Juvenile rainbow trout, Oncorhynchus mykiss, were injected with estradiol-17β (E2) in order to study the source of extra calcium needed during vitellogenesis. E2-treatment increased the calcium uptake from the external medium as well as calcium mobilization from muscle and scale. Judged by the increase in plasma protein-bound calcium levels, the E2-induced increase in calcium uptake is an apparent over-mobilization of calcium, i.e., the calcium uptake of the fish is in excess of what is found bound to plasma proteins. As the calcium excretion and calcium space (calculated from free plasma calcium levels) were unaffected, the excess calcium is suggested to be incorporated into internal calcium stores. This implies that the systems regulating vitellogenesis and calcium balance are integrated on the mechanistic or endocrine level, and that E2 causes calcium mobilization of a magnitude geared to the needs of the sexually maturing female.