The Questions of Developmental Biology

Guidance by diffusible molecules
Netrins and their receptors.
The idea that chemotactic cues guide axons in the developing nervous system was first
proposed by Santiago Ramón y Cajal (1892). He suggested that the commissural neurons of the
spinal cord might be told by diffusible factors to send axons from their dorsal positions to the
ventral floor plate. Commissural neurons are interneurons that cross the ventral midline to
coordinate right and left motor activities. Thus, they somehow must migrate to (and through) the
ventral midline. The axons of these neurons begin growing ventrally down the side of the neural
tube. However, about two-thirds of the way down, their direction changes, and they project
through the ventrolateral (motor) neuron area of the neural tube toward the floor plate cells
(Figure 13.19).
In 1994, Serafini and colleagues developed an assay that
would allow them to screen for such a diffusible molecule. When
dorsal spinal cord explants from chick embryos were plated onto
collagen gels, the presence of floor plate cells near them promoted
the outgrowth of commissural axons. Serafini and his co-workers
took fractions of embryonic chick brain homogenate and tested
them to see if any of the proteins therein mimicked this activity.
This process resulted in the identification of two proteins, netrin-1
and netrin-2. Netrin-1 is made by and secreted from the floor plate
cells, whereas netrin-2 is synthesized in the lower region of the
spinal cord, but not in the floor plate (Figure 13.19B). The
chemotactic effects of these netrins were demonstrated by
transforming chick fibroblast cells (which usually do not make these
proteins) into netrin-producing cells using a vector containing an
active netrin gene (Kennedy et al. 1994). Aggregates of these netrinsecreting
fibroblast cells elicited commissural axon outgrowth from
dorsal rat spinal cord explants, while control cells that were given
the vector without the active netrin gene did not elicit such activity
(Figure 13.20). It is possible that the commissural neurons first
encounter a gradient of netrin-2, which brings them into the domain
of the steeper netrin-1 gradient (see Figure 13.19A).