We have recently reported that during moderate intensity exercise, low muscle glycogen concentration and utilization caused by a high-fat diet is associated with a marked increase in fat oxidation with no effect on plasma glucose uptake (Rd glucose). It is our hypothesis that this increase in fat oxidation compensates for low muscle glycogen, thus preventing an increase in Rd glucose. Therefore, the purpose of this study was to determine if low muscle glycogen availability increases Rd glucose under conditions of impaired fat oxidation. Six cyclists exercised at 50% VO_2peak for 1 hour after 2 days on either a High-fat (60% fat/24% carbohydrate, HF) or Control (22% fat/65% carbohydrate, CON) diet to manipulate muscle glycogen to low and normal levels, respectively. Two hours before the start of exercise subjects ingested 80 mg of propanolol (B), a non-selective -adrenergic receptor blocker, to impair fat oxidation during exercise. HF significantly decreased calculated muscle glycogen oxidation (p<0.05) and this decrease was partly compensated for by an increase in fat oxidation (p<0.05) accompanied by an increase in whole body lipolysis (p<0.05), despite the presence of B. Although HF increased fat oxidation, plasma glucose appearance rate, Rd glucose, and glucose clearance rate were also significantly increased by 13, 15, and 26%, respectively (all p<0.05). In conclusion, when lipolysis and fat oxidation are impaired, in this case by B, fat oxidation cannot completely compensate for a reduction in muscle glycogen utilization and consequently plasma glucose turnover increases. These findings suggest that there is a hierarchy of substrate compensation for reduced muscle glycogen availability following a high-fat and low-carbohydrate diet, with fat being the primary and plasma glucose the secondary compensatory substrate. This apparent hierarchy likely serves to protect against hypoglycemia when endogenous glucose availability is low.