Anzilotti et al. Cell Death and Disease (2018)9:206 DOI 10.1038/s41419-017-0227-9 Cell Death & Disease ARTICLE Open Access Preconditioning, induced by sub-toxic dose of the neurotoxin L-BMAA, delays ALS progression in mice and prevents Na + /Ca 2+ exchanger 3 downregulation Serenella Anzilotti 1 , Paola Brancaccio 2 , Giuseppe Simeone 2 , Valeria Valsecchi 2 , Antonio Vinciguerra 2 , Agnese Secondo 2 , Tiziana Petrozziello 2 , Natascia Guida 1 , Rossana Sirabella 2 , Ornella Cuomo 2 , Pasquale Cepparulo 2 , Andrè Herchuelz 3 , Salvatore Amoroso 2,4 , Gianfranco Di Renzo 2 , Lucio Annunziato 1,2 and Giuseppe Pignataro 2 Abstract Preconditioning (PC) is a phenomenon wherein a mild insult induces resistance to a later, severe injury. Although PC has been extensively studied in several neurological disorders, no studies have been performed in amyotrophic lateral sclerosis (ALS). Here we hypothesize that a sub-toxic acute exposure to the cycad neurotoxin beta-methylamino-L- alanine (L-BMAA) is able to delay ALS progression in SOD1 G93A mice and that NCX3, a membrane transporter able to handle the deregulation of ionic homeostasis occurring during ALS, takes part to this neuroprotective effect. Preconditioning effect was examined on disease onset and duration, motor functions, and motor neurons in terms of functional declines and severity of histological damage in male and female mice. Our findings demonstrate that a sub- toxic dose of L-BMAA works as preconditioning stimulus and is able to delay ALS onset and to prolong ALS mice survival. Interestingly, preconditioning prevented NCX3 downregulation in SOD1 G93A mice spinal cord, leading to an increased number of motor neurons associated to a reduced astrogliosis, and reduced the denervation of neuromuscular junctions observed in SOD1 G93A mice. These protective effects were mitigated in ncx3+/- mice. This study established for the first time an animal model of preconditioning in ALS and candidates NCX3 as a new therapeutic target. Introduction Preconditioning (PC) is a phenomenon wherein a mild insult induces a cellular and tissue resistance to a later severe injury 1 . Over the years numerous stimuli were described as possible PC inductors. Among these, hypoxic stimuli, bacterial toxins such as LPS, small seizures, volatile anesthetics, hyperthermia, and hypothermia 1 . Protection triggered by these stimuli is usually divided by a temporal point of view in acute, rapid PC, and long-lasting, delayed PC 1 . In particular, the delayed PC includes the involve- ment of a genomic reprogramming, which wavers, in most cases, in a downregulation or upregulation of proteins involved in the pathogenesis of the disease 2 . To date, although PC has been extensively studied in several neurological disorders such as Parkinson disease, brain ischemia, and epilepsy, no evidence has been provided on the existence of this PC neuroprotection strategy in amyotrophic lateral sclerosis (ALS). The aim of this study was therefore to identify candidate stimuli and/or genes that can activate the © The Author(s) 2018 Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’ s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/. Correspondence: Giuseppe Pignataro (giuseppe.pignataro@unina.it) 1 IRCCS SDN, Via Gianturco, 80131 Naples, Italy 2 Division of Pharmacology, Department Neuroscience, School of Medicine, Federico II University of Naples, Via Pansini, 5, 80131 Naples, Italy Full list of author information is available at the end of the article Edited by A. Verkhratsky Official journal of the Cell Death Differentiation Association 1234567890():,; 1234567890():,;