18-NAD internal standard. Results reported as normalized based on the area ratio of NAD to the spiked O18-NAD internal standard to correct the variable rate of degradation of NAD in various tissue matrixes. Crude lysates were then centrifuged at 14,000 x g for 15 minutes twice and the protein concentration was determined using BioRad Protein Assay Reagent. 30 g proteins were resolved by a 412% or 12% NUPAGE Bis-Tris precast gel electrophoresis and transferred to nitrocellulose membranes. Individual proteins PubMed ID:http://www.ncbi.nlm.nih.gov/pubmed/19725016 were detected with the specific antibodies and visualized on film using horseradish peroxidase-conjugated secondary antibodies and Western Lightning Enhanced Chemiluminescence. RNA Isolation and Gene Expression Total RNA was isolated from flash-frozen mouse tissue samples by using Trizol isolation method and cleaned up with RNAEasy mini columns. cDNA was produced using a high capacity cDNA transcription kit. The primer probe sets PubMed ID:http://www.ncbi.nlm.nih.gov/pubmed/19724269 were ordered from IDT. The quantitative expression of each gene was assessed using Taqman Gene Expression Assays on an Applied Biosystems 7900HT machine. Relative expression was calculated using the delta-delta Ct method. CLAMS Indirect open circuit calorimeter was performed using the Comprehensive Lab Animal Monitoring System. In each experiment, thirty two mice were weighed before testing. Each mouse was placed in its own chamber without bedding and allowed to acclimate to its surroundings; mice were not pre-acclimated to the CLAMS cages prior to the experiment. All experiments began at 910 am and continued for up to 48 hrs. Phase 1 of the study assessed metabolic preference during a non-feeding state; food was removed from the cages. Phase 2 evaluated metabolic preference during a feeding condition; food in excess was provided at 4 PM. The system measured oxygen consumption, carbon dioxide production and calculated the Respiratory Exchange Ratio for each animal. Energy expenditure was also estimated using the formulas: Caloric Value CV 3:815 1:232 REREnergy Expenditure EE CV VO2 Activity was determined using an array of infrared beams surround each cage. Total activity was monitored continuously and any movement that produced a beam break was counted and order Indirubin-3′-oxime summed. Treadmill running Exercise endurance was determined using a Columbus Instruments modular treadmill. Eight mice could be run at a time in this system. Prior to the measurement, all mice were weighed and allowed to orient to the inclined treadmill surroundings for 35 minutes. A ramping program was then begun at 4 m/min for 5 min to allow the animals to acclimate to the movement of the belt and orient them to the shock grid. The speed was increased to 10 m/min for 10 min and then increased 2 m/min every 2 minutes until the animals run to exhaustion. Exhaustion was defined by an RER ~1.0 and the unwillingness to 
move from the shock grid after 6 shocks. Run time, run distance, work performed, VO2, VCO2, respiratory exchange rate were determined during each bout on the treadmill. Respiratory Exchange Ratio during both the CLAMs and treadmill exercise protocols was calculated as RER = VCO2/VO2. Under normal circumstances, an RER value of 0.70 indicates that fat is the predominant fuel source; an RER of 0.85 suggests a mix of fat and carbohydrates, and a value of 1.0 or above suggests that carbohydrates are the predominant fuel source. 15 / 19 CD38 and Exercise Intolerance and Metabolic Inflexibility In our exercise protocols, an RER > than 1.0 served as