Baldelli P, Forni P E, Carbone E
INFM Research Unit and Department of Neuroscience, I-University of Turin, Corso Raffaello 30, I-10125 Turin, Italy.
Eur J Neurosci. 2000 Nov;12(11):4017-32. doi: 10.1046/j.1460-9568.2000.00305.x.
Neurotrophins exert short- and long-term effects on synaptic transmission. The mechanism underlying these forms of synaptic plasticity is unknown although it is likely that intracellular Ca2+ and presynaptic Ca2+ channels play a critical role. Here we show that BDNF, NGF and NT-3 (10-100 ng/mL) exhibit a selective long-term up-regulation of voltage-gated Ca2+ current densities in developing hippocampal neurons of 6-20 days in culture. NGF and NT-3 appear more effective in up-regulating L-currents, while BDNF predominantly acts on non-L-currents (N, P/Q and R). The effects of the three neurotrophins were time- and dose-dependent. The EC50 was comparable for BDNF, NGF and NT-3 (10-16 ng/mL) while the time of half-maximal activation was significantly longer for NGF compared to BDNF (58 vs. 25 h). Despite the increased Ca2+ current density, the neurotrophins did not alter the voltage-dependence of channel activation, the kinetics parameters or the elementary properties of Ca2+ channels (single-channel conductance, probability of opening and mean open time). Neurotrophin effects were completely abolished by coincubation with the nonspecific Trk-receptor inhibitor K252a, the protein synthesis blocker anisomycin and the MAP-kinase inhibitor PD98059, while cotreatment with the PLC-gamma blocker, U73122, was without effect. Immunocytochemistry and Western blotting revealed that neurotrophins induced an increased MAP-kinase phosphorylation and its translocation to the nucleus. The present findings suggest that on a long time scale different neurotrophins can selectively up-regulate different Ca2+ channels. The action is mediated by Trk-receptors/MAP-kinase pathways and induces an increased density of newly available Ca2+ channels with unaltered gating activity.
神经营养因子对突触传递具有短期和长期影响。尽管细胞内Ca2+和突触前Ca2+通道可能起着关键作用,但这些形式的突触可塑性背后的机制尚不清楚。在这里,我们表明,脑源性神经营养因子(BDNF)、神经生长因子(NGF)和神经营养因子-3(NT-3,10-100 ng/mL)在培养6-20天的发育中的海马神经元中表现出电压门控Ca2+电流密度的选择性长期上调。NGF和NT-3在上调L型电流方面似乎更有效,而BDNF主要作用于非L型电流(N、P/Q和R型)。这三种神经营养因子的作用具有时间和剂量依赖性。BDNF、NGF和NT-3的半数有效浓度(EC50)相当(10-16 ng/mL),而与BDNF相比,NGF达到最大激活一半的时间明显更长(58小时对25小时)。尽管Ca2+电流密度增加,但神经营养因子并未改变通道激活的电压依赖性、动力学参数或Ca2+通道的基本特性(单通道电导、开放概率和平均开放时间)。与非特异性Trk受体抑制剂K252a、蛋白质合成阻滞剂茴香霉素和丝裂原活化蛋白激酶(MAP)抑制剂PD98059共同孵育可完全消除神经营养因子的作用,而与磷脂酶Cγ(PLC-γ)阻滞剂U73122共同处理则无效。免疫细胞化学和蛋白质印迹显示,神经营养因子诱导MAP激酶磷酸化增加并向细胞核转位。目前的研究结果表明,在长时间尺度上,不同的神经营养因子可以选择性地上调不同的Ca2+通道。其作用由Trk受体/MAP激酶途径介导,并诱导新可用的Ca2+通道密度增加,而门控活性未改变。