Figure 2.
KATP channel activation after glycolytic inhibition is conditional on the perfusion flow rate and the presence of mitochondrial fuels. A, The spontaneous firing rate of SNr neurons, recorded in the continuous presence of the KATP channel blocker Glib (200 nm), was significantly decreased (circle symbols; 28.8 ± 4.2 vs 19.3 ± 2.5 spikes/s; n = 8; p = 0.001, Student's paired t test) after inhibition of glycolysis with 2-DG (5 mm) in the absence of glucose but in the presence of βHB (2.5 mm). The decreased firing rate after inhibition of glycolysis with IAA (1 mm) in the presence of βHB (2.5 mm) was not reversed after addition of Glib (10 μm; square symbols; 18.2 ± 1.7 vs 17.0 ± 1.8 spikes/s; n = 6; p > 0.05, one-way ANOVA with Bonferroni's test). IAA (1 mm) significantly decreased the firing rate of Kir6.2 KO SNr neurons (triangle symbols; 33.3 ± 3.6 vs 15.7 ± 1.6 spikes/s; n = 8; p = 0.0003, Student's paired t test). B, With a flow rate of 5 ml/min, βHB (2.5 mm) sustained the spontaneous firing of an SNr neuron after inhibition of glycolysis with IAA (1 mm). Further addition of the KATP channel blocker Glib (10 μm) did not reverse the decrease in firing rate. a–c, Traces depict cell-attached recordings of spontaneous firing at the indicated times (calibration: 50 pA, 200 ms). C, With a lower flow rate of 1 ml/min, βHB (2.5 mm) was unable to sustain the firing rate of an SNr neuron. Addition of Glib (200 nm) could partially restore the firing rate. a–c, Traces depict cell-attached recordings of spontaneous firing at the indicated times (calibration: 20 pA, 200 ms). D, Cell-attached recordings of spontaneous firing rates with inhibition of glycolysis by IAA (1 mm) in the presence of glucose (10 mm). IAA completely silenced SNr firing of control neurons (n = 6; black trace). When KATP channels were inhibited using Glib (200 nm; 10 min preincubation; n = 4; blue trace) or eliminated in Kir6.2 KO mice (n = 6; red trace), SNr firing displayed a transient increase, followed by a complete silencing. E, Representative whole-cell recordings showing the time course of the effect of IAA (1 mm) application on the normalized firing rate of SNr neurons in the presence of glucose. In a control neuron, application of IAA promptly decreased the spontaneous firing rate without any increase in firing rate (black line). In a neuron preincubated in Glib (200 nm; blue line) or in a neuron from a Kir6.2 KO mouse (red line), the firing rate increased after application of IAA and then stopped firing. F, Summarized data from all whole-cell experiments with application of IAA in the presence of glucose. After application of IAA, control neurons had a hyperpolarized resting potential (−74.6 ± 3.5 mV; n = 9; black symbols). After IAA, neurons preincubated in Glib (200 nm; blue symbols) had more depolarized resting potentials (−60.5 ± 2.7 mV; n = 6; p < 0.05, one-way ANOVA with Bonferroni's test), and neurons from Kir6.2 KO animals (red symbols) also rested more depolarized (−53.2 ± 3.8 mV; n = 6; p < 0.05, one-way ANOVA with Bonferroni's test). All error bars indicate SEM; *p < 0.05. Ctrl, Control.