Familial dysautonomia (FD) is characterized by severe and progressive sympathetic and sensory neuron loss caused by a highly conserved germline point mutation of the human ELP1/IKBKAP gene. Elp1 is a subunit of the hetero-hexameric transcriptional elongator complex, but how it functions in disease-vulnerable neurons is unknown. Conditional knockout mice were generated to characterize the role of Elp1 in migration, differentiation and survival of migratory neural crest (NC) progenitors that give rise to sympathetic and sensory neurons. Loss of Elp1 in NC progenitors did not impair their migration, proliferation or survival, but there was a significant impact on post-migratory sensory and sympathetic neuron survival and target tissue innervation. Ablation of Elp1 in post-migratory sympathetic neurons caused highly abnormal target tissue innervation that was correlated with abnormal neurite outgrowth/branching and abnormal cellular distribution of soluble tyrosinated α-tubulin in Elp1-deficient primary sympathetic and sensory neurons. These results indicate that neuron loss and physiologic impairment in FD is not a consequence of abnormal neuron progenitor migration, differentiation or survival. Rather, loss of Elp1 leads to neuron death as a consequence of failed target tissue innervation associated with impairments in cytoskeletal regulation.

The rodent superior cervical ganglion (SCG) is a useful and readily accessible source of neurons for studying the mechanisms of sympathetic nervous system (SNS) development and growth in vitro. The sympathetic nervous system (SNS) of early postnatal animals undergoes a great deal of remodeling and development; thus, neurons taken from mice at this age are primed to re-grow and establish synaptic connections after in situ removal. The stereotypic location and size of the SCG make it ideal for rapid isolation and dissociation. The protocol described here details the requirements for the dissection, culture and differentiation of SCG neurons. The protocol is suitable for culturing neurons from late embryonic gestation to approximately postnatal day 3. The culture technique discussed below utilizes glass coverslips for the microscopic examination of fixed cells.

A highly conserved germline point mutation located in the donor splice site of intron 20 of the human Elp1 gene leads to loss of its encoded protein and causes familial dysautonomia (FD; Riley‐Day Syndrome; hereditary sensory and autonomic neuropathy, type 3; HSAN3), an autosomal recessive disease characterized by severe and progressive sympathetic and sensory neuron loss. The protein encoded by elp1 is required for development beyond mid‐gestation and it is part of the transcriptional elongator multi‐protein complex, but its functional role in disease pathogenesis remains poorly understood. To examine the role of elp1 in sympathetic and sensory neuron development, we generated conditional elp1 knockout mice and we used Wnt1‐Cre transgenic mice to ablate elp1 in neural crest (NC) cells, a highly migratory population of cells which gives rise to sympathetic, sensory and enteric neurons during development. Loss of elp1 in NC cells showed no impairment in their migration, but there was a significant impact on post‐migratory sensory and sympathetic neuron survival and target tissue innervation. In vitro studies using elp1‐deficient neurons showed that it has a profound impact on growth factor‐induced neurite outgrowth but no role in neuron survival. These results indicate that FD is not caused by impaired neuronal migration but rather, neuron death resulting from ineffective target tissue innervation.

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