Wnt3a ligand facilitates autophagy in hippocampal neurons by modulating a novel GSK-3β-AMPK axis

Juvenal A. Ríos, Juan A. Godoy, Nibaldo C. Inestrosa

In the adult central nervous system (CNS), Wnt signaling regulates dendritic structure and synaptic plasticity. The Wnt signaling pathway can be divided into the canonical (β-catenin-dependent) and non-canonical pathways. In the canonical pathway, the binding of canonical ligands such as Wnt3a to the Frizzled receptor induces inactivation of glycogen synthase kinase-3β (GSK-3β), which stabilizes β-catenin and allows its translocation to the nucleus. However, to date, few studies have focused on β-catenin-independent Wnt signaling or explained the underlying mechanisms connecting Wnt signaling to cellular energy metabolism. A recent study demonstrated negative regulation of 5' adenosine monophosphate-activated protein kinase (AMPK), a major target of GSK-3β that regulates cellular metabolism under diverse conditions. Mainly based on these observations, we evaluated whether Wnt3a ligand modulates autophagy by regulating the GSK-3β/AMPK axis. Cultured primary hippocampal neurons and slices of the CA1 region of rat hippocampus were used. GSK-3β inhibition, AMPK activation, PP2Ac expression, and LC3 processing were examined by western blotting. Autophagic compartments were studied using the CYTO-ID® fluorescent probe, and mature autophagosomes were observed via transmission electron microscopy (TEM). Wnt3a ligand, acting through the Frizzled receptor, promotes the rapid activation of AMPK by inactivating GSK-3β. Biochemical analysis of downstream targets indicated that Wnt3a ligand modulates autophagy in hippocampal neurons. Our results revealed new aspects of Wnt signaling in neuronal metabolism. First, AMPK is an additional target downstream of the Wnt cascade, suggesting a molecular mechanism for the metabolic effects previously observed for Wnt signaling. Second, this mechanism is independent of β-catenin, suggesting a relevant role for non-genomic activity of the Wnt pathway in cellular metabolism. Finally, these results have new implications regarding the role of Wnt signaling in the modulation of autophagy in neurons, with a possible role in the removal of accumulated intracellular proteins.