Figure 3.

Thalamic p-AMPK expression and GIRK currents following AMPK activation. (A) Thalamocortical neuron AMPKAR expression based on construct developed by Tsou et al.⁶³ and Konagaya et al.⁶⁴ Scale bar = 50 µM. (B) Thalamocortical neuron AMPKAR FRET efficiency increased during application of metformin (10 mM, purple), A-769662 (100 nM, green). FRET values are normalized to the 3-min baseline period prior to drug application. (C) Histogram (left) of FRET values for all neurons during baseline (black) and metformin (purple) application. Pairwise comparison (right) of mean FRET per slice in baseline and metformin application. (D) Histogram of FRET values (left) and mean pairwise comparisons (right) for A-769662 application. See Supplementary Table 3 for data values. (E) Top left: Whole-cell patch-clamp recording in an acute thalamic brain slice. Baclofen application pipette is on the right. Top right: Schematic of experiment. Whole-cell patch-clamp recordings were performed using recording pipettes containing either control internal solution (black) or internal solution supplemented with an AMPK activator (red). A second pipette, placed proximal to the patched neuron, contained 100 µM baclofen (blue). Bottom: Representative traces of evoked GABA_B currents with recording pipette containing AMPK activator (grey, t = 3 min; red, t = 9 min). Arrows point to capacitive transient of a voltage-step used to measure access resistance and puff onset. (F) AMP (1 mM) stabilized, but did not significantly mitigate (P = 0.06), GABA_B-receptor current rundown. (G) Metformin (1 mM) significantly prevented GABA_B current rundown relative to control. (H) A-769662 (100 nM) similarly attenuated GABA_B current rundown relative to control. For F–H, response amplitude of GABA_B-receptor mediated K⁺ current was normalized to the current amplitude at 3 min (t = 1); data are mean ± SE; *P < 0.05. Repeated-measures two-way ANOVA; unpaired t-test for pairwise comparison. See Supplementary Table 3 for details.