Martinou J C, Bierer F, Le Van Thai A, Weber M J
Laboratoire de Pharmacologie et Toxicologie Fondamentales, Centre National de la Recherche Scientifique, Toulouse, France.
Brain Res Dev Brain Res. 1989 Jun 1;47(2):251-62. doi: 10.1016/0165-3806(89)90180-6.
Motoneurons from E14 rat embryos have been retrogradely labelled with a carbocyanine derivative and purified approximately 20-fold by density gradient centrifugation. For certain experiments, motoneurons have been purified to near homogeneity by cell sorting. These neurons were then cultured at low density on various substrata. Laminin stimulated neurite outgrowth of the identified motoneurons and stimulated choline acetyltransferase (ChAT) development by 2-fold; these cultures could not be maintained for more than 4 days. Motoneurons could be cultured for at least 7 days on monolayers of both living and lysed astrocytes from newborn cortex. The cells survived equally well on both substrata but neurite outgrowth was less elaborated on lysed astrocytes on which motoneurons displayed a mono- or bipolar, rather than a multipolar, morphology. Furthermore, ChAT developed at a 2-fold higher level on lysed, rather than living, newborn astrocytes. Such a difference was not observed using astrocytes from E14 rat embryos. In the latter case, both living and lysed astrocytes supported ChAT development at the same rate as lysed newborn astrocytes. Consequently, living astrocytes from newborn rat cortex are relatively less permissive for ChAT expression. On the other hand, both living and lysed astrocytes from newborn cortex induced ChAT expression by sympathetic neurons to the same extent. We propose that, in addition to retrograde factors from muscle origin, cell surface molecules expressed by embryonic astrocytes stimulate ChAT expression by motoneurons. Such molecules would be present in newborn astrocytes in an intracellular form and could be unmasked by cell lysis. Different cell surface molecules may induce ChAT activity in sympathetic neurons.
来自E14大鼠胚胎的运动神经元已用一种碳花青衍生物进行逆行标记，并通过密度梯度离心法纯化了约20倍。在某些实验中，通过细胞分选将运动神经元纯化至接近同质。然后将这些神经元以低密度培养在各种基质上。层粘连蛋白刺激了已鉴定运动神经元的神经突生长，并使胆碱乙酰转移酶（ChAT）的发育增加了2倍；这些培养物无法维持超过4天。运动神经元可以在新生皮质的活星形胶质细胞和裂解的星形胶质细胞单层上培养至少7天。细胞在两种基质上存活情况相同，但在裂解的星形胶质细胞上神经突生长较不精细，运动神经元在其上呈现单极或双极而非多极形态。此外，ChAT在裂解的而非活的新生星形胶质细胞上发育水平高2倍。使用来自E14大鼠胚胎的星形胶质细胞未观察到这种差异。在后一种情况下，活的和裂解的星形胶质细胞支持ChAT发育的速率与裂解的新生星形胶质细胞相同。因此，新生大鼠皮质的活星形胶质细胞对ChAT表达的允许性相对较低。另一方面，新生皮质的活星形胶质细胞和裂解的星形胶质细胞诱导交感神经元ChAT表达的程度相同。我们提出，除了来自肌肉来源的逆行因子外，胚胎星形胶质细胞表达的细胞表面分子刺激运动神经元的ChAT表达。这些分子在新生星形胶质细胞中以细胞内形式存在，可通过细胞裂解而暴露。不同的细胞表面分子可能诱导交感神经元中的ChAT活性。
