Metabolic characterization of undifferentiated and differentiated PAZ6 cells. (a) Expression of mitochondrial respiratory complexes I, II, III, IV and V in PAZ6 pre-adipocytes and adipocytes. Differentiation is associated with an overall increase in the expression of mitochondrial electron transfer chain complexes. Treatment of PAZ6 adipocytes with retinoic acid (1 μM), T3 (2nM) and Forskolin (2 μM)/IBMX (125 μM) further increases the expression of complexes II, III and IV. Values are presented as the fluorescent units ratio between each respiratory complex protein and the nuclear encoded NNT protein ± SEM. (b) OCR of PAZ6 pre-adipocytes and adipocytes in unbuffered DMEM containing 5 mM pyruvate and 2.5 mM glucose. (c) Basal, uncoupled and maximal respiratory capacities are robustly increased in the adipocyte relative to the pre-adipocyte state (* p < 0.0001). (d) OCR in PAZ6 adipocytes under various treatments (24 hrs).. FCCP-dependent (State3u) respiration is mildly increased in adipocytes treated with retinoic-acid (RA) plus T3 (**p < 0.05 vs. adipocyte). Forskolin/IBMX had a synergistic effect upon State3u respiration when given in combination with RA (***p < 0.0001 vs. adipocyte; # p < 0.05 vs. adipocyte + RA). (e) Basal, uncoupled and maximal respiration were calculated. Maximal respiratory capacity is increased in adipocytes treated with RA plus T3 (*p < 0.05 vs. adipocyte) and RA plus T3 plus forskolin/IBMX (**p < 0.05 vs. adipocyte). (f) Glycolysis, shown as the rate of extracellular acidification (ECAR) in PAZ6 pre-adipocytes and adipocytes under several conditions. Rates are significantly increased in adipocytes treated with RA, RA plus T3 (**p < 0.01 vs. adipocyte) and RA plus T3 plus forskolin/IBMX (***p < 0.001 vs. adipocyte). Results expressed as mean ± SEM.