Spine length, pelvic girdle morphology and body form vary markedly among populations of the brook stickleback (Culaea inconstans). Predation may drive the evolution of well developed defensive structures because these traits increase handling times and allow captured prey to escape from small predators. Previous studies indicate that C. inconstans from Macochee Creek (Champaign Co., Ohio), a tributary of the Mad River, appear to have spine lengths that approach the extremes for the species. However, it is unclear whether long spines are common throughout the Mad River drainage or are limited to this single tributary. Morphometric analysis revealed that C. inconstans from the Mad River drainage have longer spines than other populations in its southeastern range and that long spines are consistently found throughout the Mad River drainage. The selection pressures responsible for long spines in C. inconstans from the Mad River drainage are presently not clear.