An artificial selection experiment was conducted for six generations to investigate the responses to selection for cadmium resistance in the least killifish, Heterandria formosa. There was a rapid response to selection. After only one generation of selection, two of the three selection lines had an increased resistance to cadmium, while all three selection lines had an elevated resistance by the next generation. After six generations of selection, fish from the selection lines survived about three times as long as control line fish when exposed to cadmium. Realized heritability for cadmium resistance was estimated to be 0.50. Cross-resistance to copper was studied in the F2, F3, and F6 generations; fish from the cadmium-resistant lines had a longer survival time than those from the control lines. Resistance to heat was investigated in the F2, F3, and F5 generations; the cadmium-resistant lines had a significantly shorter survival time than the control lines. Our results showed that evolution of resistance to cadmium was rapid, that cross-resistance to copper occurred, and that the evolution of resistance to cadmium was accompanied by a trade-off (fitness costs) as evidenced by the reduced heat tolerance.