Norepinephrine increases glucose transport in brown adipocytes via beta(3)-adrenoceptors through a cAMP, PKA, and PI3-kinase-dependent pathway stimulating conventional and novel PKCs

To identify the signaling pathways that mediate the adrenergic stimulation of glucose uptake in brown adipose tissue, we used mouse brown adipocytes in culture. The endogenous adrenergic neurotransmitter norepinephrine ( NE) induced 2-deoxy-D-glucose uptake 3-fold in a concentration-dependent manner (pEC(50) similar to 6.5). The uptake was abolished by high doses of propranolol. The NE effect was mimicked by isoprenaline (pEC(50) similar to 6.9), BRL 37344 (pEC(50) similar to 8.6), CL 316243 (pEC(50) similar to 9.7) and CGP 12177 (pEC(50) similar to 7.3) and was thus mediated by beta(3)-adrenergic receptors. The NE-induced effect on 2-deoxy-D-glucose uptake was mediated by adenylyl cyclase and cAMP because responses were inhibited by the adenylyl cyclase inhibitor 2', 5'-dideoxyadenosine and the protein kinase A inhibitor 4-cyano-3-methylisoquinoline. Cholera toxin and 8-bromoadenosine cAMP were both able to increase 2-deoxy-D-glucose uptake. Involvement of other adrenergic signaling pathways (alpha(1)- and alpha(2)-adrenergic receptors) were excluded. The phosphatidylinositol 3-kinase (PI3K) inhibitor LY294002, abolished beta-adrenergic- or 8-bromoadenosine cAMP-stimulated 2-deoxy-D-glucose uptake, demonstrating that a cAMP-dependent PI3K-mediated pathway is positively connected to glucose uptake. Inhibition of the beta-adrenergically stimulated response with protein kinase C (PKC) inhibitors (Go 6983, which inhibits (alpha, beta, gamma), (delta), and (zeta) isoforms and Ro-31-8220, which inhibits (alpha, beta1, beta2, gamma) and (epsilon) but not atypical isoforms) indicated that cAMP-mediated glucose uptake is stimulated via conventional and novel PKCs. These results demonstrate that adrenergic stimulation, through beta(3)-adrenergic receptors/ cAMP/protein kinase A, recruits a PI3K pathway stimulating conventional and novel PKCs, which mediate glucose uptake in brown adipocytes.