Altered substrate availability and muscle glycogen concentration may mediate exercise-induced gene transcription in skeletal muscle during and following exercise. To determine the influence of prolonged exercise and rapid glycogen repletion on regulation of peroxisome proliferator-activated receptor-gamma co-activator-1alpha (PGC-1α), a nuclear co-activator involved in coordinating metabolic gene expression and mitochondrial biogenesis, 7 male subjects (23+1.3 y, VO2max 48.4+0.8 mL kg-1 min-1) completed 2 trials consisting of exhaustive cycling exercise (∼2h) at 65% VO2max followed by ingestion of either a high-(HC) or low-carbohydrate (LC) diet for the ensuing 52 h recovery period. Biopsies were taken at rest, exhaustion, and at 2-, 24-, and 52-h of recovery. Glycogen content remained depressed throughout recovery in LC (P<0.05), while returning to resting levels by 24 h of recovery in HC. Exercise induced a 6.2-fold increase in PGC-1α mRNA (P<0.05) that returned to resting levels within 24 h of recovery. There was also a 23% increase in PGC-1α protein (P<0.05) at the end of exercise, and it remained elevated for at least 24 h (P<0.05). While there was no direct treatment effect with HC vs. LC for PGC-1α mRNA, there were trends (P=0.11 and 0.06 in exercise and recovery, respectively) between the changes in glycogen and PGC-1α protein. PGC-1α protein content is increased by a single bout of exercise and its up-regulation may be associated with changes in muscle glycogen stores.