Anatomical descriptions of the genitofemoral nerve (GFn) innervating the lower pelvic area are contradictory. Here we re-examine its origin and innervation by its various branches of principal target organs in the male rat. Using gross dissection, electrophysiological techniques and retrograde tracing of motoneurones with horseradish peroxidase, we confirm that the GFn originates from lumbar spinal nerves 1 and 2, and that at the level of the common iliac artery it divides into a lateral femoral and a medial genital branch. In contrast to previous studies, we report that the genital and not the femoral branch innervates the abdominal-inguinal skin, and not only the genital but also the femoral branch innervates the cremaster muscle (Cm) surrounding the testes. Motoneurones innervating the Cm proper are located in the ventral nucleus of L1 and L2, and those innervating the muscular transition region of the rostral Cm are located in the ventral nucleus in L1 and the ventrolateral nucleus in L2. The GFn may contribute to male reproductive performance by transmitting cutaneous information during copulation and, via contraction of the Cm to promote ejaculation, the protective displacement of the testes into the abdominal cavity during fighting and as a sperm-protecting thermoregulatory measure.