The spinal cord and brain harbor a variety of motor neurons that innervate limbs, thorax, neck, or face. Different classes
of motor neurons reside at different locations along the head-to-tail (rostrocaudal) axis of the
nervous system, reflecting a patterning that arises early in embryonic development, when the nervous system is but a simple tube. Indeed, different sections of the neural tube begin expressing distinct subsets of genes thought to govern cell fate, such as members of the
Hox gene family, shortly after the tube forms. The question is how this patterned gene expression arises. Ulrika Nordstrm, Thomas Edlund, and their colleagues have tackled this problem in the chick embryo. They report that rostrocaudal patterning of the spinal cord and
hindbrain is under the influence of the signaling molecule Wnt. Other signals refine Wnt patterning and create distinct sections in which different types of motor neurons eventually differentiate.
The embryonic nervous system of vertebrates is shaped by signals coming from surrounding tissues, in particular from the mesoderm, which gives rise to skeletal bones and muscles. The mesodermal signal fibroblast growth factor (FGF), for instance, imposes caudal fates on the developing nervous system, whereas Wnt and FGF in combination pattern the middle portions of the prospective brain. Since Wnt remains abundant in regions where the spinal cord will start to grow behind the nascent brain, Nordstrm and her colleagues reasoned that it might also pattern the posterior regions of the embryos nervous system.
The researchers cut out small fragments (called explants) of prospective neural tissue from chick embryos that had reached different developmental stages and observed the explants development in culture. At developmental stage 4, the chick embryo is a pin-head-size disc of cells that lies atop the yolk. Its most prominent feature is the primitive streak, a thin furrow that starts as an anterior dimple known as Hensens node and runs all the way to the embryos posterior end. Cells destined to become neural occupy a horseshoe-shaped halo around Hensens node. The anterior and middle portions of the brain derive from the horseshoes arch, whereas hindbrain and spinal cord come from its branches.
The researchers found that stage 4 explants taken from the horseshoes branch expressed
Hoxc9,
Hoxb8 and
Hoxb4 in its caudal half and
Krox20 in its rostral half after a day in culture. The
Hox gene combination is characteristic of the lumbar and tail region of the spinal cord, whereas expression of
Krox20 marks the anterior hindbrain. When cultured in the presence of an inhibitor of Wnt signaling, the explants failed to turn on
Krox20 and the
Hox genes, and instead expressed
Otx2, a marker of the forebrain. Therefore, Wnt signaling appears necessary for the neural tube to adopt the posterior characteristics of hindbrain and spinal cord.
Missing from the cultured explants, however, were cells expressing a combination of
Hoxb8 and
Hoxb4, which mark the thoracic and neck regions of the spinal cord, or
Hoxb4 alone, which marks the posterior hindbrain. By contrast, these cells appeared readily in cultured explants taken from stage 8 embryos. By this stage, mesoderm has crept through the backward-sliding Hensens node to lie under the elongating neural tube, where it begins to organize into somites, the precursors of vertebrae and ribs. The researchers reasoned that retinoic acid, a signal secreted by somites, might combine with Wnt to specify hindbrain and anterior spinal cord fates.
Adding retinoic acid to cultures of stage 4 explants confirmed this hypothesis. retinoic acid suppressed the expression of
Krox20 and
Hoxc9, while inducing the appearance of cells expressing
Hoxb8 and
Hoxb4, or
Hoxb4 alone. If the cultures also contained FGF, which normally diff