dissection-immunohistochemistry-drosophila-adult-leg-to-detect

Dissection and Immunohistochemistry of the <em>Drosophila</em> Adult Leg to Detect Changes at the Neuromuscular Junction for an Identified Motor Neuron

Drosophila melanogaster represents a genetically tractable model to study neuronal structure and function, and subsequent changes in disease states. The ...

Rhode Island College

Warren Alpert Medical School of Brown University

Mutations in superoxide dismutase 1 (SOD1) cause familial amyotrophic lateral sclerosis (ALS) in humans. ALS is a neurodegenerative disease characterized by progressive motor neuron loss leading to paralysis and inevitable death in affected individuals. Using a gene replacement strategy to introduce disease mutations into the orthologous  () gene, here, we characterize changes at the neuromuscular junction using longer-lived  mutant adults. Homozygous  or  flies display progressive walking defects with paralysis of the third metathoracic leg. In dissected legs, we assessed age-dependent changes in a single identified motor neuron (MN-I2) innervating the tibia levitator muscle. At adult eclosion, MN-I2 of  or  flies is patterned similar to wild-type flies indicating no readily apparent developmental defects. Over the course of 10 days post-eclosion, MN-I2 shows an overall reduction in arborization with bouton swelling and loss of the post-synaptic marker  () in mutant  adults. In addition, increases in polyubiquitinated proteins correlate with the timing and extent of MN-I2 changes. Because similar phenotypes are observed between flies homozygous for either  or  alleles, we conclude these NMJ changes are mainly associated with  loss-of-function. Together these studies characterize age-related morphological and molecular changes associated with axonal retraction in a  model of ALS that recapitulate an important aspect of the human disease.This article has an associated First Person interview with the first author of the paper.

Anthony Agudelo

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