(A) Representative image of the DMH with Prdm13 (yellow) and Pmch (green) visualized by RNAscope. Cells were counterstained with DAPI (blue). White box shows the DMH, which is divided into medial and lateral areas by dashed line. White arrow shows yellow+ and green+ cells. Scale bar indicates 100 μm. (B) Number of Cck+ or Grp+ cells within Prdm13+ cells in medial, lateral, or total DMH (n = 3–5). Values are shown as means ± S.E., **P < 0.01 by unpaired t test. (C) Expression of Cck mRNA in the hypothalamus of young and old mice (n = 3–4). Values are shown as means ± S.E., *P < 0.05 by unpaired t test. (D) Semiquantitative evaluation of Cck expression levels in Prdm13+Cck+ cells in young or old mice during SD-Cont. Signal dots derived from Cck mRNA were counted manually and categorized into four grades: 1 (1–5 dots/cell), 2 (6–10 dots/cell), 3 (11–15 dots/cell), and 4 (>16 dots/cell). Numbers of cells divided into four grades of Cck expression were shown as means ± S.E., listed P-values, and non-significant (ns) by two-way ANOVA with Bonferroni’s post hoc test.