c-fos expression and NADPH-d reactivity in spinal neurons after fatiguing stimulation of hindlimb muscles in the rat

The distribution of Fos-immunoreactive (Fos-ir) and nicotinamide adenine dinucleotide phosphate-diaphorase (NADPH-d)-reactive neurons in the rat lumbar spinal cord was examined following muscle fatigue caused by intermittent high-rate (100 s^-1) electrical stimulation of the triceps surae muscle or the ventral root L5 (VRL5) for 30 min. Following both types of stimulation, the fatigue-related c-fos gene expression was more extensive in the L2-L5 segments on the stimulated side, and the majority of Fos-ir neurons were concentrated in the dorsal horn. After direct muscle stimulation, the highest number of Fos-ir neurons were detected in two regions: layer 5, and superficial layers (1 and 2_o), although many labeled cells were also found in layers 3, 4, 6, and 7. In response to VRL5 stimulation, the maximal density of Fos-ir neurons was detected in the middle and lateral parts of layers 1 and 2_o, the zone of termination of high-threshold muscle afferents. Statistically significant prevalence of Fos-ir cell number was also found in layers 5 and 7 on the stimulated side. A few Fos-ir neurons were detected in the ventral horn (layer 8 and area 10) on both sides. The lamellar distribution of NADPH-d-reactive neurons was similar over all experimental groups of animals. In the L3-L6 segments, such reactive cells were arranged in two distinct regions: dorsal horn (layers 2_i, 3, and 5) and area 10; in the L1 and L2 segments, an additional cluster of NADPH-d positive cells was found in the intermediolateral cell column (IML). Double-labeled cells were not detected. We suggest that c-fos expression in response to muscle fatigue reveals activity of functionally different types of spinal neurons which could operate together with NOS-containing cells in pre-motoneuronal networks to modulate the motoneuron output.