Bergmann M, Lahr G, Mayerhofer A, Gratzl M
Abteilung Anatomie und Zellbiologie, Universität Ulm, F.R.G.
Neuroscience. 1991;42(2):569-82. doi: 10.1016/0306-4522(91)90399-9.
The development of the spinal cord involves the proliferation of neurons, their migration to well-defined areas, fiber outgrowth and synapse formation. The present study was designed to correlate the spatiotemporal pattern of expression of synaptophysin, an integral membrane protein of small synaptic vesicles, with these basic processes occurring during the embryonic development of the rat spinal cord. Thoracic segments of spinal cords from embryonic days 12, 14, 16, 18, 20 and of adult spinal cords were studied. S1 nuclease protection assays and immunoblots revealed minute amounts of specific mRNA and synaptophysin at embryonic day 12. There was a steep increase of mRNA between embryonic days 14 and 16, after which levels reached a plateau. A rise in the amount of synaptophysin in the spinal cord occurred between embryonic days 12 and 14, and the levels changed only slightly until the end of embryonic development. Even higher levels of synaptophysin, found in the adult spinal cord, may indicate that its biosynthesis continued after birth. In situ hybridization histochemistry revealed the localization of specific synaptophysin mRNA in the neuroepithelium. However, immunocytochemistry failed to detect synaptophysin in the neuroepithelial cells. Following migration of the neuroblasts, synaptophysins was found in neurons concomitantly with the onset of fiber outgrowth. Thus, already at embryonic day 12, outgrowing fibers of the dorsal root sensory neurons and of motoneurons were synaptophysin positive. From embryonic day 14 throughout the prenatal period, strong synaptophysin immunoreactivity was seen in the ventrolateral and dorsal parts of the marginal layer. Most likely this staining pattern indicates transient functional synaptic contacts because, in the adult spinal cord, the corresponding region, the white matter, exhibited only faint synaptophysin immunoreactivity. In the intermediate layer of the embryonic spinal cord, which corresponds to the gray matter of the adult spinal cord, synaptophysin-positive fibers were observed prior to the formation of functional synapses. The latter are most likely permanent, since synaptophysin in the adult spinal cord is mainly confined to the gray matter. Our data (i) show transcription and translation of synaptophysin within the neurons of the spinal cord and correlate these processes with proliferation, migration, fiber outgrowth and the formation of transient or permanent synapses, and (ii) prove that synaptophysin is a marker for fiber outgrowth in addition to synapse formation.
脊髓的发育涉及神经元的增殖、向特定区域的迁移、纤维长出和突触形成。本研究旨在将小突触囊泡的整合膜蛋白突触素的时空表达模式与大鼠脊髓胚胎发育过程中发生的这些基本过程联系起来。研究了胚胎第12、14、16、18、20天的脊髓胸段以及成年脊髓。S1核酸酶保护分析和免疫印迹显示在胚胎第12天有微量的特异性mRNA和突触素。在胚胎第14天到16天之间mRNA急剧增加，之后水平达到平稳状态。脊髓中突触素的量在胚胎第12天到14天之间增加，并且在胚胎发育结束前水平仅有轻微变化。在成年脊髓中发现的更高水平的突触素可能表明其生物合成在出生后仍在继续。原位杂交组织化学揭示了特异性突触素mRNA在神经上皮中的定位。然而，免疫细胞化学未能在神经上皮细胞中检测到突触素。在神经母细胞迁移后，随着纤维长出的开始，在神经元中发现了突触素。因此，在胚胎第12天，背根感觉神经元和运动神经元的长出纤维就是突触素阳性的。从胚胎第14天到整个产前阶段，在边缘层的腹外侧和背侧可见强烈的突触素免疫反应性。很可能这种染色模式表明存在短暂的功能性突触接触，因为在成年脊髓中，相应区域即白质仅表现出微弱的突触素免疫反应性。在胚胎脊髓的中间层，即相当于成年脊髓灰质的区域，在功能性突触形成之前就观察到了突触素阳性纤维。后者很可能是永久性的，因为成年脊髓中的突触素主要局限于灰质。我们的数据（i）显示了脊髓神经元内突触素的转录和翻译，并将这些过程与增殖、迁移、纤维长出以及短暂或永久性突触的形成联系起来，并且（ii）证明突触素除了是突触形成的标志物外，还是纤维长出的标志物。
