Authors/affiliations: Ufarry Alvarado, Axel*¹, Zaidi Pons, Malik*¹ and Torres Ortiz, Ceidy^1,2 Department of Biomedical Sciences¹, Department of Natural Sciences², Pontifical Catholic University of Puerto Rico, Ponce, Puerto Rico.

Introduction: Amyotrophic Lateral Sclerosis (ALS) is a motor neuron disease with 20% familial cases that are related to more than 100 missense mutations.  One of these genes is the superoxide dismutase-1 (sod-1) gene that codifies for an antioxidant protein. The sod-1(G85R)^C mutants in Caenorhabditis elegans (C. elegans) display protein inclusions and neurodegeneration in glutaminergic neurons after oxidative stress, defects in locomotion, impairment in autophagy and an increase in dysfunctional mitochondria overtime.  Therefore, we hypothesize that mitophagy, a type of selective autophagy, is impaired in an ALS model leading to an increase in behavior defects after oxidative stress induction.

Methods: The objective was to characterize worm behavior and survival of C. elegans sod-1(G85R)^C and WT (control) using nose touch avoidance, thrashing and Paraquat survival assays. Next, we evaluate possible neurodegeneration of neurons involved in the behavior test by applying a Dil staining assay. After evaluating neurodegeneration, our final objective is to construct and validate a C. elegans sod-1(G85R)^C model with ALS and fluorescent markers to determine mitophagy occurrence in neurons in the head region of the worm by measuring GFP/DsRed ratio. The worm mutant sod-1(G85R)^C was crossed with the worm strain SJZ42 that expresses mitochondrial specific Rosella in the pan-neuronal system. We validate and characterize the worm model with behavior assays along with C. elegans sod-1^G85R and WT.

Results: Behavior defects were displayed by a significant decrease in body bends in thrashing assays and a significant decrease in response in the nose-touch avoidance assays. These defects increase with oxidative stress exposure. In the Paraquat survival assay a significant decrease in survival percentage of sod-1(G85R)^C strain compared to WT was found. Only after oxidative stress exposure does the sod-1(G85R)^C strain have neurodegeneration of ASH, PHA and PHB glutaminergic neurons. The sod-1(G85R)^C and SJZ42 strain showed no mitophagy defects.

Conclusion: These results confirm that locomotion defects are a hallmark of ALS and oxidative stress exposure enhances these locomotion defects due to neurodegeneration. The neurodegeneration possibly is not related to mitophagy defects; however, further research is required.

Acknowledgements: Research was supported by an Institutional Development Award (IDeA) from the National Institute of General Medical Sciences of Health under the grant number P20GM103475, the Scientific Research Center of the PUCPR and Biomed Center of Brown University.