Glutamate induces autophagy via the two-pore channels in neural cells

// Gustavo J. S. Pereira 1 , Manuela Antonioli 2, 3 , Hanako Hirata 1 , Rodrigo P. Ureshino 1 , Aline R. Nascimento 1 , Claudia Bincoletto 1 , Tiziana Vescovo 3 , Mauro Piacentini 2, 3 , Gian Maria Fimia 3, 4 , Soraya S. Smaili 1 1 Department of Pharmacology, Federal University of Sao Paulo, (UNIFESP), Sao Paulo, Brazil 2 Department of Biology, University of Rome “Tor Vergata”, Rome, Italy 3 Department of Epidemiology and Preclinical Research, National Institute for Infectious Diseases IRCCS 'Lazzaro Spallanzani', Rome, Italy 4 Department of Biological and Environmental Sciences and Technologies (DiSTeBA), University of Salento, Lecce, Italy Correspondence to: Soraya S. Smaili, email: ssmaili@unifesp.br Gian Maria Fimia, email: gianmaria.fimia@inmi.it Keywords: autophagy, glutamate, NAADP, two-pore channels, AMPK Received: August 20, 2016 Accepted: December 27, 2016 Published: December 31, 2016 ABSTRACT NAADP (nicotinic acid adenine dinucleotide phosphate) has been proposed as a second messenger for glutamate in neuronal and glial cells via the activation of the lysosomal Ca 2+ channels TPC1 and TPC2. However, the activities of glutamate that are mediated by NAADP remain unclear. In this study, we evaluated the effect of glutamate on autophagy in astrocytes at physiological, non-toxic concentration. We found that glutamate induces autophagy at similar extent as NAADP. By contrast, the NAADP antagonist NED-19 or SiRNA-mediated inhibition of TPC1/2 decreases autophagy induced by glutamate, confirming a role for NAADP in this pathway. The involvement of TPC1/2 in glutamate-induced autophagy was also confirmed in SHSY5Y neuroblastoma cells. Finally, we show that glutamate leads to a NAADP-dependent activation of AMPK, which is required for autophagy induction, while mTOR activity is not affected by this treatment. Taken together, our results indicate that glutamate stimulates autophagy via NAADP/TPC/AMPK axis, providing new insights of how Ca 2+ signalling glutamate-mediated can control the cell metabolism in the central nervous system.