尼莫地平对脑损伤后神经细胞钙通道和超微结构的改变

采用细胞内Ｃａ２＋荧光探针Ｆｕｒａ－２／ＡＭ检测大鼠脑损伤后神经元突触体胞浆中游离Ｃａ２＋浓度（［Ｃａ２＋］ｉ），以研究神经细胞钙通道变化，并选用Ｃａ２＋通道阻断剂尼莫地平治疗，观察神经细胞Ｃａ２＋通道变化和超微结构的改变。结果表明，脑损伤后神经细胞钙通道开放，突触体胞浆中游离［Ｃａ２＋］ｉ在伤后０．５小时即已升高为１．０９±０．０８×１０－６Ｍ／Ｌ水平（Ｐ＜０．００１），伤后６、２４、４８和７２小时，［Ｃａ２＋］ｉ持续处于１０－６Ｍ／Ｌ水平以上。同时发现神经细胞Ｃａ２＋超载时，其超微结构损害。应用尼莫地平治疗后，神经细胞胞浆游离［Ｃａ２＋］ｉ明显下降，超微结构损害显著减轻。     

脑益嗪抗运动病时大鼠脑细胞钙离子超微结构定位的变化(论著)

目的：观察脑益嗪抗运动病时脑细胞钙离子（Ｃａ２＋）的变化并探讨其作用机理。方法：采用Ｃａ２＋超微结构定位方法，对ＳＤ大鼠预先给予脑益嗪后，采用正负交变加速度旋转刺激制备运动病模型。观察运动病大鼠和脑益嗪给药组大鼠大脑皮质、小脑皮质和脑干前庭区脑细胞Ｃａ２＋超微结构定位变化。结果：运动病大鼠大脑皮质、小脑皮质和脑干前庭区脑细胞质基质、线粒体和内质网中Ｃａ２＋反应产物增多。预先给予脑益嗪后，大脑皮质、小脑皮质和脑干前庭区脑细胞质Ｃａ２＋反应产物明显下降（Ｐ＜０．０１）。结论：脑益嗪抗运动病的中枢机理之一可能与抑制脑细胞Ｃａ２＋内流有关     

中长期模拟失重大鼠骨骼肌线粒体钙含量和肌浆网Ca^2＋—A…

为探讨失重对肌肉功能的影响，观察了中长期模拟失重时大鼠骨骼肌细胞内Ｃａｗ＾２＋转运功能变化，测定了比目鱼肌，腓肠肌线粒体钙含量和肌浆网（ＳＲ）Ｃａ＾２＋－ＡＴＰａｓｅ活性。结果是悬吊１５、３０ｄ大鼠比目鱼肌和腓肠肌线粒体Ｃａ＾＋２含量增加，肌浆网Ｃａ＾２＋－ＡＴＰ酶活性降低，说明中长期模拟失重肌细胞内Ｃａ＾２＋转运功能发生了改变，这可能是影响肌肉收缩特性的因素之一。     

力竭性游泳对大鼠心肌线粒体基质游离Ca~(2+)及Na~+/Ca~(2+)交换的影响

应用荧光Ｃａ２＋指示剂Ｆｕｒａ－２ＡＭ测定大鼠心肌线粒体基质游离Ｃａ２＋浓度变化，观察到力竭性游泳后心肌线粒体内游离Ｃａ２＋浓度下降（Ｐ＜０．０５），在介质中加０．５μｍｏｌ／　Ｌ　Ｃａ２＋时，表现出摄钙能力（Ｐ＜０．００１），但能力下降（Ｐ＜０．０５），对线粒体外Ｃａ２＋浓度变化的应答敏感性下降。在线粒体负载Ｃａ２＋条件下，Ｎａ＋依赖性Ｃａ２＋释放（Ｎａ＋／Ｃａ２＋交换）能力下降（Ｐ＜０．０５），因此线粒体不能及时反映心肌需能的变化，表现出氧化磷酸化的失调。文中探讨了线粒体Ｃａ２＋转动变化的机制和生理意义。     

脑缺血再灌流时〔^3H〕—三磷酸肌醇放射活性及突触体游离Ca^2…

研究大鼠脑血再灌流时〔＾３Ｈ〕－ＩＰ３放射活性及突触体（Ｃａ＾２＋）ｉ的变化，结果示缺血１ｍｉｎ就启动了肌醇脂质信使系统，引起〔＾３Ｈ〕－ＩＰ３显著增高，缺血２０ｍｉｎ突触体（Ｃａ＾２＋）ｉ增高，且持续至再灌流７ｄ，磷脂酶Ｃ抑制剂ＰＭＳＦ治疗能显著地抑制突触体（Ｃａ＾２＋）ｉ的升高，减轻海马ＣＡ１区缺血性神经元损伤。     

高温、噪声对飞行员红细胞膜ATP酶活性的影响

目的 探讨高温、噪声环境中飞行对飞行员红细胞膜ＡＴＰ酶活性的影响。方法 轰六飞行员２４名为实验组,普遍地面人员２１名为对照组。作多靶场轰炸飞行,舱内温度４５～５３℃,噪 ９７～１２２ｄＡ（Ａ）。飞行时间３ｈ,于飞行前（６：００ａ．ｍ．）飞行后即刻（１２：００ａ．ｍ．）、８ｈ（８：ｐ．ｍ）采血。对照组采血日行室内作业（２７℃）。用比色法测定红细胞膜Ｎａ＾＋－Ｋ＾＋ＡＴＰ酶、Ｃａ＾２＋－Ｍｇ＾２＋ＡＴ     

口服谷氨酰胺对烫伤大鼠小肠功能和结构的影响

测定烫伤大鼠早期口服谷氨酰胺（Ｇｌｎ）后血和组织中的游离氨基酸（ＦＡＡ）、二胺氧化酶（ＤＡＯ）、谷胱甘肽（ＧＨＳ）、肿瘤坏死因子（ＴＮＦ）和血浆内毒素（ＬＰＳ）的含量。结果表明，烫伤后１０ｈ、２ｄ、５ｄ、和８ｄ小肠ＤＡＯ水平降低，血浆ＤＡＯ升高；标准饲料（Ｃ）组血浆Ｇｌｎ在伤后２ｄ和５ｄ降低；Ｇｌｎ饲料（Ｇ）组各时点均增加，１０ｈ和８ｄ增加显著；Ｇｌｎ＋精氨酸（Ｇ＋Ａ）组在２ｄ降低。Ｃ组血浆丙氨酸、精氨酸和血氨较其它２组低。ＬＰＳ水平增高，但Ｃ与Ｇ组比较无统计学差异。病理检查见Ｃ组在伤后１０ｈ、５ｄ和８ｄ小肠绒毛萎缩变短，固有层慢性炎性细胞增多，肠壁变薄，Ｃ组和Ｇ＋Ａ组小肠粘膜厚度接近正常。提示口服Ｇｌｎ可能对烫伤大鼠小肠粘膜结构和功能有保护作用。     

兔胸部撞击伤时中性粒细胞凋亡与肺损伤的关系

目的：ＰＭＮ在炎症的发生、发展和转归中起着至关重要的作用,本实验动态地观察兔胸部撞击伤时ＰＭＮ凋亡的发生以及与肺损伤之间的关系。方法：制备兔胸部撞击伤模型,分离纯化灌洗液中ＰＭＮ,应用流式细胞术测定凋亡、坏死、存尖细胞比例及呼吸爆发功能的变化,并且观察与ＬＤＨ和胸浆游离Ｃａ＾２＋变化之间的关系。结果：肺灌洗液中ＰＭＮ的凋亡延迟持续至１２ｈ,在伤后各时相点活细胞增多。而灌洗液ＰＭＮ呼吸爆发从２ｈ即显著增强,８ｈ达到峰值。同时,灌洗液ＬＤＨ的升高从４ｈ￣２４ｈ显著高于对照。伤后ＰＭＮ胞浆游离Ｃａ＾２＋有短暂升高。结论：ＰＭＮ在肺组织中大量扣押并且正常的凋亡途径发生障碍,造成ＰＭＮ持续处于激活状态及毒性内容物的持续释放,与肺组织损伤有密切关系,并且ＰＭＮ凋亡延迟可能与Ｃａ＾２＋的短暂升高有关。     

口服谷氨酰胺对烫伤大鼠功能和结构的影响

测定烫伤大鼠早期口服谷氨酰胺（Ｇｌｎ）后血和组织中的游离氨基酸（ＦＡＡ）、二胺氧化酶（ＤＡＯ）、谷胱甘肽（ＧＨＳ）、肿瘤坏死因子（ＴＮＦ）和血浆内毒素（ＬＰＳ）的含量。结果表明，烫伤后１０ｈ、２ｄ、５ｄ和８ｄ小肠ＤＡＯ水平降低，血浆ＤＡＯ升高；标准饲料（Ｃ）组血浆Ｇｌｎ在伤后２ｄ和５ｄ降低；Ｇｌｎ饲料（Ｇ）组各时点均增加，１０ｈ和８ｄ增加显著；Ｇｌｎ＋精氨酸（Ｇ＋Ａ）组在２ｄ降低。Ｃ组血浆丙氨酸、     

氟桂利嗪,倍他司汀对膜迷路积水豚鼠蜗内电位及Ca^2＋浓度的影响

４５只豚鼠随机分为３组，造成膜迷路积水的模型，对照组右耳作空白对照，两用药组的豚鼠分别于造模后第１天口服大剂量氟桂利嗪（１０ｍｇ／ｋｇ，１／日）及倍他司汀（１０ｍｇ／ｍｇ，２／日）３０天，发现在造模３０天后，实验对照组听性脑干反应（ＡＢＲ）阈值增高，向术侧摆动试验眼震（ＳＰＶＮ）频数下降，蜗内电位（ＥＰ）下降，内淋巴Ｃａ＾２＋浓度增高，ＥＴ和Ｃａ＾２＋呈负相关，氟桂利嗪和倍他司汀均能显著抑制前庭性     

创伤性脑损伤神经细胞钙超载机制中大电导钙激活钾通道的作用

目的 探讨大电导钙激活钾通道在创伤性脑损伤神经细胞钙超载机制中的作用.方法 体外培养小鼠大脑皮层神经元细胞,应用膜片钳技术,分别在静息条件下和持续电流去极化条件下,采用自身对照观察大电导钾通道特异性阻断剂Iberiotoxin灌流前后,神经元细胞内游离钙浓度和动作电位频率的变化,并按随机数字表法分成实验组(加Iberiotoxin)和对照组,采用显微荧光测钙法,观察Iberiotoxin对高钾条件下神经细胞游离钙浓度的影响.结果 静息条件下,Iberiotoxin灌流(100 nmmol/L)对神经元细胞自发动作电位的频率、游离钙浓度无显著影响.持续去极化电流刺激实验中,Iberiotoxin灌流前动作电位频率为(10.4±3.0)Hz,灌流后为(13.8±3.7)H2(P＜0.05);Iberiotoxin灌流前游离钙浓度较静息值高(14.21±16.98)nmol/L,灌流1 min后游离钙浓度较静息值高(44.07±34.4)nmol/L,比灌流前显著增加(P＜0.05).高钾溶液(20 mmol/L KCl)诱导神经元游离钙浓度升高,而Iberiotoxin灌流(100 nmmol/L)则使游离钙浓度进一步显著升高.结论 生理条件下,大电导钾通道不参与神经元游离钙浓度和兴奋性的调节;但在受持续去极化刺激或高钾条件时,阻断大电导钾通道对神经元游离钙浓度和兴奋性具有显著的影响,提示大电导钾通道参与神经细胞游离钙水平的调节,在创伤性脑损伤神经细胞钙超载机制中发挥作用.     

Effects of contrast media on calcium transients and motion in cultured ventricular cells

To investigate the mechanisms of the negative inotropic effects of contrast media, we superfused spontaneously contracting cultured chick embryo ventricular cells with Renografin-76 and iohexol (12% solutions), and hypertonic sucrose during simultaneous measurement of [Ca2+]i transients (indo-1) and motion (video-motion detector system). Exposure to contrast agents caused a significant reduction of contractility, with Renografin-76 having a much greater effect on amplitude of motion than iohexol. Renografin-76 significantly depressed [Ca2+]i transient amplitude, whereas iohexol had no effect. Addition of Ca2+ to correct for calcium binding by Renografin-76 completely reversed its depression of [Ca2+]i transients but only partially reversed the negative inotropic effects. Hypertonic sucrose caused a significant decrease in contraction amplitude, with no significant effects on [Ca2+]i transient amplitude. We conclude that the marked negative inotropic effect of Renografin-76 is caused by both calcium binding and hypertonicity. The less marked depression of contractility produced by iohexol likely is a result of hypertonicity and not caused by alteration of [Ca2+]i.     

Calcium spiking activity and baseline calcium levels in ROS 17/2.8 cells exposed to extremely low frequency electromagnetic fields (ELF EMF)

PURPOSE: To determine whether extremely low frequency electromagnetic fields can alter average free cytosolic calcium ion concentrations [Ca2+]i and transient increases in [Ca2+]i in populations of ROS 17/2.8 cells. MATERIALS AND METHODS: Cells loaded with the calcium-selective luminescent photoprotein, aequorin, were placed in the bottom of a sample chamber, which was inserted into the gap of a previously described air gap reactor system where they were exposed either to sinusoidal magnetic fields at a variety of frequencies and flux densities or to sham conditions. Real-time recordings of photon counts due to aequorin luminescence were obtained and data were analysed with the use of probit plots. RESULTS: Probit plots of data obtained from cells exposed to the various magnetic fields were virtually superimposable over the data obtained for the same cultures during pre- and post-exposure sham or no-field periods. CONCLUSION: These experiments provided no evidence for any effects of ELF EMF, either positive or negative, on either average [Ca2+]i or on transient increases in [Ca2+]i.     

Iloprost在实验性颅脑损伤中的保护作用

目的：研究lloprost对大鼠创伤性脑损伤后神经记忆行为及突触体内[Ca^2 ]液度与脑含水量的影响。方法：液压冲击致伤装置制作大鼠脑创伤模型，爬杆回避箱测定动物的神经记忆行为，干湿重法测定脑组织含水量，Fura-2/AM荧光标记法测定突触体内[Ca^2 ]i，并采用Iloprost进行治疗，研究其对神经记忆行为、[Ca^2 ]i及脑水肿的影响。结果：创伤后动物会发生神经记忆行为的改变、脑组织水含量的增加及钙超载，Iloprost治疗后能改善动物的神经记忆行为，使[Ca^2 ]i明显下降，脑水肿明显减轻。结论：细胞内钙通道开放、钙离子超载在创伤性脑水肿的发生发展中起了重要作用，Iloprost对创伤性脑水肿有治疗和脑保护作用。     

大鼠脑组织γ刀照射后水通道蛋白4mRNA与蛋白激酶C活性变化的相关性研究

目的 探讨大鼠脑组织γ刀照射后水通道蛋白(AQP)4 mRNA的表达变化及其与蛋白激酶(PKC)活性变化的相关性。方法 Wistar大鼠30只,用γ刀照射尾状核头部(单靶点照射,中心剂量100 Gy,准直器为4 mm)制成放射外科模型,观察照射后1、3、7、15、30和45d 脑组织中AQP4 mRNA、PKC活性及细胞内游离Ca²⁺的变化和相互关系。检测方法分别为RT-PCR、液态闪烁计数以及Fura-2/AM法。结果 AQP4 mRNA的表达由正常对照的0.99±0.05逐渐增加至照射后30 d的2.32±0.10,45 d下降至2.21±0.08;PKC 活性及脑细胞内游离Ca²⁺浓度则分别由正常对照的0.5896±0.2101和455.82±20.13逐渐下降至30 d 的0.0404±0.0294和196.72±9.87,45d又回升至0.1050±0.0607和219.26±10.43。除1 d 外,AQP4 mRNA、PKC活性及细胞内游离Ca²⁺ 均与对照组差异有统计学意义(P<0.01)。AQP4 mRNA与PKC呈显著负相关(r=-0.918,P=0.003),脑细胞内游离Ca²⁺浓度与PKC 活性呈显著正相关(r=0.959,P=0.001)。结论 γ刀照射后PKC活性受抑可能是脑组织AQP4 mRNA表达上调的因素之一,而PKC活性降低则可能与高剂量电离辐射致细胞内Ca²⁺浓度降低有关。     

Intracellular calcium during excitation-contraction coupling in mammalian ventricle.

A rapid, transient, rise in cytoplasmic free calcium ion concentration ([Ca2+]i-transient) couples electrical excitation to contraction in muscle. Such [Ca2+]i-transients in muscle are actually subcellular spatio-temporal events that are determined dynamically by i) diffusional fluxes of Ca2+, ii) by the binding or unbinding of Ca2+ to ligands such as troponin c and calmodulin, and iii) by the various cellular processes, such as release of Ca2+ from sarcoplasmic reticulum, that produce fluxes of Ca2+ across the membranes bounding organelles or the cell. In heart muscle, a particularly large number of cellular processes contribute to the cytoplasmic [Ca2+]i-transient, compared with skeletal muscle. In addition, the actual change in cytoplasmic free [Ca2+]i is now known to be a small fraction of the total [Ca] transient (free plus bound) because most (98 to 99%) of the Ca2+ that enters the cytoplasm binds to ligands. In this article it is shown that under most physiological conditions the SR is the major determinant of the [Ca2+]i-transient in heart, that release of Ca2+ from the SR is induced by Ca2+ entering via L-type Ca(2+)-channels, and that the Na/Ca exchanger is the major route by which Ca2+ leaves the cell. The precise quantitative contribution of all these processes to the [Ca2+]i-transient still remains to be determined, however.     

Effect of in ovo immobilization on development of chick hind‐limb articular cartilage: An evaluation using micro‐MRI measurement of delayed gadolinium uptake

To examine the effect of immobilization on the development of articular cartilage, we assessed glycosaminoglycan (GAG) content in the chick articular surface by delayed gadolinium‐enhanced MRI of cartilage (dGEMRIC). Chick embryos were paralyzed by decamethonium bromide (DMB) from day 10 to either day 13 or day 16. The GAG content of the chick knee was compared with that of nonparalyzed chick embryos. Histologic analysis was unable to quantify GAG content; however, dGEMRIC demonstrated that GAG content was higher in the femoral condyles of the nonparalyzed embryos on day 13, and on day 16 the GAG content was lower in both the femoral condyles and the tibial plateaus of the nonparalyzed embryos. These results suggest that paralysis delays embryonic hind‐limb development. Osteoblastic activity at the cartilage canal, as demonstrated by staining for alkaline phosphatase (ALP), was present only in the nonparalyzed chick embryos on day 16. The GAG content of the cartilage decreased when the cartilage canals began to form on day 16. The effect of immobilization on hind‐limb development was indicated by the differences in the GAG content of the cartilage anlage measured by dGEMRIC in the developing knee joint of paralyzed and nonparalyzed embryonic chicks. Magn Reson Med, 2006. © 2006 Wiley‐Liss, Inc.     

Mitochondrial and intracellular free-calcium regulation of radiation-induced apoptosis in human leukemic cells.

PURPOSE: To investigate the mechanisms and pathways of X-ray apoptosis in Molt-4 cells, focusing on mitochondrial and cytosolic Ca2+ ([Ca2+]i) regulation. MATERIALS AND METHODS: X-irradiated Molt-4 cells and cell extract (CE) were used to analyse: (1) induced apoptosis (Giemsa stain), (2) p53, Bcl-2 and Bax expressions (immunoblot), (3) mitochondrial potential deltapsi(m) and (4) [Ca2+]i (flow cytometry), (5) caspase-3 activity, and (6) roles of [Ca2+]- and caspase-3-mediated pathways by inhibiting either or both pathways for induced apoptosis. RESULTS: Molt-4 cells were sensitive to apoptosis since 5 Gy induced 57 and 94% apoptosis at 6 and 24 h. After 5Gy, p53 was accumulated that upregulated Bax but which repressed Bcl-2 with time, resulting in a 7-fold increase in Bax/Bxl-2 at 6 h. Predominant Bax reduced deltapsi(m), and low-deltapsi(m) cells increased 45 min earlier than apoptosis after 5 Gy. Caspase-3 was activated in apoptotic CE. The caspase-3 inhibitor Ac-DEVD-CHO inhibited apoptosis and DNA-ladder formation by approximately 50%, suggesting a approximately 50% role of caspase-3-activated DNase (CAD). [Ca2+]i was increased after 5 Gy. [Ca2+]i-chelating BAPTA-AM (5 microM) and/or DNase gamma-inhibiting Zn2+ (0.5 mM) inhibited approximately 50% of induced apoptosis and DNA-laddering, indicating a 50% participation of Ca2+/Mg2+-dependent DNase gamma. CONCLUSIONS: The p53-Bax-mitochondria-caspase-3-CAD pathway and the [Ca+2]i-mediated DNase gamma pathway were involved in the regulation of X-ray apoptosis in sensitive Molt-4 cells.     

氧化低密度脂蛋白及辛伐他汀对平滑肌细胞信号通路蛋白激酶活性和胞浆内游离钙的影响

分别应用γ-^32P-ATP磷酸转移法和Fluo－3／Am荧光负载、流式细胞术检测氧化修饰LDL（oxLDL）及辛伐他汀对大鼠主动脉平滑肌细胞（AS MC）蛋白激酶C（PKC）和胞浆内游离钙（[Ca^2 ]i)水平的变化。结果表明,oxLDL呈剂量依赖方式促进ASMCPKC活性增加,14min时达峰值,然后缓慢下降,30min仍维持较主水平。胞浆内[(Ca^2 ]i升高分两个时相,即快速相和持续相。移去细胞外液钙,oxLDL仍引起快速相,但持续相消失,而辛伐他汀能明显抑制oxLDL引起的ASMCPKC活性增加,并显著降低持续相胞浆内钙水平,但对快速相无影响。提示oxLDL能引起ASMC内信号通路PKC及[(Ca^2 ]i的动态变化,二者密切相关。oxLDL刺激ASMC[Ca^2 ]升高的快速相是由胞浆钙池释放引起,持续相升高是由胞外钙内流引起。辛伐他汀抑制ASMCPKC活性可能是通过胞内[Ca^2 ]i变化起作用。     

Time course changes in [Ca2+]i, force, and protein content in hindlimb-suspended mouse soleus muscles

BACKGROUND: Exposure to reduced gravitational forces during spaceflight is associated with significant reductions in skeletal muscle mass and strength. The purpose of this study was to test the hypothesis that increases in resting cytosolic free calcium concentration ([Ca2+]i) would precede reductions in protein content and maximal isometric tetanic force (Po) in mouse soleus muscle after initiation of hindlimb suspension. METHODS: Female ICR mice (n = 42) were hindlimb suspended for 1, 2, 3, 5, or 7 d; weight-matched mice were used as controls. Following the hindlimb suspension, the left soleus muscle was used to determine Po in vitro and the right soleus muscle was used to determine protein content and [Ca2+]i via confocal laser scanning microscopy. RESULTS: Compared with controls, [Ca2+]i was elevated by 38% at 2 d, and 117% at 7 d. Compared with controls, soleus muscle total and myofibrillar protein contents were reduced 27-29% and 30-34%, respectively, at 5-7 d after initiation of hindlimb suspension. Compared with controls, soleus muscle Po was decreased by 24% at 3 d, and 38% at 7 d. CONCLUSION: It appears that resting cytosolic Ca2+ homeostasis is disturbed soon after the initiation of hindlimb suspension, and these elevations in [Ca2+]i may play a role in initiating soleus muscle atrophy.