Fig. 5
Oxt neurons in the PVH respond to FGF21. a Photomicrographs depict coronal PVH sections from wild-type C57BL/6 mice. Sections were subjected to in situ hybridization to identify β-Klotho mRNA (Klb, blue dots). Then, sections were immunostained to identify Oxt protein (brown) expression. Green arrows indicate Klb mRNA staining in Oxt neurons. b Photomicrographs depict coronal PVH sections from wild-type C57BL/6 mice injected with murine FGF21 (mFGF21) (1 mg/kg, IP) or water (vehicle). The sections were immunostained for c-Fos (black) and Oxt (brown). Blue arrows indicate Oxt (−) c-Fos (+) neurons; green arrows indicate c-Fos (+) Oxt (+) neurons. c The percentage of c-Fos (+) cells among PVH Oxt neurons after an IP injection of vehicle or mFGF21 (n = 4 per group). d, e A representative [Ca2+]i result, expressed by fura-2 fluorescence ratio (F340/F380), in an ex vivo single-cell Oxt-neuron. Addition of 50 ng/ml mFGF21 for 5 min to perfusate (KRB containing 10 mM glucose) increased [Ca2+]i in a single PVH neuron (e), which was subsequently shown to be IF to Oxt by immunocytochemical staining (d). f, g Percentage of mFGF21-responding Oxt neurons among PVH Oxt neurons (f) and mFGF21-responding PVH neurons (g). The number above the bar indicates the number of neurons that responded over number examined. The length of scale bar is 200 μm for the low magnification panel and 50 μm for the high magnification panel in a; 50 μm in b; and 20 μm in d. Data are shown as box and whisker plots (centre line, median; box limits, upper and lower quartiles; whiskers, the minimum and maximum value of a data set). *p < 0.05. See also Supplementary Fig. 5
