CGRP incubation for 3 h does not cause KCC2 internalization in superficial dorsal horn neurons in either female or male mice spinal cord explants. A, Representative confocal images of the dorsal horn of spinal cord explants from female and male mice incubated with either 50 nm CGRP or vehicle for 3 h, showing CGRP, IB4, KCC2, and NeuN staining. Scale bar, 20 μm. B, Average KCC2 intensity profiles from dorsal horn neurons of male (left; n = 8 mice) and female (right; n = 7 mice) mice explants incubated with vehicle (orange and violet lines) or 50 nm CGRP (red and purple lines). Results are presented as the mean per group. C, Membrane KCC2 intensity of the four experimental groups calculated at position zero. No significant differences were observed between vehicle- and CGRP-treated female or male mice spinal cord explants.