Neuroscience Department, International School for Advanced Studies (SISSA), Trieste, Italy.
Biophys J. 2012 Jun 6;102(11):2451-60. doi: 10.1016/j.bpj.2012.04.036. Epub 2012 Jun 5.
We used optical tweezers to analyze the effect of jasplakinolide and cyclodextrin on the force exerted by lamellipodia from developing growth cones (GCs) of isolated dorsal root ganglia (DRG) neurons. We found that 25 nM of jasplakinolide, which is known to inhibit actin filament turnover, reduced both the maximal exerted force and maximal velocity during lamellipodia leading-edge protrusion. By using atomic force microscopy, we verified that cyclodextrin, which is known to remove cholesterol from membranes, decreased the membrane stiffness of DRG neurons. Lamellipodia treated with 2.5 mM of cyclodextrin exerted a larger force, and their leading edge could advance with a higher velocity. Neither jasplakinolide nor cyclodextrin affected force or velocity during lamellipodia retraction. The amplitude and frequency of elementary jumps underlying force generation were reduced by jasplakinolide but not by cyclodextrin. The action of both drugs at the used concentration was fully reversible. These results support the notion that membrane stiffness provides a selective pressure that shapes force generation, and confirm the pivotal role of actin turnover during protrusion.
我们使用光镊分析了卷曲霉素和环糊精对分离的背根神经节（DRG）神经元发育生长锥（GC）的片状伪足施加的力的影响。我们发现，已知抑制肌动蛋白丝周转率的 25 nM 卷曲霉素降低了片状伪足前缘突起过程中的最大作用力和最大速度。通过原子力显微镜，我们验证了环糊精（已知可从膜中去除胆固醇）降低了 DRG 神经元的膜刚度。用 2.5 mM 的环糊精处理的片状伪足施加的力更大，其前缘可以以更高的速度前进。卷曲霉素和环糊精都不影响片状伪足回缩过程中的力或速度。力产生的基本跳跃的幅度和频率因卷曲霉素而降低，但不因环糊精而降低。两种药物在使用浓度下的作用都是完全可逆的。这些结果支持了这样一种观点，即膜刚度提供了一种选择性压力，塑造了力的产生，并且证实了肌动蛋白周转在突起过程中的关键作用。
