We report in the present study that inhibition of endogenous NO formation does not alter glucose uptake of human skeletal muscle at rest or during low intensity exercise. However, by affecting the release and uptake of free fatty acids, NO appears to contribute to the regulation of muscle energy metabolism, at least when the muscle is at rest.
The effect of nitric oxide on muscle glucose uptake at rest and during exercise
In the present study we show that glucose uptake is unaffected by prior NOS inhibition with L-NMMA both at rest and during exercise. Previous human studies that have addressed the role of nitric oxide for glucose uptake in skeletal muscle have shown discrepant findings where some have shown that glucose uptake during exercise is reduced when NOS is inhibited [8, 11], whereas others have shown no effect . The discrepancy between findings in the different studies is unclear, however, differences in experimental conditions between the studies could in part explain the findings. Firstly, McConell and co-workers  began infusion of L-NMMA ten minutes after steady state exercise, while we began infusion ten minutes before exercise. Copp and colleagues showed that the timing of NOS inhibition does have an effect on the blood flow response in relation to muscle fibre type in rats , but it may also affect glucose uptake. Another experimental difference lies in the mode and intensity of exercise. In our study single leg exercise at ~10 watts with a substantial isometric component, was used whereas in the study by Bradley and co-authors the exercise consisted of two leg cycling at the 60% of peak VO2max, corresponding to ~142 watts . Consequently, glucose uptake was increased 30-fold ~ in the study by Bradley et al. whereas we observed a ~10-fold increase in our study. The approach and results of Kingwell et al. was similar to Bradley et al. [8, 11]. Thus, our present findings combined with that of others  suggest that glucose uptake at rest and during low-to-moderate exercise intensity is not NO mediated, but NO affects glucose uptake during higher exercise intensities [8, 11]. Many previous animal studies that have addressed the effect of NO on glucose uptake in muscle have also resulted in contrasting conclusions [7, 9, 10, 13, 18], whereas most studies on cardiac muscle have all reported increased glucose uptake in parallel with increased carbohydrate metabolism during the inhibition of NOS [12, 20–22]. Thus, overall, the role of NO for glucose uptake appears to be more important in cardiac than skeletal muscle.
The effect of nitric oxide on free fatty acid exchange in muscle
In line with the results of Rottman et al. in mice , the present study demonstrates that NO inhibition alters the exchange of FFAs over the limb. At rest in the control condition, there was a release of FFA from the limb, whereas during NOS inhibition an uptake of FFA was detected (Figure 1). This finding, combined with the unaltered glucose uptake during NOS blockade, is also in line with the observation that oxygen consumption of the muscle is increased during NOS inhibition, and suggests that the overall metabolism, as indicated by the change in oxygen consumption, of the muscle was enhanced during NOS inhibition. The finding suggests that nitric oxide suppresses fatty acid metabolism in resting human skeletal muscle, which is in accordance with findings in vitro and animal studies showing that NOS inhibition increases FFA oxidation .
Whether the observed increase in FFA utilization also resulted in increased FFA oxidation is unclear as this was not determined in the present study. Nevertheless, the increase in oxygen consumption during the NOS blockade could indicate that there was an increased FFA oxidation. Many animal [36, 37] and human  studies indicate that increased rates of FFA oxidation lowers the efficiency of the muscle. However, a switch to exclusive use of fatty acids as an energy source is calculated to impair the efficiency of ATP production by only 10–15% . Our results do not suggest a complete substrate switch and, thus, the increased FFA uptake and utilization cannot fully explain the observed increase in oxygen consumption. Therefore, an additional explanation for the increase in oxygen uptake during NOS blockade probably was a reduced inhibitory influence of NO on mitochondrial respiration [40–42]. Alternatively, utilization of FFAs may have actually been enhanced secondarily to this phenomenon to fulfil increased cellular metabolism, but this possibility warrants further investigation. Finally, it has been observed that the inhibition of NOS leads to enhanced lipolysis in subcutaneous adipose tissue [43, 44]. In our study arterial FFA levels appeared to be somewhat increased both at rest and during exercise, but this did not reach statistical significance, which points to the conclusion that indeed shift in the utilization rather than increased availability accounted for the observed increase in FFA uptake.
Muscle biopsies were not obtained in the current study so direct measurements of the degree of NOS inhibition could not be obtained. The dose of L-NMMA used in the present study was similar to that used in a number of studies from our own as well as other laboratories. These previous studies, e.g. Rådegran and Saltin 1999, have shown that resting blood flow as well as the responses to acethylcoline infusions are approximately halved with use of this L-NMMA dose, indicating an effective inhibition . In the current study, resting blood flow during L-NMMA infusion was reduced to a similar extent as previously observed. Moreover, it has been previously demonstrated that infusion of the NOS blocker L-NAME that reduces limb blood flow to a similar extent as the L-NMMA dose used in the current study, reduces NO synthase activity by approximately 70% . Hence, it appears likely that the extent of NOS inhibition was similar in the present as in previous studies on humans [45, 46]. Finally, as a vehicle control group was not applied in the present study, it is not possible to completely eliminate the possibility that there may have been a carry over effect during the second bout of exercise with NOS inhibition.
In conclusion, endogenous nitric oxide does not appear to change glucose uptake of human skeletal muscle at rest or during low intensity exercise, but shifts a release of free fatty acids to uptake, thereby altering muscle energy metabolism, in particular at rest.