Effect of Cu^2＋ on K^＋ Current in Acutely Isolated Rat Hippocampal Neurons by Whole Cell Patch Clamp Technique

用整个房间补丁夹钳技术，短暂的外面的 K+ 电流(Ito ) 和推迟的整流器 K+ 电流(Idr ) 上的 Cu2+ 的效果在尖锐地孤立的老鼠被学习海马趾的神经原。当 Cu2+was 的集中比 2H 降低时， Ito 和 Idr

Effect of temperature on the activation of myocardial K_(ATP) channel in guinea pig ventricular myocytes: a pilot study by whole cell patch clamp recording

Background The myocardial ATP sensitive potassium channel （KATP channel） has been known for more than two decades, the properties of this channel have been intensively investigated, especially the myocardial protection effect by opening this channel. Numerous studies, including hypothermic, using KATP agonists to achieve a hyperpolarizing cardioplegic arrest, have shown a better myocardial protection than potassium arrest. However, there is no evidence showing that KATP channel could be opened by its agonists under profound hypothermia. We investigated the effect of temperature on activation of myocardial KATP channel by nicorandil.Methods Isolated ventricular myocytes were obtained by collagenase digestion of the hearts of guinea pigs and stored in KB solution at 4℃. With a steady ground current, the myocytes were perfused with 1 mmol/L nicorandil until a steady IKATP occurred. Then the cells were perfused with 1 mmol/L nicorandil plus 1 μmol/L glybenclamide. Currents signals were recorded on whole cells using patch clamp technique at several temperatures. The temperature of the bath solution around myocytes was monitored and was controlled at 4℃, 10℃, 20℃, 25℃ and 35℃ respectively. About 10 cells were tested at each temperature, the cells were considered useful only when the outward current could be induced by nicorandil and blocked by glybenclamide. All data were analyzed using Graphpad PRISM 3.0 （Graphpad, San Diego, CA, USA）. Nonlinear curve fitting was done in Clampfit （Axon） or Sigmaplot （SPSS）.Results At 4℃, 10℃, 20℃, 25℃ and 35℃, the time needed to open the myocardial KATP channel was （81.0±0） minutes, （50.54±11.7） minutes, （28.84±2.3） minutes, （9.4± 10.2） minutes and （2.3± 1.0） minutes respectively （P=0.003）. The linear relationship between temperature and time needed to open the channel was y （min） = （4348.790±124.277x）/60, where y （min） is time needed to open KATP channel, x is temperature, correlation coefficient r =-0.942 （P=0.00）, regression coefficient b =-124.277 （P=0.00）. The current densities among different temperatures were statistically different （P=0.022）, the current density was greater after the activation of KATP channel at higher temperatures. The lower the temperature, the fewer cells in which KATP channels could be opened. At 4℃, only one cell in which the KATP channel could be opened, took a quite long time （81 minutes）and the Ⅰ-Ⅴ curve was quite untypical.Conclusions KATP channel activated by nicorandil is temperature dependent and the temperature linearly related to time needed to open KATP channel; the lower the temperature, the longer the time needed to open channel and the smaller the current density. At profound hypothermia, it is difficult to activate KATP channels.

In vivo patch clamp recording technique in the study of neurophysiology

The patch clamp recording technique in vivois a blind patch clamp recording methods to record the current of the spinal or cereral neurons of anaes：hesia （ or awake） animals. This technique can be used to study the synaptic function and plasticity in central nervous system in vivoin order to understand the physiological properties of the ion channels from an integrated point of view. The advantage of this technique have already presented itself in the study of the synaptic transmission and nervous network. Nowadays, in vivo patch whole-cell recording technique in combination with other techniques is becoming a common method in the research fields.

Induction of pancreatic duct cells of neonatal rats into insulin-producing cells with fetal bovine serum: A natural protocol and its use for patch clamp experiments

AIM: To induce the pancreatic duct cells into endocrine cells with a new natural protocol for electrophysiological study.METHODS: The pancreatic duct cells of neonatal rats were isolated, cultured and induced into endocrine oells with 15% fetal bovine serum for a period of 20 d. During this period, insulin secretion, MTT value, and morphological change of neonatal and adult pancreatic islet cells were comparatively investigated. Pancreatic β-cells were identified by morphological and electrophysiological characteristics, while ATP sensitive potassium channels(KATP), voltage-dependent potassium channels (KV), and voltage-dependent calcium channels (KCA) in β-cells were identified by patch clamp technique.RESULTS: After incubation with fetal bovine serum, the neonatal duct cells budded out, changed from duct-like cells into islet clusters. In the first 4 d, MTT value and insulin secretion increased slowly (MTT value from 0.024±0.003 to0.028±0.003, insulin secretion from 2.6±0.6to 3.1±0.8 mIU/L). Then MTT value and insulin secretion increased quickly from d 5 to d 10 (MTT value from 0.028±0.003 to 0.052±0.008, insulin secretion from 3.1±0.8to 18.3±2.6 mIU/L), then reached high plateau (MTT value ＞0.052±0.008, insulin secretion ＞18.3±2.6 mIU/L).In contrast, for the isolated adult pancreatic islet cells,both insulin release and MTT value were stable in the first 4 d (MTT value from 0.029±0.01 to 0.031±0.011,insulin secretion from 13.9±3.1 to 14.3±3.3 mIU/L), but afterwards they reduced gradually (MTT value ＜0.031±0.011, insulin secretion ＜8.2±1.5 mIU/L), and the pancrearic islet cells became dispersed, broken or atrophied correspondingly. The differentiated neonatal cells were identified as pancreatic islet cells by dithizone staining method, and pancreatic β-cells were further identified by both morphological features and electrophysiological characteristics, i.e. the existence of recording currents from KATP KV, and KCA.CONCLUSION: Islet cells differentiated from neonatal pancreatic duct cells with the new natural protocol are more advantageous in performing patch clamp study over the isolated adult pancreatic islet cells.

Whole-cell recordings of calcium and potassium currents in acutely isolated smooth muscle cells

瞄准：在 mes 伤寒的尖锐地孤立的平滑肌房间记录钙和钾水流在老鼠的动脉的分支。方法：平滑肌房间被胶原酶文摘和机械研碎刚与擦亮的吸量管孤立。在整个房间的模式的补丁 clamp 技术被采用记录钙和钾水流。结果：过程分裂了没有损害电镀物品的平滑肌房间房间的生理的特征。电压门 Ca2+ 和钾水流成功地用整个房间的补丁 clamp 配置被记录。结论：方法分裂从老鼠 mes 伤寒的平滑肌房间动脉的分支。电压门隧道水流能在这准备被记录。

Whole-cell recordings of voltage-gated Calcium,Potassium and Sodium currents in acutely isolated hippocampal pyramidal neurons

Objective：To record Calcium, Potassium and Sodium currents in acutely isolated hippocampal pyramidal neurons. Methods：Hippocampal CA3 neurons were freshly isolated by 1 mg protease/3 ml SES and mechanical trituration with polished pipettes of progressively smaller tip diameters. Patch clamp technique in whole-cell mode was employed to record voltage-gated channel currents. Results：The procedure dissociated hippocampal neurons, preserving apical dendrites and several basal dendrites, without impairing the electrical characteristics of the neurons. Whole-cell patch clamp configuration was successfully used to record voltage-gated Ca^2＋ currents, delayed rectifier K^＋ current and voltage-gated Na^＋ currents. Conclusion：Protease combined with mechanical trituration may be used for the dissociation of neurons from rat hippocampus. Voltage-gated channels currents could be recorded using a patch clamp technique.

The Reconstructing of Low Signal-noise Ratio Single Ion Channel Signal from Patch-clamp Recordings Sampled in the Colored Background Noise

The single ion channel signal is an ionic current that can be recorded by the patch clamp technique. Hidden Markov model (HMM) algorithm has been used to convert the low signal noise ratio (SNR) noisy recording into an idealized quantal one in the case of white background noise. The traditional HMM algorithm is extended and adapted to the colored background noise. A new algorithm called EHMM (Extended HMM) algorithm is proposed, and mainly validated by simulation. Results show that it’s effective.

马钱子碱抑制豚鼠心室肌细胞钠电流的作用研究

目的本研究旨在分析马钱子碱(brucine)对分离的单个豚鼠心室肌细胞钠电流(INa+)的作用,并探索其抗心律失常的可能机制。方法共选取24只健康的成年豚鼠作为实验对象,雌雄不拘,随机分为4组,每组6只:第1组是未接受任何处理的对照组,之后通过微量加样器使培养皿中brucine药物的终浓度分别达到3μmol/L、10μmol/L、30μmol/L。采用急性酶解法分离获得单个豚鼠心室肌细胞,通过全细胞膜片钳技术测量离子通道电流,以检测不同浓度的brucine对豚鼠心室肌细胞INa+的影响。结果正常的对照组没有明显的INa+电流峰值的变化,而brucine 3μmol/L组给药前后基本不影响INa+电流峰值,当brucine的浓度增加到10μmol/L时,给药前后INa+电流峰值显著下降(P<0.05),在30μmol/L组给药前后INa+电流峰值大幅度减少(P<0.01),并且这种抑制作用呈浓度依赖性。正常对照组及brucine 3μmol/L剂量组中,电流-电压曲线(Ⅰ~Ⅴ曲线)并未受到显著的影响;然而,10μmol/L、30μmol/L剂量组与对照组比较,INa+的Ⅰ~Ⅴ曲线上移,无平行移动,且曲线形状不变。结论Brucine能够通过抑制心室肌细胞钠通道电流发挥抗心律失常作用。

无镁诱导仓鼠原代皮质神经元电生理学特性

目的无镁细胞外液建立癫痫放电模型,利用全细胞膜片钳技术,检测仓鼠原代皮质神经元的电生理学特性。方法采用生后1~2 d新生叙利亚仓鼠,分离大脑皮质培养原代神经元培养至第12天,分别予有镁细胞外液(有镁组)和无镁细胞外液(无镁组)孵育3 h,3 h后均更换为正常孵育液继续培养24 h。利用全细胞膜片钳技术,电压钳模式下记录神经元兴奋性突触后电流(excitatory postsynaptic currents,EPSC),电流钳模式下记录神经元兴奋性突触后电位(excitatory postsynaptic potentials,EPSP)。结果与有镁组相比,无镁组仓鼠原代皮质神经元EPSC[(124.38±75.15)Hz vs.(33.93±22.32)Hz,P<0.001]及EPSP[(37.05±38.37)Hz vs.(5.63±9.52)Hz,P<0.01]的频率升高,且差异有统计学意义;而两组之间EPSC及EPSP的振幅、曲线下面积和半宽度的差异无统计学意义(P>0.05)。结论无镁处理后仓鼠原代皮质神经元兴奋性升高,仓鼠原代皮质神经元可用于构建癫痫细胞模型。

Sex Difference in the Repolarization Currents of Rabbit Ventricular Cells

The current difference between male and female rabbit ventricular myocytes was investigated for elucidating the mechanism of longer QT interval and higher incidence of drug-associated torsade de pointes in female rabbits than in male rabbits. Whole cell patch clamp technique was used to record APD, I_to, I_K,tail, I_K1 and I_Ca,L of myocytes from left ventricular apex. There was no difference in the membrane capacitance between male and female rabbit myocytes. APD_90 was longer in female rabbits (560.4±26.5 ms, n=15) than in male ones (489.0±20.7 ms, n=14), P<0.05. In female rabbit myocytes, I_K,tail, I_to, I_K1 and I_Ca,L were 0.71±0.05 pA/pF (n=17), 8.28±1.03 pA/pF (n=18), 24.5±3.6 pA/pF (n=12) and 9.0±2.3 pA/pF (n=15) respectively. In male rabbit myocytes, they were 0.84±0.07 pA/pF (n=18), 8.60±1.20 pA/pF (n=18), 25.9±4.5 pA/pF (n=14) and 9.3±2.6 pA/pF (n=16) respectively. I_K,tail in female rabbits was significantly lower than that of male rabbits (P<0.05), but there was no difference in I_to, I_K1 and I_Ca,L between male rabbits and female rabbits (P>0.05). The lower I_K,tail of female rabbit myocytes may contribute to the longer repolarization and the higher incidence of drug-associated torsade de pointes.

手动膜片钳检测盐酸罗哌卡因及其右旋异构体对HEK293细胞hERG电流的影响

目的研究比较盐酸罗哌卡因和盐酸罗哌卡因右旋异构体对高表达hERG钾通道的HEK293细胞hERG电流的影响。方法用手动膜片钳检测转染后hERG钾通道稳定表达的HEK293细胞电流,多菲莱德做阳性药,将盐酸罗哌卡因和盐酸罗哌卡因右旋异构体依次稀释成30.00、10.00、3.33、1.11、0.37μmol·L^(-1),依次作用于细胞,记录电流变化,计算抑制率。结果盐酸罗哌卡因0.37、1.11、3.33、10、30μmol·L^(-1)对电流Iherg-tail的抑制率分别为(6.12±0.30)%、(13.04±1.20)%、(19.21±0.33)%、(35.56±0.66)%、(65.37±4.17)%,IC_(50)为19.482μmol·L^(-1)(n=15)。盐酸罗哌卡因右旋异构体0.37、1.11、3.33、10.00、30.00μmol·L^(-1)对电流Iherg-tail的抑制率分别为(4.13±3.43)%、(7.34±5.60)%、(9.49±2.75)%、(16.60±0.87)%、(31.36±1.45)%,IC_(50)>30μmol·L^(-1)(n=15)。阳性对照药品多菲莱德0.00185、0.00556、0.01667、0.05000、0.15000μmol·L^(-1)对电流Iherg-tail的抑制率分别为(7.81±2.77)%、(19.67±1.88)%、(57.16±4.39)%、(89.71±3.55)%、(99.66±0.89)%、IC_(50)为0.015μmol·L^(-1)(n=15)。结论和阳性对照药品多菲莱德比较,盐酸罗哌卡因对hERG通道为弱抑制作用,盐酸罗哌卡因右旋异构体对hERG通道为无明显抑制作用。

Effects of Lizhong Tang on cultured mouse small intestine interstitial cells of Cajal

AIM:To investigate the effects of Lizhong Tang,an herbal product used in traditional Chinese medicine,on mouse small intestine interstitial cells of Cajal(ICCs).METHODS:Enzymatic digestions were used to dissociate ICCs from mouse small intestine tissues.The ICCs were morphologically distinct from other cell types in culture and were identified using phase contrast microscopy after verification with anti c-kit antibody.A whole-cell patch-clamp configuration was used to record potentials(current clamp) from cultured ICCs.All of the experiments were performed at 30-32 ℃.RESULTS:ICCs generated pacemaker potentials,and Lizhong Tang produced membrane depolarization in current-clamp mode.The application of flufenamic acid(a nonselective cation channel blocker) abolished the generation of pacemaker potentials by Lizhong Tang.Pretreatment with thapsigargin(a Ca 2+-ATPase inhibi-tor in the endoplasmic reticulum) also abolished the generation of pacemaker potentials by Lizhong Tang.However,pacemaker potentials were completely abolished in the presence of an external Ca 2+-free solution,and under this condition,Lizhong Tang induced membrane depolarizations.Furthermore,When GDPβ-S(1 mmol/L) was in the pipette solution,Lizhong Tang still induced membrane depolarizations.In addition,membrane depolarizations were not inhibited by chelerythrine or calphostin C,which are protein kinase C inhibitors,but were inhibited by U-73122,an active phospholipase C inhibitors.CONCLUSION:These results suggest that Lizhong Tang might affect gastrointestinal motility by modulating pacemaker activity in interstitial cells of Cajal.

K^+channels inhibited by hydrogen peroxide mediate abscisic acid signaling in Vicia guard cells

A number of studies show that environmental stress conditions increase abscisic acid (ABA) and hydrogen peroxide (H2O2) levels in plant cells. Despite this central role of ABA in altering stomatal aperture by regulating guard cell ion transport, little is known concerning the relationship between ABA and H2O2 in signal transduction leading to stomatal movement. Epidermal strip bioassay illustrated that ABA- inhibited stomatal opening and ABA-induced stomatal closure were abolished partly by externally added catalase (CAT) or diphenylene iodonium (DPl), which are a H2O2 scavenger and a NADPH oxidase inhibitor respectively. In contrast, internally added CAT or DPI nearly completely or partly reversed ABA-induced closure in half-stoma. Consistent with these results, whole-cell patch-clamp analysis showed that intracellular application of CAT or DPI partly abolished ABA-inhibited inward K+ current across the plasma membrane of guard cells. H2O2 mimicked ABA to inhibit inward K+ current, an effect which was reversed by the addition of ascorbic acid (Vc) in patch clamping micropipettes. These results suggested that H2O2 mediated ABA-induced stomatal movement by targeting inward K+ channels at plasma membrane.

Tyrosine hydroxylase-positive cells and dopaminergic neuronal function in human embryonic stem cells: An electrophysiological validation

BACKGROUND： Induced differentiation strategies and cytochemical properties of human embryonic stem ceils （hESCs） have been investigated. However, the electrophysiological functions of tyrosine hydroxylase （TH）-positive cells dedved from hESCs remain unclear. OBJECTIVE： To investigate the differentiation efficiency of TH-positive cells from hESCs in vitro using modified four-step culture methods, including embryoid body formation, and to examine the functional characteristics of the differentiated TH-positive cells using electrophysiological techniques. DESIGN, TIME AND SETTING： Neuroelectrophysiology was performed at the Reproductive Medicine Center and Stem Cell Research Center, Peking University Third Hospital, and the Neuroscience Research Institute and Department of Neurobiology, Peking University, from September 2004 to August 2008. MATERIALS： The hESC line, PKU-1.1, a monoclonal cell line derived from a pre-implantation human blastocyst in the Reproductive Medical Center of Peking University Third Hospital. The patch clamp recording system was provided by the Neuroscience Research Institute and Department of Neurobiology, Peking University. METHODS： The hESC line was induced to differentiate into TH-positive cells in vitro using a modified four-step culture method, including the formation of embryoid body, as well as the presence of sonic hedgehog and fibroblast growth factor 8. The cell karyotype was assessed by G-banding karyotype analysis techniques and specific markers were detected immunocytochemically. Whole-cell configuration was obtained after obtaining a tight seal of over 1 GΩ. Ionic currents were detected by holding the cells at -70 mV and stepping to test voltages between -80 and 40 mV in 10-mV increments in voltage-clamp configuration. MAIN OUTCOME MEASURES： We measured the cell karyotype, specific cell markers, and the electrophysiological properties of the voltage-gated ion channels on the cell membrane of TH-positive dopaminergic cells differentiated from our hESCs line in vitro. RESULTS： The differentiated cells had a consistent appearance, and the majority of cells （〉 90%） expressed TH and β-tubulion, as well as the neural progenitor marker, nestino Cell karyotype analysis demonstrated that all of the hESCs had a stable and normal karyotype （46, XX） after differentiation. In addition, patch clamp recording showed that the 10 recorded TH-positive cells exhibited a fast inward current when the test voltage depolarized to -30 mV, and a delayed outward current when the test voltage depolarized to -10 mV. The peak of inward current was obtained at voltage between 10 mV and 0 mV, while the peak of outward current was obtained at 40 mV. The average peak of inward current density was （ -50.05 ± 15.50） pA/pF, and the average peak of outward current density was （41.98 ± 13.55） pA/pE CONCLUSION： More than 90% of the differentiated hESC-derived cells induced by the modified four-step culture method exhibit dopaminergic neuronal properties, including general electrophysiological functional properties, such as functional potassium and sodium channels.

Effects of Atractylodes Macrocephala on the Cytomembrane Ca^(2+)-activated K^+ Currents in Cells of Human Pregnant Myometrial Smooth Muscles

The study examined the inhibitory effect of Atractylodes macrocephala （AM） on the uterine contraction during premature delivery and explored its electrophysiological mechanism by studying the effects of AM on the Ca^2＋-activated K^＋ currents of pregnant human myometrial smooth muscle cells with or without the treatment with intedeukin-6. Single cells were acutely isolated from pregnant human myometrial smooth muscles. Whole-cell Ca^2＋-activated K^＋ currents were recorded by using an Axopatchl-D amplifier. The cells were divided into three groups： group A in which AM was added into perfusate, group B, in which interleukin-6 was added into perfusate） and group C in which AM was added into perfusate after addition of interleukin-6. IL-6 10 ng/mL inhibited BKca by 36.9%±13.7% as compared with control （P〈0.01）. AM at 2 mg/mL raised BKca by 36.7%±22.6% or 45.2%±13.7% with or without the treatment of IL-6, respectively （P〈0.01）. It is concluded that AM was able to enhance the BKca of pregnant human myometrial smooth muscle cells treated or untreated with interleukin-6 and its effect on the BKca IL-treated cells was stronger that its effect on BKca of untreated cells. Our results suggested that AM can help to maintain the membrane potentials and the resting status of pregnant human myometrial smooth muscle cells.

Inhibitory Effects of Neferine on Na_v1.5 Channels Expressed in HEK293 Cells

Neferine, a bisbenzylisoquinoline alkaloid in Lotus Plumule, was proved to have a wide range of biological activities. In the present study, using whole-cell patch-clamp technique, we investigated the effects of neferine on Nav1.5 channels that are stably expressed in HEK 293 cells. We found that neferine potently and reversibly inhibited Nav1.5 currents in a concentration dependent manner with a half-maximal inhibition（IC50） being 26.15 μmol/L. The inhibitory effects of neferine on Nav1.5 currents were weaker than those of quinidine at the same concentration. The steady-state inactivation curve was significantly shifted towards hyperpolarizing direction in the presence of 30 μmol/L neferine, while the voltage-dependent activation was unaltered. Neferine prolonged the time to peak of activation, increased the inactivation time constants of Nav1.5 currents and markedly slowed the recovery from inactivation. The inhibitory effect of neferine could be potentiated in a frequency-dependent manner. These results suggested that neferine can block Nav1.5 channels under the open state and inactivating state and it is an open channel blocker of Nav1.5 channels.

Effects of antigliomatin from the scorpion venom of Buthus martensii Karsch on chloride channels on C6 glioma cells

Using whole-cell patch-clamp recordings, the effects of antigliomatin were observed on chloride channels on C6 glioma cells cultured in vitro. Antigliomatin was extracted from the venom of the scorpion Buthus martensii Karsch. Chloride channels are closed under normal osmotic pressure. When osmotic pressure was reduced to 120, 110 and 100 mV, the cell volume enlarged, chloride channels opened, and the chloride channel current increased. Three minutes after antigliomatin treatment, the chloride channel current decreased in a dose-dependent manner. These results show that antigliomatin extracted from the venom of the scorpion Buthus martensii Karsch diminishes chloride channel currents on C6 glioma cells.

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

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.

Caribbean maitotoxin elevates [Ca^(2+)]i and activates non-selective cation channels in HIT-T15 cells

AIM:To investigate the cytotoxic mechanism of caribbean maitotoxin(MTX-C) in mammalian cells.METHODS:We used whole-cell patch-clamp techniques and fluorescence calcium imaging to determine the cellular toxic mechanisms of MTX-C in insulin secreting HIT-T15 cells,which is a system where the effects of MTX have been observed.HIT-T15 cells stably express L-type calcium current,making it a suitable model for this study.Using the fluorescence calcium indicator Indo-1 AM,we found that there is a profound increase in HIT-T15 intracellular free calcium 3 min after application of 200 nmol/L MTX-C.RESULTS:About 3 min after perfusion of MTX-C,a gradual increase in free calcium concentration was observed.This elevation was sustained throughout the entire recording period.Application of MTX-C did not elicit the L-type calcium current,but large cationiccurrents appeared after applying MTX-C to the extracellular solution.The current-voltage relationship of the cation current is approximately linear within the voltage range from-60 to 50 mV,but flattened at voltages at-80 and-100 mV.These results indicate that MTX-C induces a non-voltage activated,inward current under normal physiological conditions,which by itself or through a secondary mechanism results in a large amount of cationic influx.The biophysical mechanism of MTX-C is different to its isoform,pacific maitotoxin(MTX-P),when the extracellular calcium is removed.CONCLUSION:We conclude that MTX-C causes the opening of non-selective,non-voltage-activated ion channels,which elevates level of intracellular calcium concentration and leads to cellular toxicities.

Changes in intrinsic excitability of ganglion cells in degenerated retinas of RCS rats

AIM：To evaluate the intrinsic excitability of retinal ganglion cells（RGCs） in degenerated retinas.METHODS：The intrinsic excitability of various morphologically defined RGC types using a combination of patch-clamp recording and the Lucifer yellow tracer in retinal wholemount preparations harvested from Royal College of Surgeons（RCS） rats,a common retinitis pigmentosa（RP） model,in a relatively late stage of retinal degeneration（P90） were investigated.Several parameters of RGC morphologies and action potentials（APs） were measured and compared to those of non-dystrophic control rats,including dendritic stratification,dendritic field diameter,peak amplitude,half width,resting membrane potential,AP threshold,depolarization to threshold,and firing rates.RESULTS：Compared with non-dystrophic control RGCs,more depolarizations were required to reach the AP threshold in RCS RGCs with low spontaneous spike rates and in RCS OFF cells（especially A2o cells）,and RCS RGCs maintained their dendritic morphologies,resting membrane potentials and capabilities to generate APs.CONCLUSION：RGCs are relatively well preserved morphologically and functionally,and some cells are more susceptible to decreased excitability during retinal degeneration.These findings provide valuable considerations for optimizing RP therapeutic strategies.