The optimization of multisystem neurorehabilitation protocols including electrical spinal cord stimulation and multi-directional tasks training require understanding of underlying circuits mechanisms and distribution of the neuronal network over the spinal cord. In this study we compared the locomotor activity during forward and backward stepping in eighteen adult decerebrated cats. Interneuronal spinal networks responsible for forward and backward stepping were visualized using the C-Fos technique. A bi-modal rostrocaudal distribution of C-Fos-immunopositive neurons over the lumbosacral spinal cord (peaks in the L4/L5 and L6/S1 segments) was revealed. These patterns were compared with motoneuronal pools using Vanderhorst and Holstege scheme; the location of the first peak was correspondent to the motoneurons of the hip flexors and knee extensors, an inter-peak drop was presumably attributed to the motoneurons controlling the adductor muscles. Both were better expressed in cats stepping forward and in parallel, electromyographic (EMG) activity of the hip flexor and knee extensors was higher, while EMG activity of the adductor was lower, during this locomotor mode. On the basis of the present data, which showed greater activity of the adductor muscles and the attributed interneuronal spinal network during backward stepping and according with data about greater demands on postural control systems during backward locomotion, we suppose that the locomotor networks for movements in opposite directions are at least partially different.