Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disorder whose mechanisms underlying remain incompletely understood, particularly the role of glial cells. This study investigated the impact of ALS-associated genes on distinct glial populations in Drosophila. We assessed motor function and lifespan, revealing significant sexual dimorphism, with males generally showing greater declines. Our findings also underscore the importance of glial cells, particularly subperineurial glia (SPG) in the male leg femur, and provides valuable insights into the complex interplay between glial cells and ALS-associated genes.