Pituitary adenylate cyclase activating polypeptide (PACAP) a neuropeptide recently isolated from hypothalamus, exists in two biological active forms: PACAP38 and PACAP27. The PACAP precursor (preproPACAP) contains besides PACAP a 29 amino acid peptide designated PACAP related peptide (PRP) in its sequence. The hypothalamus has the highest concentration of PACAP, but PACAP mRNA and immunoreactivity are found in several other tissues including the testis. Here we demonstrate that extracts of testis contain all preproPACAP derived peptides the concentrations being: PACAP38 2.8 +/- 0.14 pmol/g wet weight, PACAP27 0.18 +/- 0.02 pmol/g and PRP 0.84 +/- 0.14 pmol/g, respectively. By in situ hybridization PACAP/PRP mRNA was located near the perimeter of the seminiferous tubules in early germ cells, and by immunocytochemistry PACAP immunoreactivity was shown in spermatids near the lumen, in spermatogonia and primary spermacytes, but not in mature spermatids or spermatozoa. The findings suggest that PACAP might be an autocrine regulator in developing germ cells during spermatogenesis.