Figure 3
Single-cell 2-photon Cl-imaging in Adeno-clomeleon-GFP virus infected mouse sensorimotor cortical slice. A. Sample image of neuron with Adeno-clomeleon-GFP virus infection. B1: Two panels on right are color-coded images showing [Cl-]i in an upper layer pyramidal neuron. Note that [Cl-]i is greater in the image acquired after L6/WM stimulation (E. Stim.) than in image acquired before L6/WM stimulation (Baseline). Each color-coded image is average of 15 consecutively acquired images. B2: Same format and neuron as in B1, but in this case the images were obtained in the presence of GABAA receptor antagonist SR95531 (gabazine). In the presence of gabazine, L6/WM stimulation was associated with little or no increase of this neuron’s [Cl-]i. C: Time course of alteration of neuronal [Cl-]i (Δ[Cl-]i) associated with L6/WM stimulation. Data points were obtained from images of same neuron before (black plot) and after exposure to ACSF containing the GABA antagonist gabazine (red plot). Error bars indicate ±1SE. D: Calibration curve of Cl- for FRET signals. Data points were obtained from standard solution containing 0, 50, 100, and 150 mM chloride respectively. E: Bar with black color (E. Stim.) shows average across-neuron [Cl-]i (n = 6) measured subsequent to L6/WM stimulation; bar on right (E. Stim. + SR95531; red color, n = 6) shows average across-neuron [Cl-]i measured subsequent to L6/WM stimulation in the presence of the GABAA antagonist SR95531. All three bars are normalized by [Cl-]i before stimulation. Each neuron was located in a different slice. ** indicates p < 0.001; * indicates p < 0.05; NS – not significant.