Intracellular calcium and cyclic nucleotide levels modulate neurite guidance by microtopographical substrate features.
Intracellular calcium and cyclic nucleotide levels modulate neurite guidance by microtopographical substrate features.
复制标题
细胞内钙和环状核苷酸水平通过微观形态底物特征调节神经突引导。
DOI：
10.1002/jbm.a.35738
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发表时间：
2016-08
期刊：
影响因子：
--
通讯作者：
Hansen MR
中科院分区：
文献类型：
--
作者：
Li S;Tuft B;Xu L;Polacco M;Clarke JC;Guymon CA;Hansen MR
微米级和纳米级的表面特征已经成为引导神经突生长到与下一代神经假体电极非常接近的潜在工具。然而，潜在的信号事件的能力，生长锥，以应对地形特征仍然在很大程度上未知。因此，本研究探讨了[Ca 2 +]i和环核苷酸水平对螺旋神经节神经元（SGNs）的神经突精确跟踪地形微图案的能力的影响。使用光聚合和光掩模来产生微图案化的甲基丙烯酸酯聚合物基底。将解离的SGN培养物铺在微图案化表面上。通过升高细胞外K+、在无钙培养基中维持或浴中应用各种钙通道阻滞剂来操纵钙内流和从内部储存中释放。环核苷酸活性增加的应用cpt-cAMP或8-Br-cGMP。升高[Ca 2 +]i的治疗培养物与高钾减少神经突对齐物理微特征。在无钙培养基中培养或用非选择性电压门控钙通道阻滞剂镉或L型钙通道阻滞剂硝苯地平处理不会显著改变SGN神经突排列。相比之下，ryanodine或xestospongin C，分别通过ryanodine敏感通道或肌醇-1，4，5-三磷酸受体阻断内部钙储存的释放，各自显着降低神经突排列。Cpt-cAMP显著降低神经突排列，而8-Br-cGMP显著增强神经突排列。[Ca 2 +]i或cAMP水平的操纵显著地破坏神经突引导，而cGMP水平的升高增加神经突排列。结果表明，细胞内信号通路类似于那些招募的趋化性线索参与神经突的地形特征的指导。
Micro- and nano-scale surface features have emerged as potential tools to direct neurite growth into close proximity with next generation neural prosthesis electrodes. However, the signaling events underlying the ability of growth cones to respond to topographical features remain largely unknown. Accordingly, this study probes the influence of [Ca2+]i and cyclic nucleotide levels on the ability of neurites from spiral ganglion neurons (SGNs) to precisely track topographical micropatterns. Photopolymerization and photomasking were used to generate micropatterned methacrylate polymer substrates. Dissociated SGN cultures were plated on the micropatterned surfaces. Calcium influx and release from internal stores were manipulated by elevating extracellular K+, maintenance in calcium-free media, or bath application of various calcium channel blockers. Cyclic nucleotide activity was increased by application of cpt-cAMP or 8-Br-cGMP. Elevation of [Ca2+]i by treatment of cultures with elevated potassium reduced neurite alignment to physical microfeatures. Maintenance of cultures in Ca2+-free medium or treatment with the non-selective voltage-gated calcium channel blocker cadmium or L-type Ca2+ channel blocker nifedipine did not signficantly alter SGN neurite alignment. By contrast, ryanodine or xestospongin C, which block release of internal calcium stores via ryanodine-sensitive channels or inositol-1,4,5-trisphosphate receptors respectively, each significantly decreased neurite alignment. Cpt-cAMP significantly reduced neurite alignment while 8-Br-cGMP significantly enhanced neurite alignment. Manipulation of [Ca2+]i or cAMP levels significantly disrupts neurite guidance while elevation of cGMP levels increases neurite alignment. The results suggest intracellular signaling pathways similar to those recruited by chemotactic cues are involved in neurite guidance by topographical features.