Nupr1 Modulates Methamphetamine-Induced Dopaminergic Neuronal Apoptosis and Autophagy through CHOP-Trib3-Mediated Endoplasmic Reticulum Stress Signaling Pathway.

Zhoumeng Lin,Xiang Xu,Wei-Bing Xie,Xu Zhao,Rui Chen,Huijun Wang,Xuebing Chen,Chao Liu,Yunchun Tai,Chuanxiang Chen,Enping Huang

doi:10.3389/fnmol.2017.00203

Abstract

Methamphetamine (METH) is an illegal and widely abused psychoactive stimulant. METH exposure causes detrimental effects on multiple organ systems, primarily the nervous system, especially dopaminergic pathways, in both laboratory animals and humans. In this study, we hypothesized that Nuclear protein 1 (Nupr1/com1/p8) is involved in METH-induced neuronal apoptosis and autophagy through endoplasmic reticulum (ER) stress signaling pathway. To test this hypothesis, we measured the expression levels of Nupr1, ER stress protein markers CHOP and Trib3, apoptosis-related protein markers cleaved-caspase3 and PARP, as well as autophagy-related protein markers LC3 and Beclin-1 in brain tissues of adult male Sprague-Dawley (SD) rats, rat primary cultured neurons and the rat adrenal pheochromocytoma cells (PC12 cells) after METH exposure. We also determined the effects of METH exposure on the expression of these proteins after silencing Nupr1, CHOP, or Trib3 expression with synthetic small hairpin RNA (shRNA) or siRNA in vitro, and after silencing Nupr1 in the striatum of rats by injecting lentivirus containing shRNA sequence targeting Nupr1 gene to rat striatum. The results showed that METH exposure increased Nupr1 expression that was accompanied with increased expression of ER stress protein markers CHOP and Trib3, and also led to apoptosis and autophagy in rat primary neurons and in PC12 cells after 24 h exposure (3.0 mM), and in the prefrontal cortex and striatum of rats after repeated intraperitoneal injections (15 mg/kg × 8 injections at 12 h intervals). Silencing of Nupr1 expression partly reduced METH-induced apoptosis and autophagy in vitro and in vivo. These results suggest that Nupr1 plays an essential role in METH-caused neuronal apoptosis and autophagy at relatively higher doses and may be a potential therapeutic target in high-dose METH-induced neurotoxicity.

Highlights

Methamphetamine (METH) is an illicit and one of the most widely used addictive central nervous system stimulants
Immunofluorescence staining results demonstrated that subacute METH treatment increased Nupr1 expression in both prefrontal cortex and striatum of rats (Figure 3C). These results suggest that METH exposure upregulates Nupr1 and CHOP protein expression, which may contribute to METH-induced endoplasmic reticulum (ER) stress in vivo
The results showed that the number of TUNEL-positive cells was increased by more than 8-fold in PC12 cells transfected with control small hairpin RNA (shRNA) and exposed to METH compared with vehicle treatment, and the number of TUNEL-positive cells was decreased by >2-fold in PC12 cells treated with METH and shNupr1 #2 compared with control shRNA group (Figures 4B,B1)

Summary

Introduction

Methamphetamine (METH) is an illicit and one of the most widely used addictive central nervous system stimulants. It has been shown that METH exposure causes neuronal death in several brain areas, such as the striatum, indusium griseum, medial habenular nucleus and amygdale (Bowyer and Ali, 2006; Kitamura et al, 2007). In addition to these results observed in vivo, other studies, including ours, have demonstrated that METH exposure can cause apoptosis and autophagy in dopaminergic neurons (Castino et al, 2008; Kongsuphol et al, 2009; Huang et al, 2015; Chen C. et al, 2016; Roohbakhsh et al, 2016; Li et al, 2017). The mechanisms underlying METH-induced apoptosis and autophagy in neuronal cells have not yet been elucidated

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