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. 2008 Mar 12;3(3):e1749. doi: 10.1371/journal.pone.0001749

Figure 6. mtNOS Is a final insulin/Akt effector regulating muscle utilization of available glucose.

Figure 6

(A) Representative western blot of disrupted and normal mtNOS, densitometries, and mtNOS activities of mitochondria isolated from right and left gastrocnemius of the same animal, 36 h after direct electroporation of respectively 10 µg of siRNA nNOS or empty pRNAT-U6.1/Neo vector to muscle, as described in Experimental Procedures. (n = 6; densitometry: t = 6.55, DF = 10, P = 0.000; activity at 12 h: t = 6.07, DF = 10, P = 0.000; activity at 24 h: t = 2.22, DF = 10, P = 0.05) (B) To test the interdependence of oxidative and intermediary metabolism as related to activation of mtNOS, metabolic studies including oxygen uptake, glycogen synthesis, complete glucose oxidation to CO2 and H2O, glucose uptake and glycogen synthase activity were performed in nNOS-silenced right gastrocnemius- and vector-administered left muscles of the same animal in parallel under the different conditions by radioactive methods (n = 6; vector: O2 uptake, F = 10.47, DF = 23, P = 0.000; 14CO2, F = 5.06, DF = 23, P = 0.02; (U-14C)-glucose to glycogen, F = 8.61, DF = 23, P = 0.006; 2-deoxyglucose uptake, F = 22.55, DF = 23, P = 0.000; glycogen synthase, F = 3.59, DF = 23, P = 0.032; with siRNAnNOS: O2 uptake, F = 13.78, DF = 23, P = 0.000; 3H2O, F = 17.34, DF = 23, P = 0.000; 14CO2, F = 4.25, DF = 23, P = 0.034; (U-14C)-glucose to glycogen, F = 8.08, DF = 23, P = 0.004; 2-deoxyglucose uptake, F = 12.90, DF = 23, P = 0.000; glycogen synthase, F = 3.62, DF = 23, P = 0.031). (C) Metabolic studies were performed by previous administration of crescent siRNAAkt2 or vehicle under conditions analogous to A and B (n = 3, p-GSK3: t = 7.47, DF = 4, P = 0.002; at 24 h, vehicle vs siRNAAkt2, 14CO2: t = −3.23, DF = 4, P = 0.032; (U-14C)-glucose to glycogen: t = 2.85, DF = 4 P = 0.05). The threshold for significance is the same as that described in Figure 4.*P<.05 vs controls or between nNOS or Akt2 silenced and not-silenced muscle samples; ** P<.05 between different times of insulin administration.