We investigated changes in renal hyaluronidase activity in streptozotocin (STZ)-induced diabetic rats during the progression of diabetes. Prior to the study, we characterized rat renal hyaluronidase activity to find that its optimum pH is 3.5 and that it consists of two isomers of 73 and 63 kDa, as detected by zymography. Hyaluronidase activity was traced in one whole kidney and in the cortex and medulla of the other kidney up to the 18th week after STZ injection. Whole kidney hyaluronidase activity started to increase on day 3 and reached a maximum level 2.4 times that of the controls in the 3rd week. Cortical hyaluronidase showed a similar tendency to that of whole kidney hyaluronidase, while medullary hyaluronidase activity continued to increase until the 8th week, suggesting their different involvements in the progression of diabetic nephropathy. In zymography, the intensities of the two isomer bands increased with the progression of diabetes, but the intensity ratio did not change significantly and no new isomer band appeared. Renal HAase activity increased only in STZ-induced diabetic rats, but not in spontaneously diabetic Goto-Kakizaki rats still without remarkable renal disorder. Based on these findings, increased renal HAase activity may serve as a useful marker for diabetic nephropathy.