Habitual, moderate exercise is associated with improved health, including reductions in illness. These benefits may stem, in part, from immune function improvements. We have previously reported that daily wheel running increases serum and peritoneal natural IgM (nIgM) in pathogen-free Sprague-Dawely rats. B-1 cells, which primarily reside in the peritoneal cavity, produce nIgM in the absence of antigen stimulation. This study examined whether physical activity would also increase B-1 cell numbers in the peritoneal cavity, mesenteric lymph nodes (MLN) and spleen. Male, pathogen-free Fischer-344 rats were sedentary (standard cages) or physically active (running wheel access) for 6-7 weeks. Peritoneal cavity, MLN and spleen cells were taken, and the number of CD5+/CD11b+ (B-1) cells were measured using two-color flow cytometry. The results were that physically active animals had increased numbers of CD5+/CD11b+ cells in the peritoneal cavity. In addition, physically active animals had increased serum and peritoneal nIgM, thus replicating our previous observations. These results indicate that voluntary running selectively increases the B-1 cell population, which is most likely responsible for the elevated serum and peritoneal nIgM in active rats. As B-1 cells are important in host defense, these changes may contribute to the health benefits of exercise.