Fig. 3.

PDE3 and PDE4 inhibitors increase Ucp1 and Pgc-1α mRNA expression in differentiated brown adipocytes. UCP1 mRNA expression was measured in differentiated brown adipocytes that were (A) pretreated with 10 μM cilostamide, 10 μM rolipram, both, or 200 μM IBMX for 30 minutes, then stimulated by vehicle or 1 nM isoproterenol for 4 hours (n = 3–13); (B) treated with 300 nM cilostamide or 10 μM cilostamide with or without 10 μM rolipram (n = 3–8); or (C) pretreated with either 200 nM PF-04957325, 200 μM IBMX, or both for 30 minutes, then stimulated by vehicle or 1 nM isoproterenol for 4 hours (n = 3). UCP1 mRNA was quantified relative to 18S mRNA with use of RT-PCR as described in Materials and Methods. (D) UCP1 mRNA expression was measured from the RNA extracted from the interscapular BAT pads harvested from mice undergoing PET scanning after the procedure was terminated (Materials and Methods). UCP1 mRNA was quantified relative to GAPDH (glyceraldehyde 3-phosphate dehydrogenase) mRNA with use of real-time (RT)-PCR as described in Materials and Methods. Data are presented as mean fold over vehicle control ± S.E.M., and statistical analyses were performed using one-way analysis of variance with Dunnet post hoc test: ***P < 0.001; *P < 0.05, versus vehicle in each group.