Fig 2: Female Pirt deficient mice have an impaired glucose tolerance. Blood glucose traces after an intraperitoneal glucose tolerance test in female chow-fed (A) and high-fat, high-sugar (HFHS) diet-fed (B) and male chow-fed (D) and HFHS diet-fed (E) wild-type (WT) (black) and Pirt−/− (KO) (grey) mice with the corresponding area under the curve (AUC) (C,F) at 26 weeks of age. Phenotyping cohorts with n = 8 per sex, diet, and genotype. Data represent mean ± s.e.m. Data presented in line graphs (A,B,D,E) were analyzed by two-way ANOVA (genotype and time) and data presented in bar graphs (C,F) were analyzed by two-way ANOVA (genotype and diet) comparing WT with Pirt−/− mice. ANOVA was followed by Bonferroni post hoc multiple comparison analysis to determine statistical significance. *P < 0.05, **P < 0.01 effects of genotype within the diets and $$P < 0.01 main effects of genotype irrespective of diet.

Fig 3: Pirt is expressed in hypothalamic nuclei. Pirt gene expression, displayed as cycle of threshold (CT), in the heart muscle, pituitary gland, hypothalamus, quadriceps muscle, brown adipose tissue (BAT), inguinal white adipose tissue (iWAT), liver, and epididymal white adipose tissue (eWAT) of wild-type (WT) mice (n = 6) (A). Immunohistochemistry in the region of the arcuate nucleus (ARH), the median eminence (ME), and part of the ventromedial hypothalamic nucleus (VMH) of brain slices from WT and Pirt−/− (KO) mice (B). Pirt gene expression in the hypothalamus of WT mice compared to Pirt−/− mice (n = 5 per genotype) (C). Scale bars in (B) are 100 μm. Data represent mean ± s.e.m. ***P < 0.001 determined by an unpaired two-tailed Student's t-test comparing WT with Pirt−/− mice. 3V: Third ventricle.

Fig 4: Pharmacological activation of TRPM8 with icilin induces energy expenditure in wild-type and Pirt−/− mice. Energy expenditure of female chow-fed wild-type (WT) (A) and Pirt−/− (KO) mice (B) after subcutaneous injections of icilin (2 μmol/kg) (grey) or saline control (black) (n = 8 per genotype) with respective area under the curve (AUC) after icilin and saline injections (0–90 min) (C) and AUC of locomotor activity in the same period (D). Data represent mean ± s.e.m. *P < 0.05, **P < 0.01 determined by an unpaired two-tailed Student's t-test comparing saline and icilin injections within genotype.