流式细胞术测定参附注射液对吗啡依赖神经细胞内Ca2+浓度的影响

目的观察参附注射液对吗啡依赖人神经母细胞瘤细胞SK-N-SH胞内Ca2+浓度的影响,探讨其戒毒作用的细胞机制.方法建立吗啡依赖神经细胞模型,Fluo-3着染细胞后,应用流式细胞仪测定参附注射液对神经细胞内Ca2+浓度的影响.结果参附注射液中、高剂量组神经细胞内Ca2+浓度显著升高,纳洛酮催促引起的细胞内Ca2+的降低抑制.结论参附注射液可能通过调节细胞内Ca2+浓度发挥其戒毒作用.     

电针百会穴对大鼠缺血区脑组织神经生长因子影响的研究

目的:研究针刺"百会"穴对局灶性脑缺血大鼠神经生长因子的影响.方法:凝闭大鼠一侧大脑中动脉10h后致局灶性脑缺血,采用免疫组化染色SP法进行观察.结果:电针治疗能使局灶性脑缺血大鼠脑组织神经生长因子免疫阳性表达在细胞数量上及强度上明显增强(P＜0.01),从而阻止神经细胞内Ca2 超载,稳定细胞内环境.结论:电针对缺血性脑损伤具有一定的保护作用,为临床针灸治疗缺血性脑血管疾病奠定了理论基础.     

补阳还五汤对全脑缺血模型大鼠再灌注不同时点皮质神经细胞钙超载的影响

目的:采用膜片钳技术和离子图像分析技术,同步观察补阳还五汤预干预对全脑缺血大鼠皮质神经细胞L型Ca2 通道及细胞内Ca2 浓度的影响,探讨Ca2 信号异常参与缺血性神经细胞损伤的机制及补阳还五汤抗脑缺血的分子机制,为临床应用本方防治缺血性脑病提供理论基础,并为中药复方研究探索新的思路和方法.方法:参照改良的Pulsinelli四血管闭塞法复制全脑缺血大鼠模型,缺血后大鼠分别在存活2,2,4,8及72 h后进行皮质神经细胞急性分离,负载荧光指示剂Fura-2/AM,用钙离子成像系统检测单个神经细胞内Ca2 浓度.单通道电流经EPC-9膜片钳放大器放大,用Pulsefit Pulse采集至计算机.用分析软件TAC进行测量.结果:补阳还五汤预干预除再灌注12 h外,可显著降低其他各时点大脑皮质神经细胞内Ca2 浓度,对缺血再灌后两次Ca2 堆积均表现明显的抑制作用.补阳还五汤再灌注72 h,皮质神经细胞L型Ca2 通道平均开放时间显著降低,再灌注2 h,皮质神经细胞L型Ca2 通道开放概率显著降低.结论:降低缺血再灌注不同时点皮质神经细胞内Ca2 浓度及在缺血再灌注不同时点分别降低L型Ca2 通道开放概率和开放时间是补阳还五汤抗脑缺血的重要机制.补阳还五汤预干预对缺血性神经细胞损伤的预防和治疗有非常积极的意义.     

Orexin-A对体外培养的大鼠海马神经细胞凋亡的影响

目的:研究Orexin-A对体外培养的海马神经细胞凋亡的影响及其可能机制.方法:取体外培养的大鼠海马神经细胞分为7组,分别给予0,1.0,10.0,100.0 nmol/L及1.0、10.0、100.0 μmol/L的Orexin-A培养.另取大鼠海马神经细胞,预先加入Orexin-A受体(OX1R)阻断剂SB334867,然后同法分组及处理.观察海马神经细胞的形态学变化;采用Western Blot、逆转录PCR(RT-PGR)法检测OX1R蛋白和mRNA的表达;原位末端标记染色评价细胞凋亡率;Fura-2/AM荧光比值成像技术测定游离钙离子浓度[Ca2+]i.结果:随着Orexin-A浓度的增大,海马神经细胞凋亡增加(F=137.49,P＜0.01),同时呈剂量依赖性地触发[Ca2+]i的升高(F=961.44,P＜0.01),但是OX1R蛋白及mRNA的表达无明显变化.SB334867可以部分阻断这种现象.结论:大剂量Orexin-A对体外培养的海马神经细胞有促凋亡作用,其机制可能与增加了细胞内Ca2+浓度蓄积有关.     

定心方对大鼠缺氧及正常心肌细胞内[Ca~(2+)]_i、细胞膜电位和线粒体膜电位的影响

目的 :探讨定心方抗心律失常的分子机制。方法 :大鼠击晕后取心脏 ,用链酶蛋白酶消化分离单个心肌细胞 ,再用不同荧光染料分别标记细胞 ,在激光共聚焦显微镜上测定定心方血清对心肌细胞内 [Ca2 + ]i、细胞膜电位和线粒体膜电位的影响。结果 :缺氧使心肌细胞内钙浓度和线粒体膜电位升高 ,而使细胞膜电位降低 ,定心方血清降低了正常和缺氧心肌细胞内 [Ca2 + ]i,改善了缺氧引起的膜电位降低 ,使缺氧状态下细胞线粒体膜电位保持在基线水平。结论 :定心方使细胞内 [Ca2 + ]i 降低 ,维持细胞膜电位和线粒体膜电位接近于正常水平     

三七总皂甙对吗啡戒断大鼠海马神经细胞内游离钙的影响

目的 研究三七总皂甙(panax notoginseng saponins,PNS)对吗啡戒断大鼠海马神经细胞内[Ca2+]i的影响,探讨PNS缓解吗啡戒断症状的可能机制.方法 应用剂量递增法大鼠皮下注射吗啡及腹腔注射纳络酮建立吗啡躯体依赖及催促戒断模型.在给予吗啡的同时分别以PNS 100、200、400 mg/kg对大鼠灌胃,记数大鼠的体质量变化和跳跃次数以确定模型建立是否成功,采用流式细胞术测定其海马神经细胞内[Ca2+]i浓度.结果 ①PNS呈剂量依赖性抑制戒断大鼠体质量减轻和跳跃的发生,②慢性吗啡作用可以使海马神经细胞内[Ca"]i明显升高;纳络酮可迅速下词这种异常增高的[Ca2+]i;PNS则可增加纳络酮催促戒断大鼠海马神经细胞内[Ca2+]i,且呈剂量依赖性.结论 PNS可能通过调节神经细胞内[Ca2+]i,抑制吗啡依赖大鼠催促戒断症状的发生.     

酸枣仁汤含药血清对皮质酮损伤的PCI2细胞[Ca2+]i、CaM的影响

目的:观察酸枣仁汤含药血清对皮质酮损伤的PC12细胞[Ca2+]i、CaM的影响.方法:以高浓度皮质酮诱导PC12细胞凋亡模拟焦虑症神经细胞损伤状态.通过激光共聚焦显微镜法检测[Ca2+]i浓度、细胞化学染色法测定CaM的表达,探讨酸枣仁汤含药血清对皮质酮诱导PC12细胞凋亡的钙信号转导通路的影响.结果:与空白对照组比较,皮质酮组细胞内[Ca2+]i浓度显著升高;与皮质酮+正常动物血清组比较,皮质酮+酸枣仁汤含药血清组细胞内[Ca2+i浓度显著降低,皮质酮+安定含药血清组'皮质酮+酸枣仁汤含药血清组细胞CaM表达明显下降.结论:胞内钙超载可能是皮质酮诱导PC12细胞凋亡的主要原因之一,酸枣仁汤含药血清可能通过减轻胞内钙超载,减少Ca2+-CaM复合物的产生,拮抗皮质酮诱导的PC12细胞凋亡.     

烧伤血清对大鼠腹腔巨噬细胞内游离钙浓度及其产生NO的影响

目的:观察烧伤血清对分离的大鼠腹腔巨噬细胞内游离钙离子浓度([Ca2+]i)和产生NO的影响.方法:收集腹腔巨噬细胞,用Fura-2/AM作为细胞内游离钙离子指示剂测定[Ca2+]i,用烧伤血清刺激体外的腹腔巨噬细胞即刻进行[Ca2+]i的测定;用同浓度的烧伤血清刺激培养的腹腔巨噬细胞,测定上清NO稳定的代谢产物[NO2+NO3-]的浓度.结果:烧伤血清能显著地促进巨噬细胞[Ca2+]i的增高,以烧伤后6 h血清的作用最为明显,可达正常对照组的9倍,维拉帕米可部分抑制其作用.烧伤血清对腹腔巨噬细胞NO的产生有明显的抑制作用,也以6 h烧伤血清为最明显.结论:烧伤血清能显著地升高腹腔巨噬细胞[Ca2+]i,而NO的产生被明显地抑制.细胞内钙超载和NO产生减少可能是烧伤后早期巨噬细胞免疫功能损伤主要因素之一.     

烫伤血清对大鼠库普弗细胞胞浆游离钙浓度的影响

目的:观察烫伤血清对大鼠分离培养的库普弗细胞(KC)胞浆内游离钙([Ca2+]i)浓度的影响.方法:胶原酶灌注消化、梯度离心分离KC,在KC悬液中加入烫伤血清,Fura-2/AM荧光染色法测定[Ca2+]i.结果:分离培养的KC静息状态下[Ca2+],为(109.29±4.17)nmol/L,烫伤血清可以使[Ca2+]i显著升高,以烫伤后6 h的血清作用最明显;[Ca2+]i的浓度随血清浓度的升高而升高;烫伤血清作用1 min KC[Ca 2+]i已明显升高,2 min达高峰,20 min时仍明显高于假烫组.结论:烫伤血清能明显升高KC[Ca2+]i,作用快速而持久,且呈浓度依赖性.     

Na+,K+-ATP酶参与缺氧所致大鼠皮质神经元内钙升高

目的:探讨Na ,K -ATP酶对缺氧所致大鼠皮质神经元内钙升高的影响.方法:采用激光共聚焦显微镜及可视化动缘探测系统,测定培养大鼠皮质神经元在缺氧不同时间和双氢哇巴因(DHO,一种Na -K -ATP酶抑制剂)不同浓度时细胞内钙离子浓度([Ca2 ]i)和细胞内钠离子浓度([Na ]i),并观察DHO对缺氧后神经元[Ca2 ]i和[Na ]i升高作用的影响.结果:DHO(10-9～10-3mol/L)和缺氧(4～20 min)均可显著升高正常皮质神经元[Ca2 ]i和[Na ]i,并分别具有剂量依赖性和时间依赖性.在缺氧4 min时皮质神经元[Ca2 ]i和[Na ]i即明显升高,此时给予DHO 10-3mol/L可使二者进一步升高;但当皮质神经元缺氧15 min后,再给予相同剂量的DHO则不能使[Ca2 ]i和[Na ]i进一步增加;若对DHO(10-3mol/L)预先升高[Ca2 ]i的正常神经元再进行缺氧灌流,则[Ca2 ]i不继续升高.结论: Na ,K -ATP酶抑制是缺氧所致大鼠皮质神经元[Ca2 ]i升高的机制之一.     

钙离子选择性微电极在钙制剂药代动力学研究中的应用

目的 用自制的钙离子选择性微电极（Ca^2 -ISME）测定游离钙,拟反映体内钙被吸收的真实水平。方法 建立了血清样品中钙离子测定方法,并进行三种具有代表意义的站钙剂的大鼠药代动力学对比研究。经灌胃给药,定时取血,制备血清并测定其钙离子浓度。结果 三种制剂在血中的达峰时间不完全一致,但其AUC间经t检验无显著性差异。结论 C^2 -ISME可用于钙制剂的药物代谢动力学研究。     

针刺留针与否对脑缺血再灌注大鼠IL-6表达的影响

目的:通过测定局灶性脑缺血再灌注大鼠血清白介素-6(IL-6)的含量,研究针刺留针与否对早期脑缺血再灌注大鼠脑的保护差异。方法:40只SD大鼠随机分为正常组、假手术组、大脑中动脉阻塞(MCAO)再灌注组、针刺留针及不留针组。针刺留针组在造模后2 h、24 h固定,针刺三阴交、内关、人中,留针30 min,针刺不留针组针刺时间、穴位均同针刺留针组,针刺后即起针而不留针。造模48 h后,测定各组大鼠血清中IL-6的含量。结果:针刺能升高脑缺血再灌注大鼠血清中IL-6的含量。早期针刺后留针与否对缺血再灌注大鼠血清中IL-6水平影响差异无统计学意义(P>0.05)。结论:针刺能升高大鼠脑缺血再灌注后血清中IL-6含量,对缺血组织起到抗损伤、促修复的作用;在缺血再灌注早期,针刺后留针与不留针可能对脑的保护作用并无差异。     

尼莫地平对脑挫裂伤后神经细胞内游离钙浓度的影响

目的〓〖HTK〗研究尼莫地平对脑挫裂伤后神经细胞胞浆内游离钙离子浓度变化的影响。〖HTW〗方法〓〖HTK〗将Wistar大鼠随机分为对照组、创伤组、治疗组；创伤组按照Feeney自由落体脑损伤模型方式制作大鼠左顶叶脑挫裂伤模型，治疗组在外伤后开始静脉滴注尼莫地平。分别对脑挫裂伤后0.5、1、3、6、12、24、48h各时相点细胞内游离钙离子浓度和细胞凋亡率进行检测。对不同时相的创伤组、治疗组和对照组进行比较分析。〖HTW〗结果〓〖HTK〗脑挫裂伤后大脑皮层神经细胞内游离钙离子浓度迅速升高，0.5h时为正常值的4倍，24h时达到高峰，48h时降低明显，但仍然维持在比较高的水平。应用尼莫地平治疗后，钙离子浓度显著降低，6h时降至接近正常水平。脑挫裂伤后神经细胞凋亡率和胞内游离钙离子浓度存在相关性。〖HTW〗结论〓〖HTK〗脑挫裂伤后神经细胞胞浆内钙离子浓度升高，导致钙超载并引起细胞凋亡。早期应用尼莫地平能抑制钙超载，抑制细胞凋亡，对神经细胞具有极强的保护作用。     

The effects of local anesthetics on intracellular Ca2+ release from ryanodine-sensitive Ca2+ stores in gerbil hippocampal neurons

Objective To examine the effects of procaine and lidocaine on intracellular Ca(2+) release from sarcoplasmic reticulum ryanodine-sensitive Ca(2+) stores. Methods The experiment was performed on hippocampal slices from 60-80 g male Mongolian gerbils. Levels of intracellular Ca(2+) concentration in the slices were measured by microfluorometry. The slices were perfused with 50 mmol/L KCl containing medium for 30 seconds. Then, the medium was switched to physiological medium. After 5 min of incubation, the slice was perfused with 20 mmol/L caffeine containing physiology medium for 2 min. Following incubation, the slice was superfused with physiological medium until the end of the experiment. The effects of procaine and lidocanin (100 μmol/L) on caffeine-evoked Ca(2+) release were evaluated by adding them to the medium after high K(+) medium perfusion. Results Caffeine induced a marked increase in intracellular Ca(2+) concentration which was then decreased 12% upon the addition of procaine (P<0.05); however, lidocaine, did not induce a similar inhibitory reaction.Conclusion Procaine inhibits ryanodine-receptor mediated Ca(2+) release from intracellular Ca(2+) stores, while lidocaine may inhibit Ca(2+) release through other mechanisms.     

Influence of Needling the Foot-Yangming Points on Intraceilular Ca^2＋ Concentration in Smooth Muscles of the Gastric Antrum in Rabbits

Objective： To investigate the influence of acupuncture at the points of Foot-Yangming Meridian on intracellular concentration of Ca^2, called the 2nd messenger of gastric smooth muscles. Methods： 45 rabbits were randomly divided into the following 5 groups： a normal saline group, a model group treated with atropine, an acupuncture group treated by needling the points of Foot-Yangming Meridian, an acupuncture group treated by needling the points of Foot-Shaoyang Meridian, an acupuncture group treated by needling the points of Foot-Taiyang Meridian, i.e. 9 rabbits in each group. After treatment, the smooth muscles of the gastric antrum were taken to make the suspension containing alive single muscular cells, and the intracellular calcium concentration （[Ca^2＋]i） was determined by a spectrofluorometer. Results： The concentration of [Ca^2＋]i in the group of Foot-Yangming Meridian was obviously higher than that of the atropine group （P〈0.01）, but with no significant differences found among all the other groups （P〉0.05）. Conclusion： The influence of acupuncture at the points of Foot-Yangming Meridian on gastric movement is related to the release of intracellnlar Ca^2＋ in the gastric smooth muscles     

柴芩承气汤对急性胰腺炎大鼠胰腺腺泡细胞的保护作用及其机制

目的：探讨柴芩承气汤（Chaiqing Chengqi Decoction,CQCQD）对急性胰腺炎（acute pancreatitis,AP）大鼠胰腺腺泡细胞的保护作用及其机制。方法：SD大鼠以CQCQD灌喂,制备柴芩承气汤含药血清（CQCQserum,CQCQS）;SD大鼠分为AP组和假手术组,分离胰腺腺泡细胞并与CQCQS共同孵育,采用甲基噻唑基四唑（methyl thiazolyl tetrazolium,MTT）法检测不同浓度CQCQS对腺泡细胞存活力的影响,并用钙荧光指示剂Fluo-3-AM加载腺泡细胞,激光共聚焦显微镜直接观测腺泡细胞内钙荧光强度（fluorescent intensity,FI）,定量分析FI并以此表示[Ca^2＋]i。结果：5%和10%CQCQS均可提高AP大鼠胰腺腺泡细胞存活力（P〈0.05）,且10%CQCQS提高细胞存活力的作用比5%CQCQS更为显著（P〈0.05）;[Ca^2＋]i随AP病程延长而升高（P〈0.05）,CQCQS可抑制AP大鼠胰腺腺泡细胞内[Ca^2＋]i升高（P〈0.05）。结论：CQCQD对AP时的胰腺细胞有保护作用,其机制可能与抑制胰腺腺泡细胞内钙超载有关。     

“贺氏三通法”不同时点针刺对脑缺血再灌注大鼠超氧化物歧化酶和丙二醛的影响

目的观察"贺氏三通法"不同时点针刺对脑缺血再灌注大鼠缺血区脑组织和血清超氧化物歧化酶(super oxide dismutase,SOD)和丙二醛(malondialdehyde,MDA)水平的影响。方法将成年雄性SD大鼠随机分为假手术组(8只)、模型对照组(10只)、针刺组(30只)。根据干预时间的不同,针刺组分为3h针刺组、6h针刺组、48h针刺组,每组各10只。采用线栓法复制大鼠脑缺血再灌注模型,针刺组在模型复制成功后分别按3、6、48h分组给予"贺氏三通法"针刺,每日1次。模型复制成功72h后测定血清及脑组织中SOD活性和MDA含量。结果与模型对照组比较,各针刺组大鼠脑组织SOD活性显著增加(P<0.01),6h针刺组血清SOD活性有升高趋势(P>0.05);而针刺组大鼠血清MDA含量显著增加(P<0.01)。6h针刺组大鼠血清SOD活性、MDA含量显著高于3h和48h针刺组(P<0.05,或P<0.01)。结论 "贺氏三通法"早期介入可调节SOD、MDA的代谢紊乱,从而对脑缺血再灌注模型大鼠脑组织起到保护作用。     

Positive correlation of tumor necrosis factor-alpha early expression in myocardium and ventricular arrhythmias in rats with acute myocardial infarction

BACKGROUND: Tumor necrosis factor-alpha (TNF-alpha) is found to play important roles in acute myocardial infarction (AMI). Ventricular arrhythmias arising from AMI are leading causes of sudden cardiac death (SCD). We sought to clarify the effect of TNF-alpha early expression on ventricular arrhythmias in rats with AMI and its mechanism. METHODS: Rats with AMI were induced by left anterior descending coronary branch ligation. The mRNA and protein levels of TNF-alpha in myocardium were detected by real-time fluorescent quantitative PCR, Western blotting and histochemistry. Meanwhile, electrocardiogram was recorded. Different concentrations of TNF-alpha were added to isolated rat hearts in isolated heart perfusions. Effect of TNF-alpha on intracellular Ca(2+) concentration was detected by laser confocal technique. RESULTS: In AMI rats, mRNA and protein levels of TNF-alpha were higher than control (p <0.05), and the occurrence time of ventricular arrhythmias coincided with the secretion of TNF-alpha. TNF-alpha may cause ventricular arrhythmias in isolated rat heart perfusion models. Intracellular Ca(2+) intensity may quickly be increased by TNF-alpha. CONCLUSIONS: Our results reveal the positive correlation between TNF-alpha early expression and ventricular arrhythmias in rats with AMI. This effect may be associated with the increased intracellular Ca(2+) intensity caused by TNF-alpha.     

简帛医书“却”字考释与相关医籍校读

简帛医书中有"却"字,与《灵枢·经脉》对读可知其所指部位与"腘"相对应,即膝后弯曲处。该"却"字在《素问》中作"郄",凡13见,皆指"郄中"而言,为膝后腘窝处,亦称"腘中""委中"。通过对"郄""却"形音义及其文献用例详细考察可知,"郄中"之"郄"当为"却"之讹写,而膝后弯曲处正为"却"之本义,故"却"引申有却曲义,引申有退却义,后作为连词亦有转折义。"脚"为"却"之后起字,在早期医籍中仍保留,有指膝后弯曲处或膝关节处的用例。由于"胠"字与"[月谷](却之异体)"形音相近,医籍中也有相混之例需加以辨析。另后世医籍中"郄中"之"郄"多见写作"郗"者,亦当为"却"之俗写体"[郗]"的一种讹体。     

肝细胞膜钙泵与钠泵活性变化在胆红素钙结石形成中的作用

OBJECTIVE: To investigate the effects of activities of Ca(2+)-ATPase and Na(+)-K(+)-ATPase in plasma membranes of hepatocytes on the formation of calcium bilirubinate gallstone. METHODS: The rabbit models for studying calcium bilirubinate gallstone were used. One hundred and three rabbits were randomly divided into the control (sham operation) group (Con, n = 28), the simple biliary obstruction group (BO, n = 36), and the biliary obstruction and infection group (BOI, n = 39). The activities of Ca(2+)-ATPase and Na(+)-K(+)-ATPase in plasma membranes of hepatocytes and the intracellular calcium content were measured on the 3rd, 7th, 14th and 20th days after operation. RESULTS: The activities of Ca(2+)-ATPase and Na(+)-K(+)-ATPase decreased remarkably in all phases of BOI and BO groups as compared with those of Con group (P < 0.01). The over-loaded intracellular calcium was found in both BOI and BO groups. The above-mentioned changes were more significant in BOI group than in BO group (P < 0.05). CONCLUSION: The progressive decrease of the activities of Ca(2+)-ATPase and Na(+)-K(+)-ATPase is in close relationship with the continuous increase of intracellular calcium content during the formation of calcium bilirubinate gallstone in rabbit models. Infection can aggravate those changes and further the formation of stone.