美蓝与Microcystine—LR对NO激活阻力血管平滑肌KCa的影响

应用膜片箝技术的细胞贴附式与膜内向外式记录大鼠肠系膜动脉Ａ４－Ａ５分支阻力血管平滑肌（ＶＳＭ）钙激活钾通道（ＫＣａ）活动，发现外源性一氧化氮（ＮＯ）不仅在细胞贴附式下，而且在游离膜片内面向外式时均能激活ＫＣａ，在细胞贴附式，鸟苷酸环化酶抑制剂美蓝（ｍｅｔｈｙｌｅｎｅｂｌｕｅ，ＭＢ）未能阻断外源性ＮＯ激活ＫＣａ的效应。蛋白磷酸酶ＰＰＩＡ、ＰＰＩＩＡ的抑制剂ｍｉｃｒｏｃｙｓｔｉｎｅ－ＬＲ可激活ＫＣａ，并能与外源性ＮＯ的激活效应相加。提示ｃＧＭＰ／ＰＫＧ并非外源性ＮＯ激活ＫＣａ的唯一途径；抑制脱磷酸化而相对增强磷酸化也可以激活ＫＣａ活动，并能与ＮＯ激活ＫＣａ产生协同效应     

鲫鱼视网膜水平细胞的分离、形态及生理特性

单个视网膜细胞已成为视网膜研究的重要标本。本文对现有的酶解方法加以改进，成功地分离了鲫鱼视网膜各类种经元和胶质细胞，并依据其形态学特征鉴定了类型．进而，应用全细胞模式的膜片箝技术，对急性分离的水平细胞上电压门控和配体门控通道进行了初步研究。水平细胞电流－电压关系呈明显的内向整流特性，对胞外钾离子和铯离子作用的进一步考察提示存在内向整流钾通道（ｉｎｗａｒｄｒｅｃｔｉｆｉｅｒ），其激活程度及通道电导受胞外钾离子浓度的影响．此外，微摩尔级的谷氨酸在水平细胞上诱导内向电流，而较高浓度的谷氨酸引起受体明显的失敏（ｄｅｓｅｎｓｉｔｉｚａｔｉｏｎ）。丙种新近鉴定的谷氨酸受体调制物ｃｙｃｌｏｔｈｉａｚｉｄｅ和伴刀豆球蛋白对谷氨酸电流有调制作用，但其效应与在哺乳类中枢神经元上不尽相同，提示水平细胞上谷氨酸受体的亚基组成有一定的特点。     

高糖、肿瘤坏死因子促进血管内皮细胞表面整合素（α_vβ_3）表达

本文采用ＥＬＩＳＡ方法检测培养的人脐静脉内皮细胞（ＨＵＶＥＣ）表面玻璃连接蛋白受体（ＶｉｔｒｏｎｅｃｔｉｎｒｅｃｅｐｔｏｒＶｎＲ即整合素家庭的α＿ｖβ＿３）的表达改变；用（５１）Ｃｒ－标记血小板（（５１）Ｃｒ－ｐｌａｔｅｌｅｔｅ，（５１）Ｃｒ－ｐＬ）检测ＨＵｖＥｃ的粘附功能；Ｆｕｒａ－２／Ａｍ负载ＥＣ，测定ＨＵＶＥＣ胞内游离钙离子浓度（［Ｃａ（２＋）］），观察了高糖、肿瘤坏死因子－α（Ｔｕｍｏｒｎｅｃｒｏｓｉｓｆａｃｔｏｒ－α，ＴＮＦ－α）对ＥＣ粘附功能的影响。结果表明：①高糖（３０ｍｍｏｌ／Ｌ）可明显促进ＥＣ与ＰＬ的粘附（ＣＰＭ值：３６１±２ｌ．９３ＶＳ２ｌ９．６７±１６．２６，ｎ＝６Ｐ＜０．０１），抗β３亚单位单抗（β３ＭｃＡｂ）可部分阻断ＥＣ与ＰＬ粘附。ＴＮＦ－α（１０００μ／ｍ１）也有相同作用（ＣＰＭ值：４ｌ０．７±１７．６ＶＳ２１９．６７±１６．２６１ｎ＝６Ｐ＜０．０１），β３ＭｃＡｂ也部分阻断ＴＮＦ－α诱导的ＥＣ－ＰＬ间的粘附。②不同浓度高糖和ＴＮＦ－α不同时间可影响ＥＣ表面的α＿ｖβ＿３表达，并且在一定范围内有浓度和时间依赖性。③高糖和ＴＮＦ－α可明显增加ＥＣ的［Ｃａ（２＋）］ｉ，上述资料提示?     

川芎嗪对慢性缺氧豚鼠右室心肌细胞钾电流的影响

目的： 探讨慢性缺氧及川芎嗪长期应用对豚鼠右室心肌细胞钾电流的影响。方法：采用全细胞膜片钳技术分别记录并比较正常对照组、缺氧组、药物（ 川芎嗪） 治疗组豚鼠单个右室心肌细胞的膜电容、延迟整流钾电流峰值和电流－ 电压关系曲线及内向整流钾电流－ 电压关系曲线。结果：上述３ 组细胞膜电容分别为（１５４ ．９ ±１６ ．７） ｐＦ、（１７７ ．３ ±１４ ．３）ｐＦ、（１６０ ．５ ±１３ ．９）ｐＦ,其中缺氧组显著大于另两组（ Ｐ＜ ０－０１） ;延迟整流钾电流峰值分别为（１ ．１３ ±０ ．２９）ｎＡ、（０ ．７９ ±０ ．２１） ｎＡ、（１ ．０３ ±０ ．２３） ｎＡ,其中缺氧组显著低于另两组（ Ｐ＜ ０－０５） ;在－ ２０ ｍ Ｖ～＋ ６０ ｍ Ｖ,慢性缺氧组延迟整流钾电流密度较另两组显著下降（ Ｐ＜ ０－０５） 。３ 组之间的内向整流钾电流无显著差异（ Ｐ＞ ０－０５） 。结论： 慢性缺氧不改变豚鼠右室心肌细胞内向整流钾电流,但能使细胞膜电容增加,延迟整流钾电流幅度和电流密度下降;长期应用川芎嗪能在一定程度上减轻这些变化     

人类ether-a-go-go相关基因通道的动力学分析和分析机制

探讨在电生理记录的过程中，由人类ether—a—go-go相关基因（HERG）编码的通道应分析的通道动力学参数及机制。使用双电极电压钳技术，记录表达于爪蟾卵母细胞的HERG通道，编制不同的刺激脉冲分析各动力学参数。结果显示：（1）HERG通道在去极化脉冲下由于存在快失活而呈内向整流性。激活曲线可通过拟合去极化脉冲及随之的尾电流峰值而得到。激活的时间依赖性可通过拟合去极化不同时程与相应的尾电流峰值而得到。（2）陋RG的去激活的I—V（电流一电压）关系曲线仍呈明显的内向整流性，尾电流的衰减路径用双指数方程拟合可得到快和慢两个时间常数。（3）HFRG通道的失活呈电压依赖性，分别用两个不同的三脉冲程序，可得到通道的失活曲线以及去除失活后近乎线性的I—V关系。因此虽然HERG通道动力学复杂，仍可设计不同脉冲程序对其通道动力学问接进行分析，为HERG的分子位点作用研究提供基础。     

从HCV蛋白一级结构预测其C,E,NS5区的CD_4~ T细胞抗原位点

我们根据ＣＤ４＋Ｔ细胞识别抗原位点的物理化学和生物学特征，设计了一个具有查找两亲性螺旋状结构（ａｍｐｈｉｐａｔｈｉｃｈｅｌｉｘｓｔｒｕｃｔｕｒｅ）肽段功能的计算机程序。用该程序对ＨＣＶ－１型病毒Ｃ、Ｅ（Ｅ１、Ｅ２／ＮＳ１）、ＮＳ５蛋白一级结构进行分析，发现这些蛋白区存在ＣＤ４＋Ｔ细胞识别位点。此结果支持了ＣＤ４＋Ｔ细胞对ＨＣＶＣ，Ｅ，ＮＳ５区可发生增殖反应的结论。提示该程序可作为一种预测ＣＤ４＋Ｔ细胞识别ＨＣＶ抗原位点的方法。     

用热启动聚合酶链反应检测血清中乙型肝炎病毒DNA技术的建立和应用

建立了热启动聚合酶链反应（ＰＣＲ）检测乙型肝炎病毒ＤＮＡ（ＨＢＶＤＮＡ）的技术。ＰＣＲ所有反应成分被２次加样。先加Ａ液（含ｄＮＴＰｓ、一对引物和ＭｇＣｌ＿２）与一粒石蜡珠。７０℃加热后冷却至室温，使蜡珠先融化后凝固形成蜡盖封住Ａ液，然后在向其上加入Ｂ液（含耐热性ＤＮＡ聚合酶、ＨＢＶＤＮＡ模板和ＫＣｌ）并开始循环扩增。当反应管内第一次变性温度升至６０℃以上时，中隔蜡层融化，蜡上浮形成防蒸发屏障，Ａ、Ｂ两液则由于热分子运动而混匀，从而保证引物与靶基因在较高温度下严格退火以减少错导非特异性扩增和引物聚体形成。提高了ＰＣＲ的特异性和灵敏性，应用这种技术对７６例肝炎血清进行了检测，结果ＨＢＶＤＮＡ检出率为６８％，其中ＨＢｓＡｇ（＋）、ＨＢｅＡｇ（＋）、抗－ＨＢｃ（＋）血清检出率为１００％；ＨＢｓＡｇ（＋）、抗－ＨＢｅ（＋）、抗－ＨＢｃ（＋）血清检出率为７０％；ＨＢｓＡｇ（＋）、抗－ＨＢＣ（＋）血清检出率为８９％；抗ＨＢＣ（＋）血清检出率为对％；标记全阴性血清检出率为３３％。     

应用DNA疫苗制备猪温病毒单克隆抗体的研究

猪瘟病毒（ｃｌａｓｓｉｃａｌ ｓｗｉｎｅ ｆｅｖｅｒ ｖｉｒｕｓ, ＣＳＦＶ）是影响畜牧业最重要的病原体之一。单克隆抗体（ｍＡｂ）以其高度的特异性和敏感性而在ＣＳＦＶ的生物学特性研究与诊断中起着重要的作用。由于不易获得高质量的ＣＳＦＶ抗原,我国开展这项工作较晚,所建立的抗ＣＳＦＶ ｍＡｂ细胞株较少。这对于研究ＣＳＦＶ的生物学特性及流行病学调查来说,在数量上还很不够,仍需继续研制抗ＣＳＦＶ ｍＡｂ。 ＤＮＡ疫苗是９０年代出现的一种新型疫苗［１］。其特点是：基因免疫后,蛋白质抗原在宿主细胞内表达,其加工处…     

SDF1基因一个多态位点在中国人群中的分布

关于趋化因子受体与人免疫缺陷病毒（ｈｕｍａｎｉｍｍｕｎｏｄｅｆｉｃｉｅｎｃｙｖｉｒｕｓ，ＨＩＶ）的相关性研究于１９９５年起步，迅速成为当前ＨＩＶ研究热点。１９９６年，Ｏｂｅｒｌｉｎ等［１］和Ｂｌｅｕｌ等［２］发现了基质细胞衍生因子（ｓｔｒｏｍａｌｄｅｒｉｖｅｄｆａｃｔｏｒ１，ＳＤＦ１）作为趋化因子受体ＣＸＣＲ４的配体在ＣＤ＋淋巴细胞和ＨＩＶ１的融合中起作用，它是嗜淋巴细胞ＨＩＶ１株感染的体外潜在抑制因子，可阻断ＨＩＶ１侵入人体的通路。本实验的目的是观察ＳＤＦ１基因多态性在中国不同地区汉族…     

淋巴细胞跨膜信号传递分子机制的研究进展

淋巴细胞的活化伴随着ｐ５６＾ｌｃｋ和ｐ５９＾ｆｙｎ酪氨酸蛋白激酶（ＰＴＫｓ）的激活，后者是淋巴细胞接受刺激触发一系列生物化学事件的关键。ＰＴＫ抑制剂可以阻断淋巴细胞的活化，而异常升高的ＰＴＫ活性有可能致淋巴细胞亚性转化。另外还发现ＧＰＩ－锚蛋白介导的信号传递、淋巴细胞活化后Ｃａ＾２＋动员等均与ＰＴＫ活性密切相关。体内有调节ＰＴＫｓ活性的蛋白酪氨酸磷酸酶（ＰＴＰａｓｅｓ），ＰＴＫｓ和ＰＴＰａｓｅｓ作     

Postnatal maturation of the hyperpolarization-activated cation current, I(h), in trigeminal sensory neurons

Hyperpolarization-activated inward currents (I(h)) contribute to neuronal excitability in sensory neurons. Four subtypes of hyperpolarization-activated cyclic nucleotide-gated (HCN) channels generate I(h), with different activation kinetics and cAMP sensitivities. The aim of the present study was to examine the postnatal development of I(h) and HCN channel subunits in trigeminal ganglion (TG) neurons. I(h) was investigated in acutely dissociated TG neurons from rats aged between postnatal day (P)1 and P35 with whole cell patch-clamp electrophysiology. In voltage-clamp studies, I(h) was activated by a series of hyperpolarizing voltage steps from -40 mV to -120 mV in -10-mV increments. Tail currents from a common voltage step (-100 mV) were used to determine I(h) voltage dependence. I(h) activation was faster in older rats and occurred at more depolarized potentials; the half-maximal activation voltage (V(1/2)) changed from -89.4 mV (P1) to -81.6 mV (P35). In current-clamp studies, blocking I(h) with ZD7288 caused membrane hyperpolarization and increases in action potential half-duration at all postnatal ages examined. ZD7288 also reduced the action potential firing frequency in multiple-firing neurons. Western blot analysis of the TG detected immunoreactive bands corresponding to all HCN subtypes. HCN1 and HCN2 band density increased with postnatal age, whereas the low-intensity HCN3 and moderate-intensity HCN4 bands were not changed. This study suggests that functional I(h) are activated in rat trigeminal sensory neurons from P1 during postnatal development, have an increasing role with age, and modify neuronal excitability.     

Potassium channels of myenteric neurons in guinea-pig small intestine

Patch-clamp recording was used to study rectifying K+ currents in myenteric neurons in short-term culture. In conditions that suppressed Ca2+ -activated K+ current, three kinds of voltage-activated K+ currents were identified by their voltage range of activation, inactivation, kinetics and pharmacology. These were A-type current, delayed outwardly rectifying current (I(K),dr) and inwardly rectifying current (I(K),ir). I(K),ir consisted of an instantaneous component followed by a time-dependent current that rapidly increased at potentials negative to -80 mV. Time-constant of activation was voltage-dependent with an e-fold decrease for a 31-mV hyperpolarization amounting to a decrease from 800 to 145 ms between -80 and -100 mV. I(K),ir did not inactivate. I(K),ir was abolished in K+ -free solution. Increases in external K+ increased I(K),ir conductance in direct relation to the square root of external K+ concentration. Activation kinetics were accelerated and the activation range shifted to more positive K+ equilibrium potentials. I(K),ir was suppressed by external Cs+ and Ba2+ in a concentration-dependent manner. Ca2+ and Mg+ were less effective than Ba2+. I(K),ir was unaffected by tetraethylammonium ions. I(K),dr was activated at membrane potentials positive to - 30 mV with an e-fold decrease in time-constant of activation from 145 to 16 ms between -20 and 30 mV. It was half-activated at 5 mV and fully activated at 50 mV. Inactivation was indiscernible during 2.5 s test pulses. I(K),dr was suppressed in a concentration-, but not voltage-dependent manner by either tetraethylammonium or 4-aminopyridine and was insensitive to Cs+. The results suggest that I(K),ir may be important in maintaining the high resting membrane potentials found in afterhyperpolarization-type enteric neurons. They also suggest importance of I(K),ir channels in augmentation of the large hyperpolarizing after-potentials in afterhyperpolarization-type neurons and the hyperpolarization associated with inhibitory postsynaptic potentials. I(K),dr in afterhyperpolarization-type enteric neurons has overall kinetics and voltage behaviour like delayed rectifier currents in other excitable cells where the currents can also be distinguished from A-type and Ca2+ -activated K+ current.     

Ca2+-channel current and its modification by the dihydropyridine agonist BAY k 8644 in isolated smooth muscle cells

The electrophysiological properties of single smooth muscle cells isolated from the longitudinal layer of the guinea-pig ileum were studied with the whole-cell patch-clamp technique. The finding of resting potentials between -45 and -50 mV and the occurrence of spontaneous electrical activity when K+ was the predominant intracellular cation indicated that the cells were not leaky or hyperpermeable. The existence of an inward Ca2+ current overlapping in time with an outward rectifying K+ current was demonstrated. The latter could be selectively blocked by replacing internal K+ with Cs+ and external Ca2+ with Ba2+. Depolarizations to potentials between -40 and +50 mV evoked time-dependent inward currents, with a maximum peak value between -20 and 0 mV. For depolarizations beyond +50 mV time-dependent outward currents appeared. These currents were inhibited by 0.1 mM CdCl2. The activation of the inward current showed a sigmoidal time course, and the rate of onset of the current increased at more positive potentials. Inactivation could be described by two exponentials. The threshold for activation was about -40 mV, and full activation was reached at 0 mV. Inactivation was complete near 0 mV, whereas the channels were fully available at -80 mV. The fully-activated Ca2+-channel current was strongly voltage dependent. The conductance decreased for potentials close to the reversal potential, and showed rectification for hyperpolarizing potentials. The Ca2+ agonist BAY k 8644 enhanced the Ca2+-channel current without a significant effect on its kinetics. The fully-activated current and the steady-state activation were enhanced in a rather voltage-independent way.     

Transient and delayed potassium currents in the Retzius cell of the leech, Macrobdella decora

The properties of a quickly inactivating transient K current (IA) and a slowly inactivating delayed K current (IK) were investigated with two-electrode voltage-clamp techniques in the isolated soma of the Retzius cell of the leech, Macrobdella decora. The two currents could be pharmacologically separated according to their different sensitivities to tetraethylammonium ions (TEA) and 4-aminopyridine (4-AP). IA was totally blocked by 3 mM 4-AP but not affected by 25 mM TEA. IK was suppressed almost completely by 25 mM TEA, whereas its peak amplitude only decreased by 10-15% in 3 mM 4-AP. IA was activated at membrane potentials more positive than -35 to -30 mV, whereas the threshold for IK was at more positive potentials of approximately -20 to -15 mV. The activation of IA was rapid with a voltage-dependent time constant [tau m(A)] that varied from 6 to 2 ms for command potentials between -20 and 10 mV (at 22-24 degrees C). The inactivation, which was independent of voltage, was somewhat slower with a time constant (tau A) of approximately 90-110 ms. The time constants for activation [tau m(K)] and the early inactivation phase (tau K) of IK were both voltage dependent. In the range of potential steps from 0 to 30 mV, tau m(K) varied from 12 to 4.5 ms and tau K from 1,500 to 700 ms. The steady-state inactivation of IA varied with holding potential and was complete at potentials more positive than -30 mV. IA was fully available from potentials more negative than -70 mV. IK did not show steady-state inactivation below its threshold of activation. The time course of IA during a maintained depolarization could be reasonably described by the expression IA(t) = IA(infinity) [1-exp(-t/tau m(A))]2 exp(-t/tau A). The time course of activation of IK without allowance for inactivation was approximated by the expression IK(t) = IK(infinity) [1-exp(-t/tau m(K))]2. The reversal potentials and magnitude of both IA and IK were dependent on extra-cellular K concentration, which suggest that a substantial part of the two currents was carried by K ions.     

Multiple T-type Ca2+ current subtypes in electrophysiologically characterized hamster dorsal horn neurons: possible role in spinal sensory integration

Whole cell patch-clamp recordings were used to investigate the contribution of transient, low-threshold calcium currents (I(T)) to firing properties of hamster spinal dorsal horn neurons. I(T) was widely, though not uniformly, expressed by cells in Rexed's laminae I-IV and correlated with the pattern of action potential discharge evoked under current-clamp conditions: I(T) in neurons responding to constant membrane depolarization with one or two action potentials was nearly threefold larger than I(T) in cells responding to the same activation with continuous firing. I(T) was evoked by depolarizing voltage ramps exceeding 46 mV/s and increased with ramp slope (240-2,400 mV/s). Bath application of 200 μM Ni(2+) depressed ramp-activated I(T). Phasic firing recorded in current clamp could only be activated by membrane depolarizations exceeding ～43-46 mV/s and was blocked by Ni(2+) and mibefradil, suggesting I(T) as an underlying mechanism. Two components of I(T), "fast" and "slow," were isolated based on a difference in time constant of inactivation (12 ms and 177 ms, respectively). The amplitude of the fast subtype depended on the slope of membrane depolarization and was twice as great in burst-firing cells than in cells having a tonic discharge. Post hoc single-cell RT-PCR analyses suggested that the fast component is associated with the Ca(V)3.1 channel subtype. I(T) may enhance responses of phasic-firing dorsal horn neurons to rapid membrane depolarizations and contribute to an ability to discriminate between afferent sensory inputs that encode high- and low-frequency stimulus information.     

GABAB agonists modulate a transient potassium current in cultured mammalian hippocampal neurons

Depolarization of voltage-clamped cultured rat hippocampal neurons from holding potentials more negative than -60 mV produced a transient outward current with the characteristics of an A-current: it was 50% inactivated at a holding potential of -85 mV and blocked by 4-aminopyridine (1 mM). In the presence of GABA or baclofen (50-200 microM), with or without bicuculline, inactivation of this current was shifted to more positive potentials so that there was little inactivation at -70 mV. Activation of the A-current was also shifted to more positive potentials by these agonists, but the voltage dependence of activation of the sodium current was unaffected. If A-currents with similar properties can influence the time course of action potentials in presynaptic terminals. GABAB agonists could make action potentials briefer by potentiating the A-current and hence depress transmitter release.     

Background nonselective cationic current and the resting membrane potential in rabbit aorta endothelial cells

The ion channel conductances that regulate the membrane potential was investigated by using a perforated patch-clamp technique in rabbit aorta endothelial cells (RAECs). The whole-cell current/voltage (I-V) relation showed a slight outward rectification under physiological ionic conditions. The resting membrane potential was -23.3 +/- 1.1 mV (mean +/- SEM, n = 19). The slope conductances at the potentials of -80 and 50 mV were 31.0 +/- 4.0 and 62.8 +/- 7.1 pS pF(-1), respectively (n = 15). Changes in the extracellular and intracellular Cl(-) concentrations did not affect the reversal potential on I-V curves. The background nonselective cationic (NSC) current was isolated after the K(+) current was suppressed. The relative permeabilities calculated from the changes in reversal potentials using the constant-field theory were P(K):P(Cs):P(Na):P(Li) = 1:0.87:0.40:0.27 and P(Cs):P(Ca) = 1:0.21. Increases in the external Ca(2+) decreased the background NSC current in a dose-dependent manner. The concentration for half block by Ca(2+) was 1.1 +/- 0.3 mM (n = 7). Through the continuous recording of the membrane potential in a current-clamp mode, it was found that the background NSC conductance is the major determinant of resting membrane potential. Taken together, it could be concluded that the background NSC channels function as the major determinant for the resting membrane potential and can be responsible for the background Ca(2+) entry pathway in freshly isolated RAECs.     

Local superfusion modifies the inward rectifying potassium conductance of isolated retinal horizontal cells

1. Horizontal cells were enzymatically and mechanically dissociated from the white perch (Roccus americana) retina and voltage clamped using patch electrodes. Steady-state current-voltage (I-V) relationships of solitary horizontal cells were determined by changing the membrane potential in a rampwise fashion. 2. The I-V curve of cells bathed in normal Ringer solution exhibited a large conductance increase at negative membrane potentials. This conductance activated near the K+ equilibrium potential, had no clear reversal potential, was enhanced by raising the extracellular concentration of K+, and was suppressed by external Cs+. These properties identify the conductance as the inward (anomalous) rectifier. 3. Continuous superfusion of the cells' local environment with drug-free Ringer reduced the magnitude of the inward rectifier current and shifted its activation point to more negative potentials. This effect developed over approximately 30 s, lasted as long as superfusion continued and was reversible upon cessation of superfusion. 4. Pressure ejection of drug-free Ringer solution onto cells bathed in the identical solution also reduced the magnitude of the inward rectifier current, although the effects were more rapid and more transient than those exerted by superfusion. Pressure ejection had little effect when cells were simultaneously superfused with Ringer, suggesting a common mode of action on the inward rectifier. 5. In the absence of superfusion, pressure ejection of Ringer containing 200 microM L-glutamate had a biphasic effect on membrane conductance. At potentials above -60 mV, glutamate caused a conductance increase with a reversal potential near +10 mV. At potentials below -60 mV, glutamate caused a conductance decrease whose reversal potential could not reliably be determined. The latter effect was similar to the suppression of the inward rectifier by application of Ringer alone, suggesting that it may represent an artifact of pressure ejection rather than a direct effect of glutamate. 6. In support of this interpretation, we found that pressure ejection of glutamate in the presence of external Cs+ (which blocks the inward rectifier) or during local superfusion with Ringer (which prevents attenuation of the inward rectifier by pressure ejection) did not cause a conductance decrease at negative potentials. Under these conditions, glutamate caused primarily a conductance increase with a reversal potential near +10 mV.(ABSTRACT TRUNCATED AT 400 WORDS)     

Modulation of transient and persistent inward currents by activation of protein kinase C in spinal ventral neurons of the neonatal rat

Neuronal excitability can be regulated through modulation of voltage threshold (Vth). Previous studies suggested that this modulation could be mediated by modulation of transient sodium currents (I(T)) and/or persistent inward current (PIC). Modulation of I(T) and PIC through activation of protein kinase C (PKC) has previously been described as a mechanism controlling neuronal excitability. We investigated modulation of I(T) and PIC by PKC in neonatal rat spinal ventral neurons. In whole cell voltage clamp, activation of PKC by application of 1-oleoyl-2-acetyl-sn-glycerol (OAG, 10-30 microM) resulted in 1) a reduction of I(T) amplitude by 33% accompanied an increase in half-width and a decrease in the maximal rise and decay rates of the I(T); 2) a reduction of PIC amplitude by 49%, with a depolarization of PIC onset by 4.5 mV. Activation of PKC caused varied effects on Vth for eliciting I(T), with an unchanged Vth or depolarized Vth being the most common effects. In current-clamp recordings, PKC activation produced a small but significant depolarization (2.0 mV) of Vth for action potential generation with an increase in half-width and a decrease in amplitude and the maximal rise and decay rates of action potentials. Inclusion of PKCI19-36 (10-30 microM), a PKC inhibitor, in the recording pipette could block the OAG effects on I(T) and PIC. The ability of serotonin to hyperpolarize Vth was not altered by PKC activation or inhibition. This study demonstrates that activation of PKC decreases the excitability of spinal ventral neurons and that Vth can be modulated by multiple mechanisms.     

A voltage-dependent transient K(+) current in rat dental pulp cells

We characterized a voltage-dependent transient K(+) current in dental pulp fibroblasts on dental pulp slice preparations by using a nystatin perforated-patch recording configuration. The mean resting membrane potential of dental pulp fibroblasts was -53 mV. Depolarizing voltage steps to +60 mV from a holding potential of -80 mV evoked transient outward currents that are activated rapidly and subsequently inactivated during pulses. The activation threshold of the transient outward current was -40 mV. The reversal potential of the current closely followed the K(+) equilibrium potential, indicating that the current was selective for K(+). The steady-state inactivation of the peak outward K(+) currents described by a Boltzmann function with half-inactivation occurred at -47 mV. The K(+) current exhibited rapid activation, and the time to peak amplitude of the current was dependent on the membrane potentials. The inactivation process of the current was well fitted with a single exponential function, and the current exhibited slow inactivating kinetics (the time constants of decay ranged from 353 ms at -20 mV to 217 ms at +60 mV). The K(+) current was sensitive to intracellular Cs(+) and to extracellular 4-aminopyridine in a concentration-dependent manner, but it was not sensitive to tetraethylammonium, mast cell degranulating peptide, and dendrotoxin-I. The blood depressing substance-I failed to block the K(+) current. These results indicated that dental pulp fibroblasts expressed a slow-inactivating transient K(+) current.