Objective To establish a practical microPET imaging procedure for the measurement of myocardial metabolic rate of glucose (MRGlu) in mice.Methods Twenty wild-type BKS mice were divided into 4 groups by random number table method.The mice were anesthetized with different concentrations of isoflurane (1.3％,1.5％,1.8％,2.0％) at the temperature between 30 ℃ to 34 ℃.The respiratory rate and the physiologic condition were monitored for adjusting the most appropriate isoflurane concentration.Then,different volumes of saline were injected to the anesthetized mice and blood glucose concentrations were measured to test the optimal injection volume.Under the optimal operating condition,18 F-FDG were injected in a group of six mice and followed by microPET imaging.Left ventricular TAC was obtained by drawing ROI and myocardial glucose SUV was also calculated.Meanwhile,the TAC from venous sampling at different time points after 18F-FDG injection was generated.Kinetic Imaging System was used to estimate the coefficients and calculate the MRGlu(Ki ×Glu/LC; Ki =k1 ×k3/(k2+k3).One-way analysis of variance and q test were used to analyze the data.Results No movement was observed in non-fasted mice anesthetized with (1.5-1.8)％ isoflurane,and their respiratory rates were all over 80 per minute.The plasma glucose concentration showed no difference at each time point between the experimental group injected with 75 μl saline and the control group (F=1.215,P＞0.05).The plasma glucose concentration of mice injected with 150 μl saline exhibited statistically significant difference at 30 min (q =2.485,P=0.024),45 min (q =2.287,P=0.036) and 60 min (q =2.709,P =0.015).When the injection volume reached 300 μl,the blood glucose concentration increased remarkably at 45 min (q =2.435,P=0.027).Mice were maintained in good condition after injected with 18F-FDG ranging from 7.4 to 11.1 MBq within 75 μl volume,meanwhile clear and stable myocardial microPET images could also be obtained.The median myocardial SUV value was 11.88(9.71-14.93),Ki value was 0.19 (0.10-0.54) ml · min-1 · g-1and MRGlu value was 19.64 (5.55-23.28) mg · kg-1 · min-1at 45-55 min after 18F-FDG injection.Conclusion The microPET imaging may be a reliable,practical method to evaluate myocardial glucose uptake rate and metabolic rate in mice under the precondition of optimal isoflurane anesthetization (1.5％-1.8％)and small volume of 18F-FDG injection (≤＜75 μl).