The effects of total sleep deprivation (SD) on the expression of the immediate-early gene NGFI-A were studied in the rat brain by in situ hybridization. Rats were manually sleep-deprived for 3, 6, 12 and 24 h starting at light onset (08:00 h) and for 12 h starting at dark onset (20:00 h). SD performed during the day induced a marked increase in NGFI-A mRNA levels with respect to sleep controls in many cerebrocortical areas and caudate-putamen, which was most evident after 6 h SD. A decrease was seen in hippocampus and thalamus, particularly after 12 h SD. Rats sleep-deprived for 12 h during the night showed an increase in NGFI-A expression in some cortical areas while rats sleep-deprived for 24 h showed few changes with respect to controls. The pattern of NGFI-A expression after forced wakefulness showed some differences from that observed after spontaneous wakefulness [M. Pompeiano, C. Cirelli and G. Tononi, Immediate early genes in spontaneous wakefulness and sleep: expression of c-fos and NGFI-A mRNA and protein, J. Sleep Res., 3 (1994) 80-96]. These observations are discussed with respect to the functional consequences of wakefulness in specific brain areas.