[beta]2-Adrenergic activation increases glycogen synthesis in L6 skeletal muscle cells through a signalling pathway independent of cyclic AMP

Diabetologia (2007) 50:158167 DOI 10.1007/s00125-006-0484-0

ARTICLE

2-Adrenergic activation increases glycogen synthesis in L6 skeletal muscle cells through a signalling pathway independent of cyclic AMP

D. L. Yamamoto & D. S. Hutchinson & T. Bengtsson

Received: 27 June 2006 / Accepted: 1 September 2006 / Published online: 22 November 2006 # Springer-Verlag 2006

AbstractAims/hypothesis In skeletal muscle, the storage of glycogen by insulin is regulated by glycogen synthase, which is regulated by glycogen synthase kinase 3 (GSK3). Here we examined whether adrenergic receptor activation, which can increase glucose uptake, regulates glycogen synthesis in L6 skeletal muscle cells.Methods We used L6 cells and measured glycogen synthesis (as incorporation of D-[U-14C]glucose into glycogen) and GSK3 phosphorylation following adrenergic activation. Results Insulin (negative logarithm of median effective concentration [pEC50] 8.20.3) and the -adrenergic agonist isoprenaline (pEC50 7.50.3) induced a twofold increase in glycogen synthesis in a concentration-dependent manner. The 1-adrenergic agonist cirazoline and 2-adrenergic agonist clonidine had no effect. Both insulin and isoprenaline phosphorylated GSK3. The -adrenergic effect on glycogen synthesis is mediated by 2-adrenoceptors and not 1-/3-adrenoceptors, and was not mimicked by 8-bromo-cyclic AMP or cholera toxin, and also was insensitive to pertussis toxin, indicating no involvement of cyclic AMP or inhibitory G-protein (Gi) signalling in the 2-adrenergic effect on glycogen synthesis. 12-O-tetradecanoylphorbol-13-acetate (TPA) increased glycogen synthesis 2.5-fold and phosphorylated GSK3 fourfold.

Inhibition of protein kinase C (PKC) isoforms with 12-(2-cyanoethyl)-6,7,12,13-tetrahydro-13-methyl-5-oxo-5H-indolo(2,3-a)pyrrollo(3,4-c)-carbazole (G6976; inhibits conventional and novel PKCs) or 2-[1-(3-dimethylaminopropyl)-5-methoxyindol-3-yl]-3-(1H-indol-3-yl)maleimide (G6983; inhibits conventional, novel and atypical PKCs) inhibited the stimulatory TPA effect, but did not significantly inhibit glycogen synthesis mediated by insulin or isoprenaline. Inhibition of phosphatidylinositol 3-kinase (PI3K) with wortmannin inhibited the effects of insulin and isoprenaline on glycogen synthesis. Conclusions/interpretation These results demonstrate that in L6 skeletal muscle cells adrenergic stimulation through 2-adrenoceptors, but not involving cyclic AMP or Gi, activates a PI3K pathway that stimulates glycogen synthesis through GSK3.

Keywords 2-Adrenoceptor . AMPK . Cyclic AMP. Glycogen . GSK3 . L6 . PI3K . Skeletal muscle

Abbreviations AICAR 5-aminoimidazole-4-carboxamide-1--4-ribofuranoside AMPK AMP activated protein kinase Bmax maximal binding Gi inhibitory G-protein Gq Gq/11 type G-protein Gs stimulatory G-protein GSK3 glycogen synthase kinase 3 KD concentration of ligand required to occupy 50% of the binding sites pEC50 negative logarithm of median effective concentration PI3K phosphatidylinositol 3-kinase PKA protein...