Developmental mechanisms of sympathetic axon pathfinding, target selection and neurotransmitter expression

Sympathetic neuron development is characterized by the progressive evolution of undifferentiated multipotent stem cells into highly differentiated neurons, governed to a large part by extrinsic instructive signals. Identification of the mechanisms responsible for these processes is critical for a comprehensive understanding of sympathetic nervous system formation. My thesis studies have addressed several of these issues. The first is the mechanism(s) by which sympathetic axons pathfind to and select appropriate targets during development. Specifically, do target cells provide instructive cues for pathfinding sympathetic axons? Analysis of tabby mutant mice in which one sympathetic target, footpad eccrine sweat glands, is genetically lesioned revealed that gland-targeted axons extend to footpads in a stereotyped fashion and that the presence of innervation target tissue is not essential for axons to pathfind to their presumptive target site. While target cells do not appear to be required for long range pathfinding, previous studies have suggested that gradients of nerve growth factor (NGF) in peripheral tissues may aid in target selection by attracting sympathetic axons to appropriate target cells. To examine this issue, I have analyzed transgenic mice (K14-NGF) in which the normal levels of NGF expressed in footpad target tissue is altered. Developmental overexpression of NGF in epidermal keratinocytes results in reorganization of innervation patterning so that sympathetic sweat gland innervation is lost in favor of ectopic innervation of the epidermis, normally a target exclusively for sensory axons. Consequently, maturational changes in neurotransmitter coding which normally occur in sweat gland innervation failed to occur in the inappropriately localized sympathetic terminals. Studies have shown previously that sweat glands are responsible for this coding change and provide an instructive signal which causes noradrenergic innervation to acquire cholinergic/peptidergic properties during development. A detailed morphological study of this coding shift via analysis of immunoreactivity for the vesicular acetylcholine transporter (VAChT) revealed that a sympathetic noradrenergic plexus is well established in association with developing sweat glands before VAChT immunoreactivity becomes apparent. Together, these studies indicate that sympathetic neuron differentiation, even during the final stages of development, is highly dependent on extrinsic instructive signals.