Pantothenic acid (PaA) is a water-soluble vitamin required to sustain various physiological functions in animals. The physiological roles of PaA on testicular function, in particular, testicular endocrinology and sperm mortility, were investigated in rats. Male rats at 3 weeks of age were fed a PaA-free diet or a 0.0016% PaA diet (control) for 7 weeks. Total body weight, as well as the weights of the liver, kidney, pituitary, testis, epididymis, seminal vesicle and prostate; sperm motility; and the plasma concentrations of luteinizing hormone (LH), follicle-stimulating hormone (FSH), testosterone and corticosterone were measured in rats at 10 weeks of age. Body weight gain decreased from 5 weeks of age in rats fed the PaA-free diet compared with the control. The relative weights of the testes were significantly higher in the PaA-deficient group compared with the control group. Several parameters of sperm motility were significantly reduced in the PaA-deficient group compared with the control group. In addition, the plasma concentrations of testosterone and corticosterone were significantly lower in the PaA-deficient group compared with the control group, whereas the plasma concentrations of FSH and LH showed no change. These results clearly demonstrate that PaA is an essential factor in testicular endocrinology and sperm motility in male rats.