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Chinese Chemical Letters 2001年第9期12卷 813-814页

作者： Pin YANG Yan Fang SHANG Zhen Ming PEI Institute of Molecular Science Shanxi UniversityTaiyuan 030006 Department of Biology Duke UniversityDurhamNC 27708

We used whole-vacuolar patch-clamp recording mode to study the action mechanism of La3+ to Slow Vacuolar (SV) channels for the first time. We recorded SV channel currents of Xinlimei (Raphanus satirus L.) vacuolars. The minimum activation potentials of voltage-dependent SV channels tied in 25+/-5 mV. The increase in cytoplasmic Ca2+ led to enhancement of SV-type currents. It was found that the threshold potential of activation shifted towards more depolarized values whenever cytoplasmic Ca2+ was increased. When 10(-10) mol/L free La3+ was added to the bath, SV-type current was suppressed by 60 similar to 75%. These data showed La3+ reduced ion permeabilities of Xinlimei root vacuolar membrane.

关键词： Whole-vacuolar recording SV-type current cytoplasmic Ca2+ La3+ patch-clamp

Study of transmembrane La^(3+) movement in rat ventricular myocytes by the patch-clamp technique

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Chinese Science Bulletin 2002年第18期47卷 1518-1522页

作者： YANG Pin, DU Huizhi & XUE ShaowuInstitute of Molecular Science, Shanxi University, Taiyuan 030006, China Institute of Molecular Science Shanxi University Taiyuan China

We have studied transmembrane La3+ movement in rat ventricular myocytes for the first time by using the whole-cell patch-clamp recording mode. La3+ (0.01-5.0 mmol/L) could not bring out inward currents through the L-type calcium channel in rat ventricular myocytes, while it could enter the cells by the same way carried by 1μmo1/L ionomycin. When the outward Na+ concentration gradient is formed, La3+ can enter the cells via Na-Ca exchange, and the exchange currents increase with the increase of external La3+ concentrations. But compared with Na-Ca exchange currents in the same concentration, the former is only 14%-38% of the latter. The patch-clamp experiment indicates that La3+ normally can not enter ventricular myocytes through L-type calcium channel, but it can enter the cells via Na-Ca exchange.

关键词： whole-cell patch-clamp recording ventricular myocyte L-type calcium channel Na-Ca exchange La3+ Ca2+.

A slice preparation of rat pancreatic islets for patch-clamp recordings in single β cells

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A slice preparation of rat pancreatic islets for patch-clamp...

Isolation of Ca^(2+) Tolerant Cardiomyocytes from Aadult Rats for patch clamp Studies

2008北京国际生理科学学术大会

作者： WU Zheng-Yong~(1,2),ZHENG Liang-Hong~1,DOU Hai-Qiang~1,ZHU Kuai-Chang~1,LIU Chang~1,YAO Li-Juan~1,YU Xiao~1,ZHANG Claire Xi~1,SHEN Ying~2,ZHOU Zhuan~1 1 Institute of Molecular Medicine,Peking University,Beijing 100871,China 2 Department of Neurobiology,Zhejiang University School of Medicine,Hangzhou 310031,China

Pancreatic isletβcells are responsible for blood glucose control by releasing *** cells sense blood glucose by ATP-receptor,which controls Ca entry and insulin *** disorder inβcell may lead to the severe disease *** fighting diabetes,patch clamp recording in cultureβcells is a powerful tool to examine mechanisms of insulin ***,βcell is metabolic type;it is difficult to prepare primary culture of rat or mouseβcells for *** overcome problem inβcell culture,we aim to prepare slices of pancreas,which enables patch clamp *** gelling-agarose was injected into the distally clamped bile duct and cooled it with an ice-cold extracellular solution,which enhance the mechanical strength for ***βcells in slices are easily patch-clamped,and its voltage-gated currents and evoked secretion can be recorded under whole-cell configuration.

关键词： Pancreatic islet b cells patch-clamp voltage-gated currents ATP-receptor

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Journal of Nanjing Medical University 2004年第1期18卷 4-6页

作者：徐华娥陶金陈洁汪红仪李胜男南京医科大学药理学系

Objective: To investigate the factors affecting the viability and Ca 2+ tolerance of isolated rats' cardiac myocytes for patch clamp research. Methods: Hearts were firstly perfused by the Langendorff perfusion apparatus with normal Tyrode's solution, then with Ca 2+-free Tyrode's solution and subsequently with low Ca 2+ enzyme solution containing collagenase 0.1-0.2 g/L. All the solutions were saturated with oxygen and the perfusion temperature was kept at 37 ℃. Finally hearts were washed by Ca 2+-free Tyrode's solution, after which the ventricles were minced into small pieces in KB solution, dispersed and filtered. The isolated myocytes were stored in KB solution at room temperature for 1 h and recovered to normal calcium concentration before patch clamp ***: When all the factors such as water, enzyme, Ca 2+,pH, and oxygen were well controlled, the well constructed and rod-like cardiac myocytes with a yielding rate of 30%-50% came ***: All the factors should be well controlled, which ensured the isolated cells Ca 2+ tolerant and appropriate for patch clamp experiments.

关键词： isolation cardiac myocyte patch-clamp Tyrode

MicroRNA-502-3p regulates GABAergic synapse function in hippocampal neurons

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Neural Regeneration Research 2024年第12期19卷 2698-2707页

作者： Bhupender Sharma Melissa MTorres Sheryl Rodriguez Laxman Gangwani Subodh Kumar Center of Emphasis in Neuroscience Department of Molecular and Translational MedicinePaul L.Foster School of MedicineTexas Tech University Health Sciences CenterEl PasoTXUSA Bond Life Sciences Center and Department of Veterinary Pathobiology University of MissouriColumbiaMOUSA L.Frederick Francis Graduate School of Biomedical Sciences Texas Tech University Health Sciences CenterEl PasoTXUSA

Gamma-aminobutyric acid(GABA)ergic neurons,the most abundant inhibitory neurons in the human brain,have been found to be reduced in many neurological disorders,including Alzheimer's disease and Alzheimer's disease-related *** previous study identified the upregulation of microRNA-502-3p(miR-502-3p)and downregulation of GABA type A receptor subunitα-1 in Alzheimer's disease *** study investigated a new molecular relationship between miR-502-3p and GABAergic synapse *** vitro studies were perfo rmed using the mouse hippocampal neuronal cell line HT22 and miR-502-3p agomiRs and *** silico analysis identified multiple binding sites of miR-502-3p at GABA type A receptor subunitα-1 *** assay confirmed that miR-502-3p targets the GABA type A receptor subunitα-1 gene and suppresses the luciferase ***,quantitative reve rse transcription-polymerase chain reaction,miRNA in situ hybridization,immunoblotting,and immunostaining analysis confirmed that overexpression of miR-502-3p reduced the GABA type A receptor subunitα-1 level,while suppression of miR-502-3p increased the level of GABA type A receptor subunitα-1 ***,as a result of the overexpression of miR-502-3p,cell viability was found to be reduced,and the population of necrotic cells was found to be *** whole cell patch-clamp analysis of human-GABA receptor A-α1/β3/γ2L human embryonic kidney(HEK)recombinant cell line also showed that overexpression of miR-502-3p reduced the GABA current and overall GABA function,suggesting a negative correlation between miR-502-3p levels and GABAergic synapse ***,the levels of proteins associated with Alzheimer s disease were high with miR-502-3p overexpression and reduced with miR-502-3p *** present study provides insight into the molecular mechanism of regulation of GABAergic synapses by *** propose that micro-RNA,in particular miR-502-3p,could be a potential the

关键词： Alzheimer's disease GABAergic synapse gamma-aminobutyric acid type A receptor subunitα-1(GABRα1) microRNA-502-3p(miR-502-3p) miRNA in situ hybridization patch-clamp

L-and T-type Ca^(2+)channels dichotomously contribute to retinal ganglion cell injury in experimental glaucoma

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Neural Regeneration Research 2023年第7期18卷 1570-1577页

作者： Hong-Ning Wang Wen-Jing Qian Guo-Li Zhao Fang Li Yan-Ying Miao Bo Lei Xing-Huai Sun Zhong-Feng Wang State Key Laboratory of Medical Neurobiology and MOE Frontiers Center for Brain Science Institutes of Brain ScienceFudan UniversityShanghaiChina Institutes of Neuroscience and Third Affiliated Hospital Henan Provincial People’s HospitalHenan Eye InstituteHenan Eye HospitalPeople’s Hospital of Zhengzhou UniversityZhengzhou UniversityZhengzhouHenan ProvinceChina Eye Institute and Department of Ophthalmology Eye&ENT HospitalNHC Key Laboratory of MyopiaShanghai Key Laboratory of Visual Impairment and RestorationFudan UniversityShanghaiChina

Retinal ganglion cell apoptotic death is the main pathological characteristic of glaucoma,which is the leading cause of irreversible *** of Ca^(2+)homeostasis plays an important role in ***-gated Ca^(2+)channel blockers have been shown to improve vision in patients with ***,whether and how voltage-gated Ca^(2+)channels are involved in retinal ganglion cell apoptotic death are largely *** this study,we found that total Ca^(2+)current densities in retinal ganglion cells were reduced in a rat model of chronic ocular hypertension experimental glaucoma,as determined by whole-cell patch-clamp electrophysiological *** analysis showed that L-type Ca^(2+)currents were downregulated while T-type Ca^(2+)currents were upregulated at the later stage of *** blot assay and immunofluorescence experiments confirmed that expression of the Ca_(V)1.2 subunit of L-type Ca^(2+)channels was reduced and expression of the Ca_(V)3.3 subunit of T-type Ca^(2+)channels was increased in retinas of the chronic ocular hypertension *** tumor necrosis factor-α,an important inflammatory factor,inhibited the L-type Ca^(2+)current of isolated retinal ganglion cells from control rats and enhanced the T-type Ca^(2+)*** changes were blocked by the tumor necrosis factor-αinhibitor XPro1595,indicating that both types of Ca^(2+)currents may be mediated by soluble tumor necrosis factor-α.The intracellular mitogen-activated protein kinase/extracellular signal-regulated kinase pathway and nuclear factor kappa-B signaling pathway mediate the effects of tumor necrosis factor-α.TUNEL assays revealed that mibefradil,a T-type calcium channel blocker,reduced the number of apoptotic retinal ganglion cells in the rat model of chronic ocular *** results suggest that T-type Ca^(2+)channels are involved in disrupted Ca^(2+)homeostasis and apoptosis of retinal ganglion cells in glaucoma,and application of T-type

关键词： apoptosis CaV1.2 CaV3.3 chronic ocular hypertension extracellular signal-regulated kinase mitogen-activated protein kinase nuclear factor-kappa B patch-clamp retina tumor necrosis factor-α

Two outward potassium current types are expressed during the neural differentiation of neural stem cells

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Neural Regeneration Research 2013年第28期8卷 2656-2665页

作者： Ruiying Bai Guowei Gao Ying Xing Hong Xue Department of Physiology and Neurobiology Xinxiang Medical University Department of Radiotherapy Center Hospital of Xinxiang Stem Cell Research Center Zhengzhou University Basic Medical Sciences of Henan University of Traditional Chinese Medicine

The electrophysiological properties of potassium ion channels are regarded as a basic index for determining the functional differentiation of neural stem cells. In this study, neural stem cells from the hippocampus of newborn rats were induced to differentiate with neurotrophic growth factor, and the electrophysiological properties of the voltage-gated potassium ion channels were observed. Immunofluorescence staining showed that the rapidly proliferating neural stem cells formed spheres in vitro that expressed high levels of nestin. The differentiated neurons were shown to express neuron-specific enolase. Flow cytometric analysis revealed that the neural stem cells were actively dividing and the percentage of cells in the S ＋ G2/M phase was high. However, the ratio of cells in the S ＋ G2/M phase decreased obviously as differentiation proceeded. Whole-cell patch-clamp re- cordings revealed apparent changes in potassium ion currents as the neurons differentiated. The potassium ion currents consisted of one transient outward potassium ion current and one delayed rectifier potassium ion current, which were blocked by 4-aminopyridine and tetraethylammonium, respectively. The experimental findings indicate that neural stem cells from newborn rat hippo- campus could be cultured and induced to differentiate into functional neurons under defined condi- tions in vitro. The differentiated neurons expressed two types of outward potassium ion cur＇ents similar to those of mature neurons in vivo.

关键词： neural regeneration neural stem cells hippocampus proliferation differentiation neurons patch-clamp electrophysiological properties transient outward potassium ion current delayed rec-tifier potassium ion current inactivation nestin neuron-specific enolase grants-supported paper neuroregeneration

Endothelium-derived Relaxing Factor Activates Calcium-activated Potassium Channels of Resistance Vessel Smooth Muscle Cells

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Science China Chemistry 1993年第4期36卷 439-450页

作者：汤云贵郑永芳 Department of Physiology Institute of Basic Medical Sciences CAMS Faculty of Basic Medicine PUMC Beijing 100005 PRC

Direct observation was made by using the patch-clamp technique with a specially designed microperfusion system to investigate the effect of acetylcholine (Ach 10^(-6) mol/L) elicited endothelium-derived relaxing factor (EDRF) on the calcium-activated potassium channel (IK(Ca))in the smooth muscle cells of mesenteric resistance vessels in Wistar rats. Activation of IK(Ca) was firstly observed by inducing the elicited EDRF or sodium nitroprusside (SNP 10^(-8) mol/L) under various clamping voltages in cell-attached configuration. While the pipette solution contained KCl 126 mmol/L and the bath solution contained KCl 5.9 mmol/L, two types of conductances of calcium-activated potassium current being 76.4±2.3 pS(mean±S.E. n = 7) and 160.3±7.5 pS (mean±S.E. n= 7) were recorded during the EDRF activation, one type of conductance being 100.5±2.8 pS (mean±S.E. n = 6) was activated by nitric oxide (NO) which is an effective component from SNP. Differences in kinetic characteristics of these channels between EDRF and NO activation were found, particularly the probability of the channel being open in EDRF activation was obviously greater than that in NO stimulation. It has been shown that the potassium channel mechanisms involved in the EDRF and NO actions might be different.

关键词： endothelium-derived relaxing factor (EDRF) calcium-activated potassium channel (IK(Ca)) mesenterie resistance vessel smooth muscle patch-clamp technique open microperfusion system.

Effects of Muscle Electrical Activity on the Transmission of Developing Neuromuscular Junction

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Science China Chemistry 1994年第5期37卷 573-580页

作者：汤云贵谢佐平毛健何展全赵南明 State Key Laboratory of Biomembrane and Membrane Biotechnique Department of Biological Sciences and Biotechnology Tsinghua University Beijing 100084 PRC

Miniature endplate potentials (MEPPS) caused by the spontaneous release of ACh from the growth cone of cholinergic neurons, are recorded by the whole-cell patch-clamp technique on a large number of 1-day cultured myoballs which have contact neurites of co-cultured neurons. Both muscle cell and neuron are dissociated from the 1-day-old (about stage 20) Xenopus embryo. Frequency and/or amplitude of MEPPs can obviously increase after the repetitive high-level depolarization caused by the stimuli on muscle cells. No detectable changes of single ACh receptor channel property are observed by using the single-channel recording technique. These results suggest that the mechanism of the increase of MEPPs after electrical activity of postsynaptic muscle cells probably involve some alteration of presynaptic membrane.

关键词： electrical activity of muscle cell patch-clamp technique miniature endplate potentials (MEPPs) acetylcholine (ACh) receptor channel.

Effects of lanthanum on calcium-activated K^+ currents and its kinetics in MC3T3 cells

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Science China Chemistry 2010年第6期53卷 1281-1286页

作者： ZHANG LiPing1,2 & YANG Pin1 1Key Laboratory of Chemical Biology and Molecular Engineering of Ministry of Education Institute of Molecular Science,Shanxi University,Taiyuan 030006,China 2College of Environment and Safety,Taiyuan University of Science and Technology,Taiyuan 030024,China Key Laboratory of Chemical Biology and Molecular Engineering of Ministry of Education Institute of Molecular Science Shanxi University Taiyuan China College of Environment and Safety Taiyuan University of Science and Technology Taiyuan China

Using the whole cell patch-clamp technique,we studied the effects of La3+ on calcium-activated K+ currents and its kinetics of activation and inactivation in non-excitable MC3T3 *** results showed that the calcium-activated outward K+ currents were promoted with increasing concentration of Ca2+ in the pipette solution and a voltage- and Ca2+-dependent ***3+ in the bath solution inhibited the currents in a concentration-dependent manner and the inhibition EC50 was 8.23 ± 1.45 μmol/*** the concentration of 50 μmol/L,La3+ significantly changed the Vh of the activation curve and shifted the activation curve to more positive potentials,but shifted the inactivation curve to more negative *** had no effect on the slope factor k of the activation and inactivation *** currents inhibition could induce a series of physiological and molecular biological functions,presumably because of its ability to depolarize the plasma membrane and enhance cell excitability,resulting in increasing Ca2+ influx and cytoplast Ca2+ *** process may be one of the molecular mechanisms by which La3+ affects the cell growth and function of MC3T3 cells.

关键词： MC3T3 cell potassium ion channel La3+ patch-clamp technique