Locomotion is a primary means by which animals interact with the world. To understand the contribution of genotype x environment interactions to individual differences in D. melanogaster larval locomotion we investigated phenotypic sensitivity to environmental changes in four strains of this species and their F-1 hybrids. We also investigated to what extent flexibility and plasticity of locomotion depend upon larval age. Specifically, we examined larval locomotion at 48 and 96 h of development on three different substrates. Locomotion was influenced by the structure of the substrate, but this depended on both the genotype and larval age. At 48 h of larval development phenotypic variation in locomotion was attributable to both genotype x environment interactions and genotypic differences among the larvae, while at 96 h of age, differences were mainly due to genotype x environment interactions. An analysis of variance of the 4 x 4 diallel cross made at 48 and 96 h of development showed, depending on the cross, either dominance to increase/decrease locomotion, overdominance to increase/decrease locomotion, or no dominance to increase/decrease locomotion. Furthermore, the diversity of behavioral responses in the F-1 hybrids was greater at 96 than at 48 h of larval development. Our results demonstrate that genotype x environment interactions result in plasticity of D. melanogaster larval locomotion, which makes sense in light of the fact that larvae, in the wild, develop in heterogeneous and rapidly changing environments.