布比卡因对大鼠背根神经节钠电流的影响

目的 观察布比卡因对大鼠背根神经节神经元钠电流的影响，探讨椎管内麻醉的作用机制。方法 以全细胞膜片钳技术记录临床浓度的布比卡因0～1000μmol/L对急性分离大鼠背根神经元TTX-S和TTX-R钠电流的影响。钳制电压分别为-100mv和-90mv，刺激电压分别为-10mv和0mv。结果 临床浓度的布比卡因对TTX-S和TTX-R钠电流均有明显的抑制作用，布比卡因的浓度与其抑制强度呈正相关；布比卡因对TTX-S钠电流抑制的半数有效量IC50为17．069μmol/L(Hill系数为0．8988)，而TTX-R钠电流的半数有效量IC50为34．511μmol/L(Hill系数为0．9003)。结论 临床浓度的布比卡因对背根神经节神经元TTX-S和TTX-R钠电流具有抑制作用，对TTX-S钠电流的阻滞比对TTX-R钠电流的阻滞强1倍左右，其抑制程度随局麻药浓度的增加而增加。     

The effect of dibutyryl cAMP on tetrodotoxin-sensitive and -resistant voltage-gated sodium currents in rat dorsal root ganglion neurons and the consequences for their sensitivity to lidocaine

Tetrodotoxin-sensitive (TTXS) sodium currents in dorsal root ganglia (DRG) neurons were enhanced by DcAMP applied acutely or by pre-treatment. Pre-treatment increased peak TTXS by 28%. This compared to the increase of tetrodotoxin-resistant sodium currents (TTXR) of 123%. In both cases the increase was associated with a hyperpolarizing shift in activation potentials. Slow inactivation was slower for both TTXR and TTXS in DcAMP treated neurons but rates of recovery from inactivation were not altered. Lidocaine blocked TTX-R with an IC(50) of 0.51+/-0.15mM (n=9) which was reduced to 0.14+/-0.05mM (n=8, P<0.05) in DcAMP treated cells. The sensitivity of TTX-S currents to lidocaine was not altered by DcAMP (control EC(50)=0.89+/-0.16mM, n=9; DcAMP EC(50)=0.73+/-0.19mM, n=6). It is concluded that TTXS currents in DRG are, like TTX-R currents, enhanced by cAMP but whilst the pharmacology of TTXR channels with respect to lidocaine is altered, that to TTXS channels is not.     

龙血素B抑制大鼠背根神经节细胞辣椒素诱发的电流反应

目的探讨龙血素B对大鼠背根神经节细胞辣椒素诱发的辣椒素受体电流的影响。方法采用全细胞膜片钳技术在急性分离的大鼠背根神经节细胞上观察龙血素B对辣椒素诱发的辣椒素受体电流的影响。结果①在-60mV的钳制电位下,辣椒素受体拮抗剂辣椒卓平可以完全抑制辣椒素受体电流;②不同浓度的龙血素B溶液对辣椒素诱发的受体电流具有浓度依赖的抑制作用;2.0、4.0、8.0和16.0μmol.L-1的龙血素B溶液对辣椒素诱发的受体电流峰值的抑制率分别为15.36%±2.12%、36.41%±2.43%、76.26%±2.16%和96.69%±3.21%(n=10,P<0.05),半数抑制浓度(IC50)为4.9μmol.L-1,Hill系数为2.29。结论龙血素B可以明显抑制辣椒素诱发的辣椒素受体电流,影响痛觉信息的传入,这可能是以龙血素B为重要成分的中药血竭产生镇痛作用的机制之一。     

神经病理性疼痛小鼠背根神经节河豚毒素敏感型钠电流的变化特征

目的 在小鼠坐骨神经慢性压迫性损伤(CCI)模型上,观察背根神经节(DRG)细胞河豚毒素敏感型(TTX-S)钠电流的变化特征。方法 采用全细胞膜片钳技术分别记录CCI模型及正常小鼠DRG细胞TTX-S钠电流,比较其电流变化特征。结果 CCI 模型小鼠热疼痛阈值较正常小鼠下降约50%。在CCI小鼠DRG细胞上,TTX-S钠电流较正常组明显增加,最大激活电压由-10 mV左移为-20 mV,最大激活电流也由正常组的-(96.3±7.7)pA/pF增加至-(146.6±12.7)pA/pF,其稳态激活和失活曲线分别向较负和较正电压方向偏移14.9和5.1 mV,但通道失活后恢复动力学特征不变。结论 CCI小鼠增加TTX-S钠电流。     

和厚朴酚对小鼠背根神经节细胞河豚毒素不敏感钠电流的影响

目的 通过观察和厚朴酚（honokiol, Hon）对河豚毒素不敏感（TTX-R）钠电流的作用,探讨它可能的镇痛机制。方法 应用酶解法急性分离小鼠背根神经节细胞,全细胞膜片钳技术记录河豚毒素不敏感钠电流。结果 和厚朴酚对河豚毒素不敏感钠电流的抑制呈现浓度依赖性,半抑制浓度（IC₅₀）为28.1 μmol·L^-1。和厚朴酚（30 μmol·L^-1）使河豚毒素不敏感钠电流密度下降48.1%,稳态激活和失活曲线分别向右和左偏移约7 mV和11.1 mV,但不影响通道的恢复时间。结论 和厚朴酚明显抑制河豚毒素不敏感钠电流,其作用可能与它的镇痛机制有关。     

Modulation by oxytocin of ATP-activated currents in rat dorsal root ganglion neurons

The modulatory effect of oxytocin (OT) on ATP-activated currents (I(ATP)) was studied in freshly isolated dorsal root ganglion (DRG) neurons of rats using whole cell clamp technique. In most of the neurons examined (50/70, 71.4%) extracellular application of OT (10(-9)-10(-5) mol/L) suppressed I(ATP) while in the rest (20/70, 28.6%) no modulatory effect was observed. OT shifted the ATP concentration-response curve downwards with a decrease of 39.8+/-4.2% in the maximal current response and with no significant change of Kd value. This OT-induced inhibition of I(ATP) showed no voltage dependence, and could be blocked by [d(CH(2))(5),Tyr(Me)(2),Thr(4),Tyr-NH(2)(9)]-OVT (d(CH(2))(5)-OVT) (10(-8) mol/L), a specific OT receptor antagonist. Intracellular application of H-9 (4 x 10(-5) mol/L, an inhibitor of protein kinase A) (n=12), BAPTA (10(-2) mol/L, a chelator of calcium ions) (n=4) could reverse the inhibitory effect of extracellular OT (10(-7) mol), while inclusion of H-7 (2 x 10(-5) mol/L, a protein kinase C inhibitior) (n=8) and KN-93 (10(-5) mol/L, an inhibitor of CaMKII) (n=9) in the recording pipette did not affect this effect. The results suggested that OT inhibition on ATP-activated currents was mediated by OT receptors in the membrane of DRG neurons; and this inhibitory effect involved the transduction of intracellular cAMP-PKA and Ca(2+).     

CGP37157 modulates mitochondrial Ca homeostasis in cultured rat dorsal root ganglion neurons

The effects of 7-chloro-3,5-dihydro-5-phenyl-1H-4,1-benzothiazepine-2-on (CGP37157), an inhibitor of mitochondrial Na⁺/Ca²⁺ exchange, on depolarization-induced intracellular free Ca²⁺ concentration ([Ca²⁺]_i) transients were studied in cultured rat dorsal root ganglion neurons with indo-1-based microfluorimetry. A characteristic plateau in the recovery phase of the [Ca²⁺]_i transient resulted from mitochondrion-mediated [Ca²⁺]_i buffering. It was blocked by metabolic poisons and was not dependent on extracellular Ca²⁺. CGP37157 produced a concentration-dependent decrease in the amplitude of the mitochondrion-mediated plateau phase (IC₅₀=4±1 μM). This decrease in [Ca²⁺]_i was followed by an increase in [Ca²⁺]_i upon removal of the drug, suggesting that Ca²⁺ trapped in the matrix was released when the CGP37157 was removed from the bath. CGP37157 also inhibited depolarization-induced Ca²⁺ influx at the concentrations required to see effects on [Ca²⁺]_i buffering. Thus, CGP37157 inhibits mitochondrial Na⁺/Ca²⁺ exchange and directly inhibits voltage-gated Ca²⁺ channels, suggesting caution in its use to study [Ca²⁺]_i regulation in intact cells.     

葛根素抑制大鼠背根神经节细胞河豚毒素不敏感性钠电流(英文)

目的:测定葛根素(Pue)对大鼠背根神经节(DRG)细胞河豚毒素不敏感性(TTXr)钠电流的作用。方法:采用全细胞钳制技术,记录成年Wistar大鼠DRG神经元中TTXr钠电流。结果:Pue在0.01-2mmol·L~(-1)浓度范围内,对TTXr钠电流的抑制率为9.5％-83.2％。该抑制作用为浓度依赖性,可部分洗脱,但非频率依赖性或电压依赖性。Pue不影响失活曲线,但使1/2最大激活电压由-26mV升至-16mV,说明抑制了激活过程。结论:Pue抑制大鼠DRG细胞中TTXr钠电流。     

Prostaglandin E2 has no effect on two components of tetrodotoxin-resistant Na+ current in mouse dorsal root ganglion

One possible mechanism underlying inflammation-induced sensitization of the primary afferent neuron is the upregulation of tetrodotoxin-resistant (TTX-R) Na(+) current by inflammatory mediators such as prostaglandins. This notion is based on reports that showed an augmentation of TTX-R Na(+) current following an application of prostaglandin E(2) (PGE(2)) in dorsal root ganglion (DRG) neurons. However, no information was available on the properties of the novel type of TTX-R Na(+) channel, Na(V)1.9, at times when these reports were published. Hence, the contribution of Na(V)1.9 to the PGE(2)-induced upregulation of TTX-R Na(+) current remains to be elucidated. To further examine the modulation of TTX-R Na(+) current by PGE(2), we recorded two components of TTX-R Na(+) current in isolation from small (<25 microm in diameter) DRG neurons using wild-type and Na(V)1.8 knock-out mice. Unexpectedly, neither the component mediated by Na(V)1.8 nor the persistent component mediated by Na(V)1.9 was affected by PGE(2) (1 and 10 microM). Our results raise a question regarding the well-known modulatory role of PGE(2) on TTX-R Na(+) current in inflammatory hyperalgesia.     

苯佐卡因对大鼠背根神经节神经元河豚毒素不敏感型钠离子通道的影响

目的研究苯佐卡因(BZC)对大鼠背根神经节(DRG)神经元河豚毒素不敏感型(TTX-r)钠电流的影响,探讨其镇痛作用的机制。方法酶解法分离新生大鼠单个DRG神经元,应用全细胞膜片钳技术记录不同浓度BZC对TTX-r钠电流的影响。结果 BZC浓度依赖性静息阻断TTX-r钠电流,30、100和300μmol.L-1的BZC分别使TTX-r钠电流峰值抑制率达(18.83±8.51)%、(33.08±9.19)%、(58.91±12.02)%,并使TTX-r钠电流稳态失活曲线浓度依赖性向超极化方向移动。结论 BZC浓度依赖性阻断DRG神经元TTX-r钠离子通道并改变通道的失活,可能是其影响痛觉传导通路以及产生镇痛作用的机制之一。     

Effects of flunarizine on induced calcium transients as measured in fura-2-loaded neurons of the rat dorsal root ganglion

Summary The effect of the calcium entry blocker flunarizine on a high-potassium induced increase of intracellular free calcium was studied. The experiments were done with neurons isolated from rat dorsal root ganglia and loaded with the calcium-sensitive dye fura-2. The increase of calcium induced by 60 mmol/1 potassium was abolished after removal of extracellular calcium, was reversibly reduced by 50 mol/l cadmium (76% inhibition), 50 mol/1 nickel (25% inhibition) and 10 mol/1 nifedipine (18°10 inhibition), and reversibly increased after removal of extracellular sodium (26% increase). The potassium induced increase of intracellular calcium is, therefore, mediated by transmembrane calcium influx, probably to a large extent through cadmium-sensitive calcium channels. Flunarizine (5 min incubation followed 1 min wash-out) reduced the amplitude of the high-potassium induced calcium increase in a dose-dependent manner (K _d = 370 ± 100 nmol/l; mean ± SEM; n = 8), causing complete inhibition at a concentration of 10 mol/1 in the majority of cells. Flunarizine ( 1 mol/1) caused a reversible increase of the resting level of intracellular calcium in some cells, an effect which disappeared in the absence of extracellular calcium. The drug (1 mol/1 had no influence on the time course of recovery of intracellular calcium subsequent to a rise induced by high-potassium or by the calcium ionophore A23187. It is concluded that flunarizine acts as an inhibitor of depolarization-mediated calcium influx. At a concentration of 1 mol/1, the drug presumably has no effect on cellular calcium extrusion and/or sequestration mechanisms. Correspondence to L. Leybaert at the above address     

芍药苷对小鼠背根神经节TTX-S钠电流的影响

目的 本文通过观察芍药苷（Paeoniflorin,Pae）对小鼠背根神经节（DRG）细胞电压门控河豚毒素敏感型（TTX-S）钠电流的影响,探讨芍药苷的镇痛机制。方法 应用全细胞膜片钳技术在急性分离背根神经节细胞上记录河豚毒素敏感型钠电流。结果 芍药苷浓度依赖地抑制河豚毒素敏感型钠电流,其半抑制浓度（IC₅₀）为1 224.1 mmol·L^-1。芍药苷1 mmol·L^-1使河豚毒素敏感型钠通道的失活曲线向超极化方向转移约7.1 mV,并且延迟失活后通道的恢复,但对激活曲线没有影响。结论 芍药苷可能通过抑制河豚毒素敏感型钠通道,改变钠通道的动力学特征,从而发挥其镇痛作用。     

神经营养因子4在成年大鼠背根节神经元的分布

目的 探讨神经营养因子4（NT－4）与成年大鼠背根节神经元的关系。方法 本文用特异的NT－4抗血清以免疫组织化学方法观察了NT－4在成年大鼠背根L6节段神经元中的颁由。结果 NT－4的免疫阳性反应物分布背根节的各型神经元,胞浆染色。结论 NT－4可能与成年大鼠背根节神经元的生理功能有关。     

Inhibition of acid‐sensing ion channel currents by propofol in rat dorsal root ganglion neurons

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NO供体硝普纳损伤脊髓背根神经元模型的建立

目的在乳鼠背根神经节神经元原代纯化培养方法的基础上,采用硝普钠(SNP)制备背根神经节神经元损伤模型。方法取乳鼠背根神经节,加入5-氟-2'-脱氧尿苷(5-FUDR)以纯化细胞,MEM-F12培养基培养,用神经元特异性烯醇化酶(NSE)免疫组织化学染色鉴定神经元。神经元培养至第八天分别加入不同浓度SNP(500,50,5μmol·L-1)制备背根神经节神经元损伤模型,用台盼蓝染色进行细胞计数,MTT法测定培养神经元活力。结果培养的脊髓背根神经节神经元生长状态正常,纯化培养纯度高于90%,可存活两周以上。培养的背根神经节神经元加入SNP作用后,随着浓度增加,神经元死亡率逐渐升高。结论采用SNP制备的脊髓背根神经节神经元损伤模型稳定、可靠,可以作为相关实验研究的理想的体外实验模型。     

Effect of 8-bromo-cAMP on the tetrodotoxin-resistant sodium (Nav 1.8) current in small-diameter nodose ganglion neurons

We examined whether 8-bromo-cAMP (8-Br-cAMP)-induced modification of tetrodotoxin-resistant (TTX-R) sodium current in neonatal rat nodose ganglion neurons is mediated by the activation of protein kinase A (PKA) and/or protein kinase C (PKC). In 8-Br-cAMP applications ranging from 0.001 to 1.0mM, 8-Br-cAMP at 0.1mM showed a maximal increase in the peak TTX-R Na(+) (Nav1.8) current and produced a hyperpolarizing shift in the conductance-voltage (G-V) curve. The PKC inhibitor bisindolylmaleimide Ro-31-8425 (Ro-31-8425, 0.5microM) decreased the peak Nav 1.8 current. The Ro-31-8425-induced modulation of the G(V)(1/2) baseline (a percent change in G at baseline V1/2) was not affected by additional 8-Br-cAMP application (0.1mM). The maximal increase in Nav 1.8 currents was seen at 0.1microM after the application of a PKC activator, phorbol 12-myristate 13-acetate (PMA) and forskolin. The PMA-induced increase in Nav 1.8 currents was not significantly affected by additional 0.1mM 8-Br-cAMP application. Intracellular application of a PKA inhibitor, protein kinase inhibitor (PKI, 0.01mM), inhibited the baseline Nav 1.8 current, significantly attenuated the 8-Br-cAMP-and PMA-induced increase in the peak Nav 1.8 current, and caused a significant increase in the slope factor of the inactivation curve. The PKI application at a higher concentration (0.5mM) greatly inhibited the PMA (0.1microM)-induced increase in the peak Nav 1.8 current amplitude and further enhanced the Ro-31-8425-induced decrease in the current. These results suggest that the 8-Br-cAMP-induced increase in Nav 1.8 currents may be mediated by activation of both PKA and PKC.     

Phosphorylation of TRPV1 by neurokinin-1 receptor agonist exaggerates the capsaicin-mediated substance P release from cultured rat dorsal root ganglion neurons

The present study was conducted to determine whether the activation of neurokinin-1 receptor (NK-1R) by its agonist (GR73632) enhances the capsaicin-evoked substance P (SP) release using a radioimmunoassay. A pre-exposure to GR73632 enhanced the capsaicin-evoked SP release in a time- and dose-dependent manner. The augmentation of capsaicin-evoked SP release by GR73632 was completely inhibited by pharmacological blockade of NK-1R or transient receptor potential vanilloid receptor subtype 1 (TRPV1), and was partially attenuated by the inhibition of either protein kinase C (PKC), cyclooxygenase (COX) or phospholipase C (PLC), p38 or p42/44 mitogen-activated protein (MAP) kinase, but not protein kinase A. This augmentation of SP release was further increased by inhibition of c-Jun NH2-terminal kinase. A short-term (10min) exposure to GR73632 resulted in an increase in the TRPV1 phosphorylation. The increase in the TRPV1 phosphorylated forms induced by a 60-min exposure to GR73632 was completely abolished by the inhibition of either PKC, COX or PLC, p38 or p42/44 MAP kinases. Immunocytochemistry study demonstrated that the NK-1R and TRPV1 were mainly co-expressed in the small-sized neurons. These findings suggest that the activation of NK-1R by its agonist, by sensitizing the TRPV1 through the PKC phosphorylation of TRPV1, may play a role in the enhancement of the capsaicin-evoked SP release from cultured rat DRG neurons.     

Lipidosis-like alterations in dorsal root ganglion cells of rats treated with tricyclic antidepressants

Summary Prolonged treatment of rats with high doses of the tricyclic antidepressants iprindole, clomipramine, or 1-chlor-amitriptyline induced the formation of lamellated and crystalloid cytoplasmic inclusion bodies within dorsal root ganglion cells. The ultrastructural observations, which are compatible with the concept of a drug-induced generalized phospholipidosis, indicate that under experimental conditions also nerve cells can be affected by this type of drug side effect.     

地西泮抑制交感神经节细胞钠通道电流的机制

目的　研究地西泮对交感神经节细胞钠通道电流的抑制作用机制。方法　酶消化法急性分离SD大鼠(7～10　d）颈上交感神经节细胞，全细胞膜片钳技术记录地西泮对钠通道电流的影响。结果　在钳制电压(Vh)－80　mV,　刺激电压(Vt)0　mV条件下，0.3　μmol