Distribution of tracers within the thoracic and sacral spine under awake and anesthetized conditions. (A) Representative pictures of tracer within the thoracic region of the spine after sacrifice at 15, 30, 60, and 90 min after P40D680 infusion into the right lateral ventricle of anesthetized (isoflurane and ket/med) and awake group mice. Scale bars, 1 mm. (B) Quantification of fluorescence intensity in the thoracic region at the different time points. (C) Representative pictures of tracer within the sacral region of the spine. Scale bars, 1 mm. (D) Quantification of the number of tracer-enhanced segments between vertebrae at the different time points in anesthetized and awake mice. (E) Quantification of fluorescence intensity in the sacral region at the different time points in anesthetized and awake mice. Data are mean ± SD and representative of two independent experiments (n = 5 each group, one-way ANOVA with Tukey post hoc test; *, P < 0.05; **, P < 0.01; ***, P < 0.001).