Figure 1.

Chemogenetic Activation of mRGCs

(A and B) After intravitreal injection of a viral vector (AAV2-hSyn-DIO-hM3Dq-mCherry) to Opn4^Cre:Z/EGFP mice, immunohistochemical staining revealed transgene (mCherry; red) expression in GFP-positive neurons (green) in an en face view of retinal whole mounts (A) and retinal section (B). The retinal section shows the expression of transgene in cells of the retinal ganglion and inner nuclear cell layers (GCL and INL) of hM3Dq Opn4^Cre/+ mice. Notice the different soma sizes of transduced cells (arrows), with DAPI stain in blue. A monochrome version of mCherry staining and more details on hM3Dq expression are provided in Figure S1.

(C) Representative images of eyes under infrared illumination from hM3Dq-expressing mice held in darkness, prior to (top) and at 20, 120, and 180 min after intraperitoneal (i.p.) injection of CNO (5 mg/kg). For more details, see Figure S2.

(D) Representative double-plotted actogram of wheel running activity of a hM3Dq mouse housed under a light:dark cycle (the light phase is indicated in yellow) until day 12 followed by constant darkness. The star shows the start of a 2 hr presentation of 0.25 mg/mL CNO in drinking water; the red line to left represents the eye fit through activity onsets. A representative double-plotted actogram of a control mouse is shown in Figure S3.

(E) Change in circadian phase CNO application to hM3Dq-expressing (open bars; n = 4) and control (filled bars; n = 5) mice (means ± SEM; Mann-Whitney U test, p = 0.015).

(F) Representative micrographs of coronal section through the SCN labeled for c-Fos (dark) from hM3Dq and control mice after CNO administration (5 mg/kg, i.p., at CT14).

(G) Mean (±SEM) number of c-Fos positive cells mm⁻² in SCN sections from hM3Dq (n = 6) and control (n = 6) mice (two-tailed unpaired t test,^∗∗p < 0.01).