Effects of postnatal development, surgical cryptorchidism, and hypophysectomy on the activity of calmodulin and of a 32,000-molecular weight (32) calmodulin-binding protein (Ono et al., J. Biol. Chem. 259: 9011-9016, 1984) were examined in rat testis. In adult rats, approximately 95% of their activities distributed in seminiferous tubule preparation, and most of them were in the 105,000-g supernatant. Both activities were low early in the development, but the calmodulin activity increased between 25 and 45 days of age, and the 32 K calmodulin-binding protein activity increased between 35 and 45 days of age. In contrast, surgical cryptorchidism caused a rapid decline in their activities in the abdominal testis, whereas their activities in a scrotal testis did not decline. Hypophysectomy similarly resulted in a gradual decline in their activities. Replacement of follicle-stimulating hormone or testosterone for 7 days from 30 days after hypophysectomy did not quantitatively restore either the testes weight or the activities of the proteins. The results indicate a primary distribution of both proteins in mature germ cells.