We have investigated the form I cyclic nucleotide phosphodiesterase (PDE) fromDrosophila melanogasterand shown that whereas heads and male thoraces and abdomens contain high levels of Ca2 +-stimulated enzyme, female thoraces and abdomens contain little Ca2 +-stimulated activity. The electrophoretic patterns of form I PDE from these 3 sources have also been studied and reveal that heads, and male thoraces and abdomens, produce two bands of form I PDE both of which are stimulated by Ca2+. Extracts of female thoraces and abdomens, on the other hand, show only a single, faster running band of PDE activity which is only marginally stimulated by Ca2+, if at all. Surveying wild-type strains ofDrosophilahas revealed that one strain, Swedish, shows altered electrophoretic mobility of the PDE band from female thoraces and abdomens. The alteration is such that the Swedish PDE band runs more anodally than the Oregon-R and Canton-S PDE activities. Mixing experiments, using co-homogenization of heads with female thoraces and abdomens, yeild a single faster running band on electrophoresis. This band contains only Ca2+-insensitive PDE. Attempts to reconstruct this loss of Ca2+-sensitive PDE without electrophoresis have failed. The Swedish electrophoretic variation of the PDE from female thoraces and abdomens has been found to be recessive with respect to the Canton-S phenotype, but the variation is observed to re-emerge and segregate with the third chromosome in the F2generation. The results indicate that electrophoretic variation in the form I PDE is, by itself, insufficient to allow the location of the structural gene for this enzyme.