The analysis of the role of transcription factors in the specification of spinal motor neurons position certainly yielded some important molecular handles on the problem of motor neuron soma migration. These are now being linked to specific effectors of neuronal migration that mediate signals from the outside environment to the neuronal cytoskeleton. Perhaps the best-known pathway that transduces such a signal is the one responding to the extracellular matrix protein Reelin (see also Chap.​ 1). It is a large, secreted protein, originally identified through a mutation in its gene, which leads to a locomotor behavioural phenotype due the aberrant localisation of cerebellar neurons (D’Arcangelo et al. 1995; Hamburgh 1963b). In the cortex, Reelin is deposited in the superficial layer by Cajal-Retzius neurons (Hirotsune et al. 1995; D’Arcangelo et al. 1995) while ApoER2 and Very Low Density Lipoprotein Receptor (VLDLR) are two principal Reelin receptors that transduce the signal to the cytoskeleton through the Dab1 adaptor phosphoprotein (Hiesberger et al. 1999; Trommsdorff et al. 1999). Mice, mutant for genes encoding these components, display severe disruptions in cortical layering, and on the basis of these phenotypes, Reelin has been postulated to act as a neuronal migration stop signal (Falconer 1951; Hamburgh 1963a; Hiesberger et al. 1999; Howell et al. 1997; Sheldon et al. 1997; Tissir and Goffinet 2003; Trommsdorff et al. 1999; Jossin and Cooper 2011). The downstream targets of this pathway include the cytoskeleton-associated proteins Lissencephaly1 (LIS1) and Doublecortin (DCX) (Tissir and Goffinet 2003). Reelin signalling, and its role in cortical projection neuron migration, is described in greater detail in Chap.​ 1.

Reelin is also expressed in the spinal cord and is required for the normal migration of PG neurons as well as the layering of the dorsal horn laminae (Yip et al. 2000, 2003a, 2004, 2009; Villeda et al. 2006; Phelps et al. 2002). Interestingly, the altered position of PG motor neurons in Reeler mutants appears to have no bearing on their connectivity to their ganglionic targets (Yip et al. 2003b) arguing that cell body position and axon projections of motor neurons can be uncoupled. This is also paralleled in more rostral regions of the nervous system where despite being inappropriately positioned, many neurons in the Reeler mouse project their axons to appropriate targets (Caviness 1976; Caviness and Frost 1980, 1983). Interestingly, the change in spinal dorsal horn layering organisation seen in Reeler mutants does have another important functional consequence: Reeler mutants have abnormal nociceptive responses (Wang et al. 2012), although in this case it is somewhat difficult to unequivocally exclude the contribution to this effect of inappropriately-developed rostral regions of the nervous system.

The specification of LMC soma position by transcription factors expressed in specific subpopulations of spinal motor neurons raised the question of whether these effects might be mediated by the Reelin pathway. Indeed, Reelin is expressed in the ventral spinal cord, and its receptors as well as the signalling intermediate Dab1 are expressed by LMC neurons (Palmesino et al. 2010). Moreover, Reelin and Dab1 mutants display LMC soma position defects: while LMC cell bodies are found in their appropriate general ventrolateral location in the spinal cord, the lateral LMC and medial LMC neuron positions are inverted along the mediolateral axis. One intriguing explanation for this phenotype is that Dab1 is expressed differentially in these neurons, with lateral LMC neurons having higher levels of the protein, compared to medial LMC. The model that emerged from additional gain and loss of function experiments is that Dab1 expression gates the LMC neurons’ sensitivity to Reelin, such that lateral LMC neurons display high Dab1 levels and thus are sensitive to Reelin located close to the ventricular zone. In contrast, medial LMC neurons display lower levels of Dab1, and thus are relatively insensitive to Reelin. This model, which is further supported by Reelin overexpression experiments (E. Palmesino and A. Kania, unpublished), posits that Reelin is not an absolute migration stop signal (D’Arcangelo et al. 1995; Ogawa et al. 1995), but rather a repulsive or migration promoting cue for LMC motor neurons, an idea in line with previous experiments in which ectopic overexpression of Reelin rescues cortical migration defects (Magdaleno et al. 2002).

The differential Dab1 expression in medial and lateral LMC neurons suggested an intriguing possibility that these levels are controlled by the same transcription factors that control LMC soma position. Indeed, Foxp1 and Lim1 gain or loss of function result in, respectively, increase or decrease of Dab1 expression in LMC neurons, and are the first example of the control of Reelin pathway protein expression by specific transcription factors (Palmesino et al. 2010). Moreover, since the same transcription factors also control LMC axon trajectory in the limb, these observations suggest a molecular hierarchy where specific transcription factors co-ordinately control both cell body position and axon trajectory, and thus establish myotopic organisation. At this point, it is unclear whether this idea can be extended to cortical or cerebellar neurons, two populations in which Reelin signaling and migration as well as molecular differentiation have been studied.

This large family of proteins has been associated with many types of adhesive cell-cell interactions (Nollet et al. 2000). Within the nervous system, cadherins are selectively expressed in a combinatorial manner and thus, can be used to subdivide many neuronal populations (Suzuki et al. 1997). This property, combined with their adhesive function, endows them with the ability to control neuronal soma position, as in the case of hindbrain motor neurons (Garel et al. 2000). In the context of spinal motor neurons, type II cadherin expression is used to delineate specific motor pools, suggesting a possible role in motor pool sorting. Indeed, gain and loss of function experiments in this system, where the expression of a single cadherin protein is altered, result in inappropriate clustering of motor neurons such that motor neurons from one transcription factor-defined motor pool are mixed with those from another pool (Price et al. 2002). Additional experiments argue that cadherin-mediated control of LMC neuron migration occurs through catenins, cytoplasmic effectors that link the membrane-bound cadherins to the underlying cytoskeleton (Bello et al. 2012; Price 2012). Extending this idea further, Jessell and his colleagues used mouse genetics to block catenin function and cadherin signalling in spinal motor neurons (Demireva et al. 2011). In such mice, the LMC neuron pool and division positions, as well as those of PG neurons are scrambled, while molecular identity is maintained. Moreover, the limb axonal trajectory of such motor neurons appears normal, providing further evidence that motor neuron soma position and axon trajectory can be uncoupled through the inactivation of specific molecular effectors. Conversely, in such experiments, the link between transcriptional identity and axon trajectory is maintained.

One outstanding question raised by these experiments is how cadherins are acting to selectively position spinal motor neuron cell bodies. In order for the selective adhesion mediated by combinations of cadherins to exert its effect on soma position and thus to allow for the different combinations of cadherins to interact with each other, motor neuron cell bodies need to have an opportunity to interact with one another. For this to happen, cell bodies of motor neurons could be mobilised randomly, like particles exhibiting Brownian motion, or in a more directed fashion, perhaps away from a source of a repellent, such as Reelin. Thus the relative contribution of Reelin signaling and cadherin-mediated adhesion in the specification of motor neuron position becomes an important question. The LMC soma positioning defects in catenin inactivation mutants are apparently more severe than those found in Reelin and Dab1 mutants. It might be that in order for Reelin signaling to occur, close cell-cell contacts need to form, something that could be precluded by inhibition of cadherin signalling. Thus, Reelin signalling in spinal motor neurons, might be dependent on cadherin function, and it will be interesting to evaluate in the future if in motor neurons the intersection of Reelin and cadherin signaling occurs in a mechanistically similar manner like in cortical projection neurons (see also Chap.​ 1).

The Eph receptor tyrosine kinases and their ephrin ligands are also important protein families that are expressed in many developing tissues, including the nervous system. They have been implicated in cell sorting, where cells that are initially intermingled become sorted into two separate compartments and thus form a boundary (Klein 2012). One seminal observation has been that of the function of Eph receptors in the compartmentalisation of hindbrain, where molecularly distinct neurons in adjacent rhombomeres form a sharp boundary (Xu et al. 1999). Unlike cadherins, there is ample evidence of the involvement of Eph receptors and their ligands in motor axon trajectory selection (Kao et al. 2012). Thus, ephrin-Eph signaling is well positioned to influence both motor neuron soma position and axon trajectory, coordinating their myotopic organisation and raising the question whether Eph signalling is involved in spinal motor neuron migration. In mice lacking EphA, a receptor important for the correct targeting of lateral LMC axons to dorsal limb muscles, the tibialis motor neuron pool is displaced within the lumbar spinal cord, arguing that this receptor is important for the normal positioning of spinal motor neuron cell bodies (Coonan et al. 2003). Interestingly, in these animals, the tibialis motor neurons still innervate their correct muscle targets, suggesting that Eph receptors can specify LMC soma position independently of axonal trajectory. This is particularly interesting given the expression patterns of ephrins and Eph receptor proteins at the level of spinal motor neuron cell bodies, and is very suggestive of their restriction to specific motor pools (Iwamasa et al. 1999).

Although at this point there is no evidence of their function in neuron soma migration, semaphorins and their transmembrane receptors neuropilins have elaborate expression patterns at the level of hindbrain and spinal motor neuron somata (Cohen et al. 2005; Huber et al. 2005). There is accumulating evidence that these families of proteins function in the specification of motor axon trajectory in the limb (Huber et al. 2005; Moret et al. 2007; Huettl et al. 2011; Sanyas et al. 2012). Interestingly, since semaphoring-neuropilin signaling has been previously implicated in cortical neuron migration (Chen et al. 2008), it would not be at all surprising if such signaling also impacts spinal motor neuron soma localisation, although so far no evidence of this function has been reported.