白介素1β抑制培养的大鼠皮层神经元钠电流峰值和动作电位幅度

白介素1β(interleukin-1β,IL-1β)是重要的促炎细胞因子,在中枢神经系统的生理学和病理学过程中发挥关键作用。电压门控钠通道是可兴奋细胞电学活动的基础,控制神经元的兴奋性和动作电位。最近的研究又显示了IL-1β与电压门控通道之间的相互作用。为考察中枢神经元中IL-1β与电压门控钠通道之间的相互作用,本研究使用10ng/mL的IL-1β处理培养的大鼠皮层神经元24h,通过电压钳技术测定电压门控钠电流,结果表明IL-1β处理抑制钠电流幅度,但不改变其激活和失活性质。与电压钳记录结果相一致,电流钳记录表明IL-1β降低动作电位幅度但不影响阈值。这些结果显示长时间的IL-1β处理可以抑制电压门控钠电流,这种抑制作用减小了动作电位幅度,这可能改变神经元的电学性质、突触传导等基本功能,并提示了IL-1β在神经系统损伤和疾病中作用的新的思路。     

白介素1β对培养的大鼠皮层神经元钙通道电流的抑制作用

白细胞介素1β（IL-1β）是重要的促炎细胞因子,在中枢神经系统（CNS）中发挥广泛的生物学功能。在病理条件下,细胞膜上电压门控钙通道的变化与疾病发展过程密切相关。虽然IL-1β和钙通道都在脑损伤和脑疾病过程中发挥重要作用,但目前还很少有两者之间相互关系的研究报道。该研究使用了培养的大鼠胎鼠皮层神经元和膜片钳记录技术,研究了长时间的IL-1β处理对电压门控钙通道电流的作用。结果表明,IL-1β在10和50 ng/mL剂量下都可以抑制钙电流,这种抑制作用具有时间和剂量依赖性的模式,并且不改变钙通道的激活性质。     

白细胞介素-6对新生大鼠海马神经元电压依赖离子通道和NMDA电流的影响

目的: 研究白细胞介素-6对海马神经元电压依赖离子通道和NMDA电流的影响.方法: 应用全细胞膜片钳技术观察IL-6对电压依赖性钠通道电流(INa),延迟整流性钾通道电流(IK),电压依赖性钙通道电流(ICa),NMDA(N-methyl-D-aspartate)受体通道电流的影响.结果: 50 ng/ml IL-6作用24 h后IK 和ICa明显减小,Cm明显增大.50,500 ng/ml时减小NMDA电流.结论: IL-6通过作用于电压依赖钾通道,钙离子通道及NMDA通道影响神经元功能.     

二氧化硫衍生物对大鼠背根神经元瞬间外向钾电流和延迟整流钾电流的影响

运用全细胞膜片钳技术研究二氧化硫衍生物对大鼠背根神经元瞬间外向钾电流(IA和ID)和延迟整流钾电流(IK)的影响。结果发现二氧化硫衍生物剂量依赖性地增大钾通道的电导,电压依赖性地增大钾电流的幅度,且这种增大作用部分可逆。二氧化硫非常显著地使延迟整流钾电流的激活过程向超极化方向移动,使瞬间外向钾电流的失活过程向去极化方向移动。10μmol/L二氧化硫衍生物作用前后,延迟整流钾电流的半数激活电压分别是(20.3±2.1)mV和(15.0±1.5)mV;IA和ID的半数失活电压分别朝去极化方向移动了6mV和7.4mV。这些结果表明二氧化硫改变了钾通道的特性,改变了神经元的兴奋性。     

多不饱和脂肪酸对成年雪貂心肌钾通道的作用

本研究是在成年雪貂的心肌上研究多不饱和脂肪酸（PUFA）对电压门控钾通道的效应。我们观察到,n-3 PUFA能抑制短时性外向钾电流（Ito)和延迟整流钾电流（IK),而对内向整流钾电流（IK1）则没有明显影响。二十二碳六烯酸（DHA）对Ito和Ik能产生浓度依赖性的抑制作用,其IC50分别为7.5和20μmol/L,但不影响IK1。二十碳五烯酸（EPA）对这三种钾通道的作用与DHA相似。花生四烯酸（5或10μmol/L)先引起IK的抑制,然后引起IK,AA的激活;用环氧合酶抑制剂消炎痛可以阻断花生四烯酸激活IK,AA的作用。不具有抗心律失常作用的单不饱和脂肪酸和饱和脂肪酸都不明显影响这些钾通道的活性。上述实验结果证明,n-3 PUFA能抑制心肌细胞的Ito和IK,但和我们以前报道的PUFA对心肌钠电流和钙电流的作用相比,其对Ito和IK抑制作用的效能较低。n-3 PUFA的抗心律失常效应可能与它们抑制心肌钠、钙、钾通道的作用有关。     

大鼠发育过程中下丘脑神经元钾离子通道温度效应变化的研究

为了探讨出生后钾离子通道在下丘脑神经元热敏感分化过程中的作用,采用膜片钳技术研究出生一个月内SD大鼠急性分离神经元的温度效应,结果表明IK电流密度在出生后一个月内变化不大(P>0.05),而IA电流密度则呈现为升高趋势(P<0.05).同时升高温度,不同出生日期的钾通道NPo都有不同程度的升高,但相较P1d的神经元来说,温度对P18d的电压依赖性影响更大一些.同时温度对IK和IA的影响是不一样的,IA的Q10>2,所有这些显示IA通道在神经元温度敏感性的发育分化过程中起着重要的作用.     

大鼠海马CA1区锥体神经元I_A和I_K在出生后早期的变化

大脑快速发育期(brain growth spurt,BGS)是神经元生长、突触连接的关键时期;电压门控性K+通道是维持细胞兴奋性和神经元间信息传递的关键通道。本文旨在探究BGS期内大鼠海马CA1区锥体神经元电压门控性K+通道电流及其通道动力学特性的变化,以期找出大鼠海马CA1区锥体神经元电压门控性K+通道发育的关键期。采用全细胞膜片钳技术,研究出生后0~4周大鼠海马CA1区脑片上的锥体神经元全细胞电压门控性K+通道电流及其通道动力学特性。结果显示:在测试电压为+90mV下,以出生后0周为参照,出生后1~4周的瞬时外向K+通道电流(IA)的最大电流密度的增幅分别为(16.14±0.51)%、(81.73±10.71)%、(106.72±5.29)%、(134.58±8.81)%(n=10,P<0.05);延迟整流K+通道电流(IK)的最大电流密度增幅分别为(16.75±3.88)%、(134.01±2.85)%、(180.56±8.49)%、(194.5±8.53)%(n=10,P<0.05),显示K+通道电流密度于1~2周增幅最大;IA的激活曲线向左移,半数激活电压随周龄增加逐渐减小,分别为14.67±0.75、13.46±0.64、8.39±0.87、4.60±0.96、0.54±0.92(mV,n=10,P<0.05);IK的激活曲线向左移,半数激活电压随周龄增加逐渐减小,分别为8.94±0.85、6.65±0.89、0.47±1.15、1.80±0.89、8.56±1.08(mV,n=10,P<0.05)。IA的失活曲线向左移,0周龄与1周龄之间的半数失活电压没有显著性差异,而出生后1~4周随周龄增加半数失活电压逐渐减小(P<0.05),分别为45.68±1.26、46.81±0.78、48.64±0.81、51.96±1.02、58.31±1.35(mV,n=10)。以上结果表明,随着鼠龄的增加,IA和IK电流密度逐渐增加,电压门控性K+通道半数激活、失活电压降低,尤其是出生后1周至2周变化明显,上述变化与海马神经元的逐渐发育成熟及其功能的完善有关。     

SO2代谢衍生物对大鼠海马CA1区神经元瞬间外向钾电流和延迟整流钾电流的影响

本文利用全细胞膜片钳技术研究了SO2 代谢衍生物———NaHSO3 和Na2 SO3 (二者分子比为 1∶3)对大鼠海马CA1区神经元瞬间外向钾电流 (IA)和延迟整流钾电流 (IK)的影响。结果表明 ,SO2 代谢衍生物可显著增大IA 和IK,且呈剂量依赖性关系 ,使IA 和IK 增大 5 0 %的剂量分别为 2 6 19μmol/L和 14 5 0 μmol/L。此外还与电压呈依赖性关系 ,但不具有频率依赖性。结果还表明 ,10 μmol/LSO2 代谢衍生物不影响IA 的激活过程 ,而对IK 的激活过程有非常显著的影响 ,给药前后IK 的半数激活电压分别为 17 6 4± 7 31mV和 13 43± 2 0 0mV (n=10 ,P <0 0 1) ,但不改变其斜率因子。另外 ,10 μmol/LSO2 代谢衍生物还非常显著地影响IA 的失活过程 ,给药前后其半数失活电压分别为 - 6 5 93± 1 97mV和 - 5 9 2 2± 3 83mV (n =10 ,P <0 0 1) ,但不改变其斜率因子。由此推断 ,SO2 代谢衍生物增大大鼠海马CA1区神经元的IA 和IK,促进IK 的激活过程 ,并抑制IA 的失活过程 ,可导致胞内K 通过K 通道的外流增加 ,胞内K 浓度降低 ,造成中枢神经元功能紊乱 ,诱导神经细胞凋亡。这意味着SO2 代谢衍生物对中枢神经系统具有损伤作用 ,从而提示大气SO2 污染可能与一些中枢神经系统疾病的发生以及衰老有关 [动物学报 49(1) :73     

脱氧鬼臼毒素对美洲大蠊背侧不成对中间神经元电压

目的:探讨脱氧鬼臼毒素(DOP)对美洲大蠊背侧不成对中间神经元(DUM)电压依赖性钾电流IK的影响。方法:采用全细胞膜片钳技术研究脱氧鬼臼毒素对美洲大蠊背侧不成对中间神经元电压依赖性钾电流的电流幅度,电流-电压关系以及激活曲线的影响。结果:DOP能够抑制电压依赖性钾通道电流的幅度,而且此抑制作用具有浓度依赖性(5、10、20、40μmol/L)。DOP抑制IK的半数抑制浓度(IC50)值为18.064μmol/L。20μmol/L DOP能使IK的电流-电压关系曲线下移,并能使IK的激活曲线向去极化方向移动。结论:DOP对美洲大蠊背侧不成对中间神经元(DUM)电压依赖性钾电流具有抑制作用,这可能是其杀虫作用的机制之一。     

白细胞介素-6对新生大鼠海马神经元电压依赖离子通道和NMDA电流的影响

目的 :研究白细胞介素 6对海马神经元电压依赖离子通道和NMDA电流的影响。方法 :应用全细胞膜片钳技术观察IL 6对电压依赖性钠通道电流 (INa) ,延迟整流性钾通道电流 (IK) ,电压依赖性钙通道电流 (ICa) ,NMDA(N methyl D aspartate)受体通道电流的影响。结果 :5 0ng/mlIL 6作用 2 4h后IK和ICa明显减小 ,Cm明显增大。 5 0 ,5 0 0ng/ml时减小NMDA电流。结论 :IL 6通过作用于电压依赖钾通道 ,钙离子通道及NMDA通道影响神经元功能。     

Inhibition of NMDA-induced outward currents by interleukin-1beta in hippocampal neurons

There is increasing evidence that a functional interaction exists between interleukin-1β (IL-1β) and N-methyl-d-aspartate (NMDA) receptors. The present study attempted to elucidate the effect of IL-1β on the NMDA-induced outward currents in mechanically dissociated hippocampal neurons using a perforated patch recording technique. IL-1β (30-100 ng/ml) inhibited the mean amplitude of the NMDA-induced outward currents that were mediated by charybdotoxin (ChTX)-sensitive Ca²⁺-activated K⁺ (K_Ca) channels. IL-1β (100 ng/ml) also significantly increased the mean ratio of the NMDA-induced inward current amplitudes measured at the end to the beginning of a 20-s application of NMDA. In hippocampal neurons from acute slice preparations, IL-1β significantly inhibited ChTX-sensitive K_Ca currents induced by a depolarizing voltage-step. IL-1 receptor antagonist antagonized effects of IL-1β. These results strongly suggest that IL-1β increases the neuronal excitability by inhibition of ChTX-sensitive K_Ca channels activated by Ca²⁺ influx through both NMDA receptors and voltage-gated Ca²⁺ channels.     

Glycosylation is required for maintenance of functional voltage-activated channels in growing neocortical neurons of the rat.

Voltage-activated currents were studied in whole-cell patch-clamped rat neocortical neurons growing in culture and treated with tunicamycin (TU), an inhibitor of protein N-glycosylation. The size of the Na+ current decreased progressively in the presence of TU (1-2 microM). This decrease was faster in growing 5-14 day-old neurons (to ca. 40% of control after 24 hours of treatment) than in fully grown 20-40-day-old neurons (to ca. 40% of control after 68 hours of treatment). The fast transient K+ current (A-current) was abolished, and the delayed rectifier K+ current was markedly reduced by a 24 hour treatment with TU (1-2 microM) in growing neurons. In contrast, in fully grown neurons these currents were unaffected by the same TU treatment. The size of the Ca2+ current was significantly reduced following a 24 hour treatment with TU (1-2 microM) in neurons at early stages of differentiation, but remained stable in 20-40-day-old neurons. It is concluded that protein glycosylation, presumably of the channel proteins themselves, is important for the functional expression of voltage-activated channels in embryonic cortical neurons during the early stages of cell growth in culture; the channels become less dependent on glycosylation in mature neurons.     

Interleukin-1β inhibits voltage-gated sodium currents in a time- and dose-dependent manner in cortical neurons

Interleukin-1β (IL-1β) is a multifunctional proinflammatory cytokine that plays a key role in the injuries and diseases of the central nervous system (CNS). A voltage-gated Na⁺ channel is essential for the excitability and electrical properties of neurons. However, it is not known whether IL-1β directly affects the central Na⁺ channels. In the present study, we examined the effects of IL-1β on Na⁺ currents in cultured cortical neurons using patch-clamp recording. Our results showed that IL-1β suppressed Na⁺ currents through its receptor in a time- and dose-dependent manner, but did not alter the voltage-dependent activation and inactivation. PKC and then p38 MAPK were involved in this inhibition. The spike amplitude was also inhibited by IL-1β in the doses that decreased the Na⁺ currents. Our findings revealed the inhibition of chronic IL-1β treatment on voltage-gated Na⁺ channels in the CNS, and showed that the action potential (AP) amplitude was reduced by IL-1β due to a decrease of Na⁺ currents.     

Modulation of voltage-activated ion currents on identified neurons ofHelix pomatia L. by interleukin-1

1. The effect of interleukin-1 (IL-1) was studied on voltage-activated ion currents of the identified central neurons of Helix pomatia L. using a two-microelectrode voltage clamp. The voltage-activated inward current (ICa) was decreased, whereas the outward current (I(net) K) was increased by IL-1. 2. IL-1 affects both the transient and the delayed rectifying potassium currents. The IL-1 modulatory effect on the voltage-activated ion currents was voltage and dose dependent. The threshold concentration for IL-1 was 2 U/ml. 3. The proposed modulatory effect of IL-1 appears to have more than one site of action on the neuron membrane ion channels. 4. Rabbit anti-human IL-1 polyclonal antiserum eliminated the IL-1 effects on the voltage-activated inward and outward currents. This is the first report demonstrating a direct effect of IL-1 modulation of voltage-activated ion currents on neurons of mollusks.     

Interleukin-1beta downregulates the L-type Ca2+ channel activity by depressing the expression of channel protein in cortical neurons

Interleukin-1beta (IL-1beta), a proinflammatory cytokine, has been involved in various diseases of the central nervous system (CNS). Due to the diverse, "contradictory" effects of IL-1beta on neurons during insults to the brain, the mechanisms underlying these effects have not been elucidated. Calcium influx through the L-type Ca2+ channels (LCCs) is believed to play a critical role in the cascade of biochemical events leading to neuron death in these pathophysiological conditions. So far, the mechanism of the interaction of IL-1beta and LCCs in the initiation and progression of these diseases is unclear. In this study, we investigate systemically the effects of IL-1beta on the LCCs current, which are believed to be implicated in the cascade of biochemical events leading to neuron death in neuropathological conditions. Using patch clamp, we observe that IL-1beta treatment (10 ng/ml, 24 h) suppresses LCC currents by approximately 38%, which made up half of the whole-cell Ca2+ current determined by nifedipine. IL-1beta does not alter the characteristics of single LCC including current amplitude, open probability, and conductance, but decreases the number of the functioning channel by 40%. Moreover, immunoblot assay exhibits that IL-1beta reduces the expression of LCC proteins by 38 approximately 42% in both whole neuron and plasma membrane fraction, and demonstrates that IL-1beta downregulates the LCC activity via the reduction of LCC density. According to early research pretreatments longer than 12 h may play a crucial role in the neuroprotective effects of IL-1beta, our findings may establish an explanation for the protective effects of this interleukin on neurons in the late stage of injury, and could raise a new issue to clinical treatment for insults to brain.     

Estradiol feedback alters potassium currents and firing properties of gonadotropin-releasing hormone neurons

GnRH neurons are regulated by estradiol feedback through unknown mechanisms. Voltage-gated potassium channels determine the pattern of activity and response to synaptic inputs in many neurons. We used whole-cell patch-clamp to test whether estradiol feedback altered potassium currents in GnRH neurons. Adult mice were ovariectomized and some treated with estradiol implants to suppress reproductive neuroendocrine function; 1 wk later, brain slices were prepared for recording. Estradiol affected the amplitude, decay time, and the voltage dependence of both inactivation and activation of A-type potassium currents in these cells. Estradiol also altered a slowly inactivating current, I(K.) The estradiol-induced changes in I(A) contributed to marked changes in action potential properties. Estradiol increased excitability in GnRH neurons, decreasing both threshold and latency for action potential generation. To test whether estradiol altered phosphorylation of the channels or associated proteins, the broad-spectrum kinase inhibitor H7 was included in the recording pipette. H7 acutely reversed some but not all effects of estradiol on potassium currents. Estradiol did not affect I(A) or I(K) in paraventricular neurosecretory neurons, demonstrating a degree of specificity in these effects. Potassium channels are thus one target for estradiol regulation of GnRH neurons; this regulation involves changes in phosphorylation of potassium channel components.     

Regulation of TRPM7 Function by IL-6 through the JAK2-STAT3 Signaling Pathway

Aims

Previous studies have demonstrated that expression of the TRPM7 channel, which may induce delayed cell death by mediating calcium influx, is precisely regulated. However, functional regulation of TRPM7 channels by endogenous molecules has not been elucidated. The proinflammatory cytokine IL-6 contributes to regulation of Ca²⁺ influx in cerebral ischemia, but the role of IL-6 in regulating TRPM7 functioning is unknown. Thus, we here investigated the interaction between IL-6 and TRPM7 channels and the relevant mechanisms.

Materials and Methods

Using whole-cell patch-clamping, we first investigated the effect of IL-6 on TRPM7-like currents in primary cultured cortical neurons. Next, TRPM7-overexpressing HEK293 cells were used to confirm the effect of IL-6/sIL-6R on TRPM7. Finally, we used specific signaling pathway inhibitors to investigate the signaling pathways involved.

Results

IL-6 or IL-6/sIL-6R dose-dependently inhibited inward TRPM7 currents, in both primary cultured neurons and HEK293 cells overexpressing TRPM7. In intracellular Mg²⁺-free conditions, extracellular Ca²⁺ or the α-kinase domain of TRPM7 did not participate in this regulation. The inhibitory effect of IL-6 on TRPM7 could be blocked by specific inhibitors of the JAK2−STAT3 pathway, but not of the PI3K, ERK1/2, or PLC pathways.

Conclusions

IL-6 inhibits the inward TRPM7 current via the JAK2−STAT3 signaling pathway.     

HIV-1gp120 induces neuronal apoptosis through enhancement of 4-aminopyridine-senstive outward K+ currents

Human immunodeficiency virus type 1 (HIV-1)-associated dementia (HAD) usually occurs late in the course of HIV-1 infection and the mechanisms underlying HAD pathogenesis are not well understood. Accumulating evidence indicates that neuronal voltage-gated potassium (Kv) channels play an important role in memory processes and acquired neuronal channelopathies in HAD. To examine whether Kv channels are involved in HIV-1-associated neuronal injury, we studied the effects of HIV-1 glycoprotein 120 (gp120) on outward K+ currents in rat cortical neuronal cultures using whole-cell patch techniques. Exposure of cortical neurons to gp120 produced a dose-dependent enhancement of A-type transient outward K+ currents (IA). The gp120-induced increase of IA was attenuated by T140, a specific antagonist for chemokine receptor CXCR4, suggesting gp120 enhancement of neuronal IA via CXCR4. Pretreatment of neuronal cultures with a protein kinase C (PKC) inhibitor, GF109203X, inhibited the gp120-induced increase of IA. Biological significance of gp120 enhancement of IA was demonstrated by experimental results showing that gp120-induced neuronal apoptosis, as detected by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay and caspase-3 staining, was attenuated by either an IA blocker 4-aminopyridine or a specific CXCR4 antagonist T140. Taken together, these results suggest that gp120 may induce caspase-3 dependent neuronal apoptosis by enhancing IA via CXCR4-PKC signaling.     

Enhancement of (45)Ca(2+) influx and voltage-dependent Ca(2+) channel activity by beta-amyloid-(1-40) in rat cortical synaptosomes and cultured cortical neurons. Modulation by the proinflammatory cytokine interleukin-1beta

Beta-amyloid protein is thought to underlie the neurodegeneration associated with Alzheimer's disease by inducing Ca(2+)-dependent apoptosis. Elevated neuronal expression of the proinflammatory cytokine interleukin-1beta is an additional feature of neurodegeneration, and in this study we demonstrate that interleukin-1beta modulates the effects of beta-amyloid on Ca(2+) homeostasis in the rat cortex. beta-Amyloid-(1-40) (1 microM) caused a significant increase in (45)Ca(2+) influx into rat cortical synaptosomes via activation of L- and N-type voltage-dependent Ca(2+) channels and also increased the amplitude of N- and P-type Ca(2+) channel currents recorded from cultured cortical neurons. In contrast, interleukin-1beta (5 ng/ml) reduced the (45)Ca(2+) influx into cortical synaptosomes and inhibited Ca(2+) channel activity in cultured cortical neurons. Furthermore, the stimulatory effects of beta-amyloid protein on Ca(2+) influx were blocked following exposure to interleukin-1beta, suggesting that interleukin-1beta may govern neuronal responses to beta-amyloid by regulating Ca(2+) homeostasis.