Figure 2. Pharmacological analysis of O2 use on different subcellular mechanisms. A, Schematic showing experimental protocol, with three stimulation sets in each condition applied at 6 min intervals (see Materials and Methods): during the first stimulation phase, no drug is present for all slices. In the second phase, some slices are exposed to drug, while some remain drug free as controls. In the third phase, 1 μm TTX is applied (to allow subtraction of the stimulation artifact from the field potential data). Responses in phase 2 are reported as a percentage of the response in phase 1, to normalize to the number of axons activated in each slice. Comparing the normalized responses for control and drug in phase 2 defines the effect of the drug tested. B–D, Amplitude of presynaptic action potential volley (B), postsynaptic field EPSP (C), and postsynaptic action potential population spike (D) measured as in Figure 1B, in the presence of NBQX+AP5 or Cd2+ together with the amplitude seen on the second application of 20 Hz stimulation in the absence of drugs (Con). The p values are from one sample t tests comparing with 100% (the response in the first stimulation phase), unless marked with an asterisk, in which case the comparison is versus zero. E–G, Amplitude of the [O2] decrease (E), decrease in NADH fluorescence (F), and overshoot of NADH fluorescence above its baseline value (G) in the presence of NBQX+AP5, Cd2+, and TTX, compared to the amplitude seen on the second application of 20 Hz stimulation in the absence of drugs (Con; these values are not significantly different from the responses to the first set of stimuli). Incrementally blocking postsynaptic activity (NBQX+AP5), transmitter release (Cd2+), and all evoked activity (TTX) incrementally decreased the [O2] decrease. The p values are from unpaired t tests, corrected for multiple comparisons. H, I, Specimen traces (H) and mean values (I) for the decrease of [O2] evoked by 500 μm glutamate in 250 μm Cd2+ and in Cd2+ + 1 μm TTX. The resting [O2] was not significantly affected by Cd2+ (reduced by 42 ± 36%; p = 0.29) nor by TTX (in Cd2+; reduced by 12 ± 36%; p = 0.75) in seven slices.