Introduction: Sleep deprivation (SD) impairs neurocognitive functions. Assuming that this effect is mediated by reduced cerebral glucose supply due to prolonged wakefulness inducing a progressive depletion of cerebral glycogen stores, we hypothesized that short-term sleep loss amplifies the deteriorating effects of acute hypoglycemia on neurocognitive functions. Methods: Seven healthy men were tested in a randomized and balanced order on 3 different conditions spaced 2 weeks apart. After a night of total SD (total SD), 4.5. h of sleep (partial SD) and a night with 7. h of regular sleep (regular sleep), subjects were exposed to a stepwise hypoglycemic clamp experiment. Reaction time (RT) and auditory evoked brain potentials (AEP) were assessed during a euglycemic baseline period and at the end of the clamp (blood glucose at 2.5. mmol/l). Results: During the euglycemic baseline, amplitude of the P3 component of the AEP was lower after total SD than after partial SD (9.2 ± 3.2. μV vs. 16.6 ± 2.9μV; t(6)= 3.2, P=0.02) and regular sleep (20.2±2.1μV; t(6)=18.8, P<0.01). Reaction time was longer after total SD in comparison to partial SD (367 ± 45 ms vs. 304 ± 36. ms; t(6) = 2.7, P=0.04) and to regular sleep (322 ± 36 ms; t(6) = 2.41, P=0.06) while there was no difference between partial SD and regular sleep condition (t(6) = 0.60, P=0.57). Hypoglycemia decreased P3 amplitude by 11.2 ± 4.1 μV in the partial SD condition (t(6) = 2.72, P=0.04) and by 9.3 ± 0.7 μV in the regular sleep condition (t(6) = 12.51, P<0.01), but did not further reduce P3 amplitude after total SD (1.8 ± 3.9μV; t(6)= 0.46, P=0.66). Thus, at the end of hypoglycemia P3 amplitudes were similar across the 3 conditions (F(2,10) = 0.89, P=0.42). RT generally showed a similar pattern with a significant prolongation due to hypoglycemia after partial SD (+42 ± 12 ms; t(6)= 3.39, P=0.02) and regular sleep (+37 ± 10 ms; t(6)=3.53, P=0.01), but not after total SD (+15 ± 16; t(6)=0.97, P=0.37), resulting in similar values at the end of hypoglycemia (F(1,6)=1.01, P=0.36). Conclusions: One night of total SD deteriorates neurocognitive function as reflected by indicators of attentive stimulus processing, but does not synergistically aggravate the impairing influence of acute hypoglycemia. The findings are not consistent with the view that neurocognitive deteriorations after SD result from challenged cerebral glucose metabolism.