Na+/Ca2+ exchanger isoform 1 takes part to the Ca2+-related prosurvival pathway of SOD1 in primary motor neurons exposed to beta-methylamino-l-alanine

Tiziana Petrozziello,Agnese Secondo,Anna Pannaccione,Beatrice Severino,Valeria De Rosa,Angela Corvino,Valentina Tedeschi,Francesca Boscia,Lucio Annunziato

doi:10.1186/s12964-021-00813-z

Abstract

BackgroundThe cycad neurotoxin beta-methylamino-l-alanine (L-BMAA), one of the environmental trigger factor for amyotrophic lateral sclerosis/Parkinson-dementia complex (ALS/PDC), may cause neurodegeneration by disrupting organellar Ca2+ homeostasis. Through the activation of Akt/ERK1/2 pathway, the Cu,Zn-superoxide dismutase (SOD1) and its non-metallated form, ApoSOD1, prevent endoplasmic reticulum (ER) stress-induced cell death in motor neurons exposed to L-BMAA. This occurs through the rapid increase of intracellular Ca2+ concentration ([Ca2+]i) in part flowing from the extracellular compartment and in part released from ER. However, the molecular components of this mechanism remain uncharacterized.MethodsBy an integrated approach consisting on the use of siRNA strategy, Western blotting, confocal double- labeling immunofluorescence, patch-clamp electrophysiology, and Fura 2-/SBFI-single-cell imaging, we explored in rat motor neuron-enriched cultures the involvement of the plasma membrane proteins Na+/Ca2+ exchanger (NCX) and purinergic P2X7 receptor as well as that of the intracellular cADP-ribose (cADPR) pathway, in the neuroprotective mechanism of SOD1.ResultsWe showed that SOD1-induced [Ca2+]i rise was prevented neither by A430879, a P2X7 receptor specific antagonist or 8-bromo-cADPR, a cell permeant antagonist of cADP-ribose, but only by the pan inhibitor of NCX, CB-DMB. The same occurred for the ApoSOD1. Confocal double labeling immunofluorescence showed a huge expression of plasmalemmal NCX1 and intracellular NCX3 isoforms. Furthermore, we identified NCX1 reverse mode as the main mechanism responsible for the neuroprotective ER Ca2+ refilling elicited by SOD1 and ApoSOD1 through which they promoted translocation of active Akt in the nuclei of a subset of primary motor neurons. Finally, the activation of NCX1 by the specific agonist CN-PYB2 protected motor neurons from L-BMAA-induced cell death, mimicking the effect of SOD1.ConclusionCollectively, our data indicate that SOD1 and ApoSOD1 exert their neuroprotective effect by modulating ER Ca2+ content through the activation of NCX1 reverse mode and Akt nuclear translocation in a subset of primary motor neurons.7n_RSb6LJnzKN1GMcoyFawVideo

Highlights

The cycad neurotoxin beta-methylamino-l-alanine (L-BMAA), one of the environmental trigger factor for amyotrophic lateral sclerosis/Parkinson-dementia complex (ALS/PDC), may cause neurodegeneration by disrupting organellar C a2+ homeostasis
Plasma membrane N a+/Ca2+ exchanger isoform 1 (NCX1) mediates a rapid [Ca2+]i increase induced by SOD1 and ApoSOD1 in rat primary motor neurons In consideration of the modulatory role exerted on C a2+ signaling by purinergic P 2X7 receptor [26], cADP-ribose receptor [27, 28], and Na+/Ca2+ exchanger (NCX) [29,30,31], rat primary motor neurons were exposed to SOD1 or ApoSOD1 in presence of their specific antagonists [32,33,34] A430879 (1 μM), 8-bromo-cyclic adenosine diphosphate-ribose (cADPR) (10 μM), or CBDMB (1 μM)
Our results indicated that only the NCX pan inhibitor CB-DMB significantly reduced the early increase of [Ca2+]i induced by SOD1 (Fig. 1A, B) and ApoSOD1 (Fig. 1C, D)

Summary

Introduction

The cycad neurotoxin beta-methylamino-l-alanine (L-BMAA), one of the environmental trigger factor for amyotrophic lateral sclerosis/Parkinson-dementia complex (ALS/PDC), may cause neurodegeneration by disrupting organellar C a2+ homeostasis. Through the activation of Akt/ERK1/2 pathway, the Cu,Zn-superoxide dismutase (SOD1) and its non-metallated form, ApoSOD1, prevent endoplasmic reticulum (ER) stress-induced cell death in motor neurons exposed to L-BMAA. This occurs through the rapid increase of intracellular C a2+ concentration ([Ca2+]i) in part flowing from the extracellular compartment and in part released from ER. The activation of Akt/ERK1/2 pathway via a transient [Ca2+] rise may underline the protective effect of SOD1 [12] This neuroprotective effect is independent from the catalytic activity of the enzyme, since the non-metallated form ApoSOD1, lacking dismutase activity, may induce protection of motor neurons from L-BMAA toxicity likewise SOD1 [12]. The correlation between this neuroprotective increase in [Ca2+]i and Akt activation has been investigated

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Discussion

Conclusion