Fig. 5: Two-photon imaging of l-lactate on astrocytes in acute brain slices.

a Schematic illustration of AAV injection into the somatosensory cortex for the brain slice experiments. ITR inverted terminal repeat, GFAP human glial fibrillary acidic protein promoter, WPRE woodchuck hepatitis virus posttranslational regulatory element, pA human growth hormone polyA signal. b Representative two-photon images of eLACCO1.1 expressed on astrocytes in brain slice before and after 10 mM l-lactate stimulation. Parallel experiments with deLACCO produced baseline fluorescence that was not significantly higher than background and did not produce a significant change in fluorescence relative to baseline (ΔF/F = 0.85 ± 1.6, P = 0.25, two-tailed paired t-test). Similar results were observed from more than 3 slices. Scale bar represents 50 μm. c Fluorescence traces of eLACCO1.1-expressing astrocytes in response to bath application of l-lactate (mean ± s.e.m.). 1 mM l-lactate (n = 5 slices from 4 mice), 2.5 mM l-lactate (n = 5 slices from 4 mice), or 10 mM l-lactate (n = 6 slices from 4 mice). d ΔF/F plots for eLACCO1.1 at each dose of l-lactate. ΔF/F was calculated by: ΔF/F = ((F_x − F_b)/F_b), where F_x is the peak fluorescence intensity and F_b is the baseline fluorescence intensity. 1 mM l-lactate (n = 5 slices from 4 mice, P = 0.01, two-tailed paired t-test), 2.5 mM l-lactate (n = 6 slices from 4 mice, P = 0.02, two-tailed paired t-test), or 10 mM l-lactate (n = 6 slices from 4 mice, P = 0.01, two-tailed paired t-test). The horizontal line is the median; the top and bottom horizontal lines are the 25th and 75th percentiles for the data; and the whiskers extend one standard deviation range from the mean represented as black filled circle. One slice data in the 2.5 mM group was omitted from the trace in c because it was not a complete time course, but ΔF/F of the slice could be measured and was included in d. Source data of c, d are provided as a Source Data file.