Locomotor behavior in terrestrial organisms is crucial for burrowing, avoiding predators, food seeking, migration, and reproduction; therefore, it is a parameter with ecological relevance. Acetylcholinesterase (AChE) is a nervous system enzyme inhibited by several compounds and widely used as an exposure biomarker in several organisms. Moreover, changes in energy reserves also may indicate an exposure to a stress situation. The aim of this study is to link biomarkers of different levels of biological organization in isopods exposed to increasing doses of dimethoate in semifield conditions. Locomotor parameters, AChE activity, and energy reserves (lipid, glycogen, and protein contents) were evaluated in the isopod Porcellio dilatatus after 48-h and 10-d exposure to dimethoate-contaminated soil. Results showed a clear impairment of both locomotor and AChE activity during the entire study, although effects were more pronounced after 48 h. Most locomotor parameters and AChE activity showed a clear dose-response relationship. By contrast, no clear trend was observed on energetic components. A positive and significant relationship was found between AChE activity and those locomotor parameters indicating activity, and the opposite was observed with those locomotor parameters indicating confusion and disorientation. The results obtained in this study enhance the importance of linking biochemical responses to parameters with ecological relevance at individual level, the value of locomotor behavior as an important marker to assess effects of toxicants, and also the usefulness and the acquisition of ecological relevance by AChE as a biomarker, by linking it with ecologically relevant behavioral parameters.