Institut Pasteur, Laboratory for Perception and Memory, F-75015 Paris, France.
J Neurosci. 2011 May 18;31(20):7551-62. doi: 10.1523/JNEUROSCI.6716-10.2011.
Neuronal precursors are continuously integrated into the adult olfactory bulb (OB). The vast majority of these precursor cells originates from the subventricular zone and migrates along the rostral migratory stream (RMS) en route to the OB. This process, called postnatal neurogenesis, results from intricate pathways depending both on cell-autonomous factors and extrinsic regulation provided by the local environment. Using electroporation in postnatal mice to label neuronal precursors with green fluorescent protein (GFP) and to reduce the expression levels of doublecortin (DCX) with short-hairpin (Sh) RNA, we investigated the consequences of impairing migration on the fate of postnatal-formed neurons. First, we showed that electroporation of Dcx ShRNA plasmid efficiently knocks down the expression of DCX and disrupts cells migration along the RMS. Second, we found misplaced anomalous migrating cells that displayed defects in polarity and directionality. Third, patch-clamp recordings performed at 5-7 days post-electroporation (dpe) revealed increased density of voltage-dependent Na(+) channels and enhanced responsiveness to GABA(A) receptor agonist. At later time points (i.e., 12 and 30 dpe), most of the Dcx ShRNA(+) cells developed in the core of the OB and displayed aberrant dendritic length and branching. Additional analysis revealed the formation of GABAergic and glutamatergic synaptic inputs on the mispositioned neurons. Finally, quantifying fate determination by numbering the proportion of GFP(+)/calretinin(+) newborn neurons revealed that Dcx ShRNA(+) cells acquire mature phenotype despite their immature location. We conclude that altering the pace of migration at early stages of postnatal neurogenesis profoundly modifies the tightly orchestrated steps of neuronal maturation, and unveils the influence of microenvironment on controlling neuronal development in the postnatal forebrain.
神经前体细胞不断整合到成年嗅球(OB)中。这些前体细胞的绝大多数来源于侧脑室下区,并沿着嗅球迁移的吻侧迁移流(RMS)迁移到 OB。这个过程被称为出生后神经发生,是由细胞自主因素和局部环境提供的外在调节的复杂途径所决定的。我们在出生后的小鼠中使用电穿孔将绿色荧光蛋白(GFP)标记神经元前体细胞,并使用短发夹(Sh)RNA 降低双皮质素(DCX)的表达水平,以研究破坏迁移对出生后形成的神经元命运的影响。首先,我们表明,Dcx ShRNA 质粒的电穿孔有效地降低了 DCX 的表达,并破坏了细胞沿 RMS 的迁移。其次,我们发现错位的异常迁移细胞表现出极性和方向性缺陷。第三,在电穿孔后 5-7 天(dpe)进行的膜片钳记录显示电压依赖性 Na(+)通道的密度增加,并且对 GABA(A)受体激动剂的反应增强。在较晚的时间点(即 12 和 30 dpe),大多数 Dcx ShRNA(+)细胞在 OB 的核心中发育,并表现出异常的树突长度和分支。进一步的分析显示,在错位的神经元上形成了 GABA 能和谷氨酸能突触输入。最后,通过对 GFP(+)/calretinin(+)新生神经元的比例进行计数来定量命运决定,表明 Dcx ShRNA(+)细胞尽管位于不成熟的位置,但仍获得成熟的表型。我们的结论是,在出生后神经发生的早期改变迁移速度会深刻改变神经元成熟的协调步骤,并揭示了微环境对控制出生后前脑神经元发育的影响。