Figure 4.

CaMKK2 deficiency does not affect contraction-stimulated AMPK phosphorylation/activityand glucose uptake in mouse skeletal muscles. (A) Analysis of LKB1 and AMPK subunit/isoform expression in gastrocnemius muscles from wild-type (WT) or CaMKK2 knock-out (KO) mice (male, 17–18 weeks old). Muscle extracts were subjected to immunoblot analysis using the indicated antibodies. (B) Isolated extensor digitorum longus (EDL) muscles from WT or CaMKK2 KO mice (male, 14–15 weeks old) were incubated in Krebs–Ringer bicarbonate buffer ex vivo for 1 h. During the last 10 min of incubation, muscle contractions were evoked by electrical stimulation. Muscle tissue extracts were subjected to immunoblot analysis with the indicated antibodies. Representative immunoblots are shown. (C and D) Quantification of AMPKα and ACC2 phosphorylation shown in (B). (E and F) Isoform-specific AMPK activity in WT and CaMKK2 KO mice (male, 17 weeks old). (G) Soleus muscle from the indicated genotypes was analyzed using the same experimental setup as (B). (H and I) Quantification of AMPKα and ACC2 phosphorylation shown in (G) and (J and K) isoform-specific AMPK activity from WT and CaMKK2 KO mice (male, 17–18 weeks). (L) [³H]-2-deoxy-glucose uptake in isolated EDL muscle from the indicated genotypes in response to contractions ex vivo. (M and N) Quantification of peak force production and force kinetics during the 10-min contractions. Data are mean ± S.E.M. n = 3 for (C and D) and n = 5–6 (E and F, H–M). A two-way ANOVA test was performed with Tukey's correction for multiple comparisons. ∗P < 0.05 (Basal vs. contracted).