Assessing potential effects of flow regulation on southeastern warmwater fish assemblages is problematic because of high species richness and our poor knowledge of habitat requirements for most species. A previous attempt to reduce the complexity of describing habitat requirements for diverse assemblages defined five 'key habitat' types based on quantitative descriptions of depth, velocity, substrate, and cover for assessing the effects of streamflow alteration on fish communities. Our study investigated relationships between availability and temporal stability of key habitats and fish abundances at regulated and unregulated sites in the Tallapoosa River system. Fish assemblage characteristics at seven sites were quantified based on 1,400 electrofishing samples collected during 1994 and 1995. Simulations were used to model availability and temporal stability of key habitats at regulated and unregulated sites, Associations between fish assemblages and availability or stability of key habitats were identified using correlation analysis. We found that hydropeaking dam operation reduced the average length of time that shallow-water habitats were stable during the spring and summer and also reduced year-to-year variation in the stability of shallow-water habitats compared to unregulated sites. Within-site comparisons of fish and habitat variables indicated that differences in fish abundances correlated with differences in the availability and temporal stability of shallow-water habitats. Additionally, groups of stream fishes defined by taxonomy or differences in orientation to the substrate and feeding mode responded similarly to changes in key habitat availability. These findings demonstrate that the temporal and spatial availability of key habitats could serve as a useful measure of the potential effects of flow alteration on lotic fish assemblages, and suggest that both short-term temporal stability of key habitats as well as annual variation in key habitat availability are important for maintaining diverse fish assemblages.