Fig. 3: Optogenetic elevation of astrocytic [Cl⁻]_i shortens activation-induced neuronal [Ca²⁺]_i.

a The optogenetic Cl⁻ pump was expressed in astrocytes to manipulate [Cl⁻]_i, while astrocytic [Cl⁻]_i or neuronal [Ca²⁺]_i was imaged. As a negative control, light stimulation was applied to mice not expressing NpHR3.0. Mice were voluntarily running on a Styrofoam sphere. b Average astrocytic [Cl⁻]_i traces during transition from stationary to mobile, recorded during continuous light stimulation of NpHR3.0 in astrocytes with or without NpHR3.0; shading indicates ±SEM. N = 6 ctrl/5 NpHR3.0. c Peak amplitude of astrocytic [Cl⁻]_i upon movement onset; time to peak of astrocytic [Cl⁻]_i; time to baseline; slope of decay, and area under the curve (AUC) from 6 to 8 s, data represent mean ± SEM. N = 5 ctrl/5 NpHR3.0, un-paired two-tailed t-test. p = 0.9238, p = 0.6626, **p = 0.0068, **p = 0.0049 *p = 0.0119. d Average neuronal [Ca²⁺]_i trace during transition from stationary to mobile, recorded during continuous light stimulation of NpHR3.0 in astrocytes with or without NpHR3.0; shading indicates ±SEM. N = 3 mice. e Peak amplitude of neuronal [Ca²⁺]_i upon movement onset; time to peak; recovery time to baseline; slope of decay, data represent mean ± SEM. N = 6 ctrl/5 NpHR3.0, un-paired two-tailed t-test, p = 0.5665, p = 0.1167, **p = 0.0049, *p = 0.0384. f Using the same protocol as in Fig. 2, whiskers were stimulated using air puffs. g astrocytic [Cl⁻]_i trace during whisker stimulation, recorded during continuous light stimulation of NpHR3.0 in astrocytes with or without NpHR3.0; shading indicates ±SEM. N = 4 ctrl/5 NpHR3.0. h Peak amplitude of astrocytic [Cl⁻]_i upon whisker stimulation; time to peak; period of maximal changes, 6–8 s after onset of stimulation. AUC during 6–8 s after onset of stimulation, data represent mean ± SEM. N = 4 ctrl/5 NpHR3.0, un-paired two-tailed t-test, **p = 0.0030, **p = 0.0017, **p = 0.0071, *p = 0.022. i Average neuronal [Ca²⁺]_i trace during whisker stimulation, recorded during continuous light stimulation of NpHR3.0 in astrocytes with or without NpHR3.0; shading indicates ±SEM. N = 8 ctrl/10 NpHR3.0. j Peak amplitude of neuronal [Ca²⁺]_i upon whisker stimulation; time to peak; period of maximal astrocytic [Cl⁻]_i changes, 6–8 s after onset of stimulation. AUC of neuronal [Ca²⁺]_i during 5 s after peak, data represent mean ± SEM. N = 8 ctrl/10 NpHR3.0, un-paired two-tailed t-test, p = 0.061, p = 0.7249, *p = 0.0147, *p = 0.0155. [Cl⁻]_i = mClY − ΔF/F (%). Source data are provided as a Source Data file.