The objective of this study was to characterize proteinase activities expressed during development and maturation of the prostate gland and seminal vesicles of the rat by using gelatin- and casein-containing SDS polyacrylamide gel zymography. The prostatic complexes of 2- and 10- day-old animals and the individual lobes of the prostate (ventral, dorsolateral, and anterior [coagulating gland]) and the seminal vesicles of 15-day-old animals expressed prominent gelatinolytic activities of approximately 64, 71, and 76 kDa. These activities had properties of metalloproteinases; i.e., they were stimulated by Ca2+ and inhibited by EDTA and EGTA. They were greatly diminished by 52 days of age (immediately postpuberty) and were not detected in the dorsal lobe of the adult. Less active gelatinolytic proteinases with molecular masses of approximately 34 and 43 kDa were expressed in the developing prostatic complexes and individual lobes and seminal vesicles, but they were not detected in postpubertal animals. Weak gelatinolytic activities of 82, 85, and 89 kDa were found in the prostatic complexes; these activities were greatly diminished in all prostate lobes with sexual maturation but were expressed in the seminal vesicles at all ages. A large-molecular-mass Ca2+- independent proteinase of 130 kDa or greater was first detected in the dorsolateral prostate at 21 days of age. This activity was expressed in both the lateral and dorsal lobes of the adult but was greater in the lateral lobe. Proteinase activities of about 22 and 26 kDa that were not stimulated by Ca2+ were detected in the ventral prostate at 15 days of age by means of both gelatin and casein gels. These two proteinases were substantially increased in activity in sexually mature animals. Other caseinolytic activities of 67, 78, 87, and 90 kDa, which were only weakly detectable, were found at all ages. However, a stronger caseinolytic activity of 22 kDa was found in the dorsolateral prostate, anterior prostate/coagulating gland, and seminal vesicles of animals 15-28 days of age. These studies show that the 64-, 71-, and 76-kDa metalloproteinase activities were expressed during prostate development, whereas other proteinases, like the 22-, 26-, and 130- kDa Ca2+-independent activities were first found after cytodifferentiation of the epithelium. These findings support the contention that the metalloproteinases may participate in tissue organization changes of the developing gland while the Ca2+-independent proteinases are involved in the secretory function of the mature gland.