the induced effect for TRPM4 activity.
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Objective To use the whole-cell patch clamp recording to observe the effect of adenosine on hippocampus pyramidal neurons epileptiform discharge from lithium chloride-pilocarpine induced epileptic rats.Methods Twenty adult male and female SD rats (weighing about 200 g) were selected and were narcotized by 10% chloral hydrate intraperitoneal injection.The whole brain was removed, then chopped into 350 μm thick slices.The brain slices were transferred to 37 ℃ preheated artificial cerebrospinal fluid for 1 h and then transferred to 23 ℃ for 30 min.Action potentials (AP) and evoked excitatory postsynaptic currents (eEPSCs) of brain slices neurons were recorded by whole-cell patch clamp, in addition, 25 μmol/L adenosine was used to observe its effect on AP and eEPSCs.Results With the application of current clamp technique, the application of adenosine (25 μmol/L) significandy decreased the numbers of AP in epileptic neurons (P < 0.01) compared with the normal neurons ((0.50 ± 0.06) nA), and eEPSCs amplitude of epileptic neurons increased significantly ((1.44 ± 0.06) nA;independent sample t test, t =30.99, P < 0.01) by voltage clamp.Bath application of adenosine (25 μmol/L) significantly reduced eEPSCs amplitude ((0.66 × 0.06) nA), while application of A1 receptor inhibitor DPCPX partially reversed this effect ((0.92 × 0.06) nA;one-way analysis of variance and q test, F =266.3, q =9.81, P < 0.01).Conclusion Adenosine possesses a strong inhibitory effect on epileptic hippocampal brain slices discharge, which is mediated by its effect on eEPSCs.
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PURPOSE: Alpha1 (alpha1)-adrenoceptor antagonists are widely used to treat lower urinary tract symptoms. These drugs not only act on peripheral tissues, but also cross the blood-brain barrier and affect the central nervous system. Therefore, alpha1-adrenoceptor antagonists may enhance brain functions. In the present study, we investigated the effects of tamsulosin, an alpha1-adrenoceptor antagonist, on short-term memory, as well as spatial learning and memory, in rats. METHODS: The step-down avoidance test was used to evaluate short-term memory, and an eight-arm radial maze test was used to evaluate spatial learning and memory. TUNEL (terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling) staining was performed in order to evaluate the effect of tamsulosin on apoptosis in the hippocampal dentate gyrus. Patch clamp recordings were used to evaluate the effect of tamsulosin on ionotropic glutamate receptors, such as N-methyl-D-aspartate (NMDA), amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA), and kainate receptors, in hippocampal CA1 neurons. RESULTS: Tamsulosin treatment improved short-term memory, as well as spatial learning and memory, without altering apoptosis. The amplitudes of NMDA-induced ion currents were dose-dependently increased by tamsulosin. However, the amplitudes of AMPA- and kainate-induced ion currents were not affected by tamsulosin. CONCLUSIONS: Tamsulosin enhanced memory function by activating NMDA receptor-mediated ion currents in the hippocampus without initiating apoptosis. The present study suggests the possibility of using tamsulosin to enhance memory under normal conditions, in addition to its use in treating overactive bladder.
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Animals , Rats , Apoptosis , Blood-Brain Barrier , Brain , Central Nervous System , Dentate Gyrus , Hippocampus , In Situ Nick-End Labeling , Learning , Lower Urinary Tract Symptoms , Memory , Memory, Short-Term , N-Methylaspartate , Neurons , Patch-Clamp Techniques , Receptors, Ionotropic Glutamate , Receptors, Kainic Acid , Receptors, N-Methyl-D-Aspartate , Urinary Bladder, OveractiveABSTRACT
Background:Visceral hypersensitivity is considered to be one of the major pathophysiological mechanisms of irritable bowel syndrome. Aims:To investigate the expression of TRPV1 and electrophysiological characteristics of colon-specific dorsal root ganglion( DRG)neurons in rat model of visceral hypersensitivity. Methods:Twenty 10-day-old rats were randomly divided into two groups. In model group,visceral hypersensitivity was induced by colorectal administration of acetic acid;while in control group the same amount of saline was administered. Colon-specific DRG neurons were labeled retrogradely by injection of DiI,a fluorochrome,into the colon wall. Expression of TRPV1 in DRG neurons was detected by immunofluorescence and the electrophysiological characteristics of DRG neurons was detected by using patch-clamp technique. Results:In model group,the expression rate of TRPV1 in colon-specific DRG neurons was significantly higher than that in controls(46. 1% vs. 36. 6% ,P <0. 01),the average rheobase was significantly decreased[(57. 80 ±1. 32)pA vs.(73. 45 ± 4. 51)pA,P < 0. 05],while the frequency of action potentials(APs)in response to doubling rheobase stimulation was significantly increased[(8. 20 ± 1. 10)Hz vs. (4. 54 ± 0. 66)Hz,P < 0. 05]. Score of abdominal withdrawal reflex(AWR)under a 60 mm Hg colorectal distention was positively correlated with the expression rate of TRPV1 and the frequency of APs in response to doubling rheobase stimulation(r = 0. 87 and r = 0. 73,P < 0. 01),but was negatively correlated with the rheobase(r = - 0. 81,P < 0. 01)in model group. Conclusions:Increased expression of TRPV1 and excitability in colon-specific DRG neurons might be a crucial step in formation of visceral hypersensitivity.
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Objective To evaluate the role of calcium channel in the mechanism of the generation and maintenance of bursting firing of substantia nigra pars compacta (SNc) dopaminergic neurons in rats.Methods Using the patch clamp technique,we observed the firing pattern switching features after adding 10 μmol/L N-methyl-D-aspartic acid (NMDA),compared the changes of whole-calcium current and L-type calcium current with or without NMDA,and analyzed the correlation between the generation of burst firing and L-type calcium channel activation.Results After NMDA treatment,the firing pattern of SNc dopaminergic neurons changed to burst firing,which was compromised by a charastistic high plateau potential and series of action potential on it.The current density of L-type calcium current increased significantly after adding NMDA,which,from (2.86 ±0.26) pA/pF (n =28),significantly increased to (3.75 ± 0.18) pA/pF (n =34 ; t =7.52,P =0.002 8).The high plateau potential was almost abolished with the application of verapamil,a specific antagonist of L-type calcium channel.Consiusion NMDA could induce the firing pattern changed to burst firing in SNc dopaminergic neurons,while L-type calcium channel contributes to the process of generation and maintenance of burst firing.
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AIM@#To study the effects of crebanine on voltage-gated Na(+) channels in cardiac tissues.@*METHODS@#Single ventricular myocytes were enzymatically dissociated from adult guinea-pig heart. Voltage-dependent Na(+) current was recorded using the whole cell voltage-clamp technique.@*RESULTS@#Crebanine reversibly inhibited Na(+) current with an IC50 value of 0.283 mmol·L(-1) (95% confidence range: 0.248-0.318 mmol·L(-1)). Crebanine at 0.262 mmol·L(-1) caused a negative shift (about 12 mV) in the voltage-dependence of steady-state inactivation of Na(+) current, and retarded its recovery from inactivation, but did not affect its activation curve.@*CONCLUSION@#In addition to blocking other voltage-gated ion channels, crebanine blocked Na(+) channels in guinea-pig ventricular myocytes. Crebanine acted as an inactivation stabilizer of Na(+) channels in cardiac tissues.
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Animals , Female , Male , Aporphines , Pharmacology , Cells, Cultured , Down-Regulation , Drugs, Chinese Herbal , Pharmacology , Guinea Pigs , Heart Ventricles , Cell Biology , Metabolism , Myocytes, Cardiac , Metabolism , Stephania , Chemistry , Voltage-Gated Sodium Channel Blockers , Pharmacology , Voltage-Gated Sodium Channels , MetabolismABSTRACT
Objective To study the effect of subarachnoid hemorrhage (SAH) on voltagedependent potassium channels (Kv) currents of smooth muscle cells,which is hypothesized to be different between cerebral pial arteries and penetrating arteries.Methods Smooth muscle cells from cerebral pial arteries and penetrating arteries in rats were enzymatically isolated 72 h after SAH,and patch clamp was used to test the relative cell size,resting potential and Kv currents.Results Resting potential of either pial ((45.63 ±1.18) mV) or penetrating artery ((41.55-± 1.19) mV) was shifted positively by SAH,even more significantly in latter (F =8.24,P < 0.05 ; F =9.36,P < 0.01) ; Resting potential of pial artery of control ((38.76 ± 1.03) mV),penetrating artery of control ((38.53 ± 0.67) mV),pial artery of SAH ((36.87 ± 1.49) mV) and penetrating artery of SAH((37.89 ± 1.24) mV) were shifted positively to the same level by 1 μmol/L 4-aminopyridine (4AP; F =3.08,P >0.05).Maximum Current Density (Imax) of either pial ((20.82 ±0.59) pA/pF) or penetrating artery ((15.15 ±0.37) pA/pF) was compromised by SAH,also more significantly in latter (F =6.22,P < 0.05) ; Imax of pial artery of control (9.15 ± 0.16),penetrating artery of control (9.04 ± 0.36),pial artery of SAH (8.77 ± 0.26) and penetrating artery of SAH (9.12 ± 0.17) were decreased to the same level by 1 μmol/L 4AP (F =2.96,P > 0.05).Conclusions SAH probably shares the similar pathway with Kv blocker (4AP) in Kv currents inhibition.Further,SAH differently inhibits smooth muscle cells Kv currents and resting potential of cerebral pial arteries and penetrating arteries,which may be related with their different sensitivity towards cerebral vasospasm following SAH.
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AIM:To study the effects and mechanisms of ethanol on chloride channels in poorly differentiated nasopharyngeal carcinoma CNE-2Z cells.METHODS:The effect of ethanol on the cell growth was analyzed by MTT as-say.The technique of whole-cell patch-clamp was used to detect the chloride current .The characteristics of the chloride current were analyzed by using the chloride channel blockers .The siRNA technique was used to analyze the molecular basis of the ethanol-sensitive chloride channels .RESULTS: Under isotonic conditions , the background current was weak and stable.Ethanol at concentrations of 0.17~170 mmol/L activated a chloride current in a concentration-dependent manner (an inverted U-shape), with a maximum effect at the concentration of 17 mmol/L.The currents showed obviously outward rectification and were susceptible to extracellular hypertonicity and the chloride channel blocker , 5-nitro-2-(3-phenylpropyl-amino) benzoic acid ( NPPB) .ClC-3 siRNA obviously decreased the currents activated by ethanol .CONCLUSION:Ex-tracellular ethanol induces chloride currents through activating the ClC-3 chloride channels .
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[ABSTRACT]AIM:Toinvestigatetheeffectsofmechanicalstretchontransientoutwardpotassiumcurrent(Ito), inward rectifier potassium current ( IK1 ) and action potential duration ( APD) of cultured neonatal rat atrial myocytes . METHODS:Neonatal rat atrial myocytes were isolated and cultured on silicone sheeting with or without stretch for 24 h. The silicone membrane area was increased by 12%in stretched group.The cells without stretch served as control .Ito, IK1 and APD were recorded by the technique of whole-cell patch clamp.RESULTS:Compared with control group, Ito density in stretched myocytes was significantly reduced [(1.6 ±0.4) pA/pF vs (12.1 ±2.9) pA/pF, P<0.01], whereas IK1 density was increased [(-10.8 ±0.8) pA/pF vs (-8.8 ±0.9) pA/pF, P<0.01].The APDs at 50%and 90%levels of repolarization ( APD50 and APD90 ) in the stretched cells were obviously decreased than those in non-stretched cells [(10.5 ±1.4) ms vs (15.5 ±2.4) ms, (30.0 ±2.8) ms vs (56.3 ±3.6) ms, P<0.01].CONCLUSION: Stretch stimulation leads to the reduction of Ito density, the increase in IK1 density and the shortness of APD in cultured rat atrial neonatal myocytes , which may contribute to atrial electrical remodeling induced by pressure overload .