Mechanistic Interplay Between Autophagy and Apoptotic Signaling in Endosulfan-Induced Dopaminergic Neurotoxicity: Relevance to the Adverse Outcome Pathway in Pesticide Neurotoxicity

Abstract

Chronic exposure to pesticides is implicated in the etiopathogenesis of Parkinson's disease (PD). Previously, we showed that dieldrin induces dopaminergic neurotoxicity by activating a cascade of apoptotic signaling pathways in experimental models of PD. Here, we systematically investigated endosulfan's effect on the interplay between apoptosis and autophagy in dopaminergic neuronal cell models of PD. Exposing N27 dopaminergic neuronal cells to endosulfan rapidly induced autophagy, indicated by an increased number of autophagosomes and LC3-II accumulation. Prolonged endosulfan exposure (>9 h) triggered apoptotic signaling, including caspase-2 and -3 activation and protein kinase C delta (PKCδ) proteolytic activation, ultimately leading to cell death, thus demonstrating that autophagy precedes apoptosis during endosulfan neurotoxicity. Furthermore, inhibiting autophagy with wortmannin, a phosphoinositide 3-kinase inhibitor, potentiated endosulfan-induced apoptosis, suggesting that autophagy is an early protective response against endosulfan. Additionally, Beclin-1, a major regulator of autophagy, was cleaved during the initiation of apoptotic cell death, and the cleavage was predominantly mediated by caspase-2. Also, caspase-2 and caspase-3 inhibitors effectively blocked endosulfan-induced apoptotic cell death. CRISPR/Cas9-based stable knockdown of PKCδ significantly attenuated endosulfan-induced caspase-3 activation, indicating that the kinase serves as a regulatory switch for apoptosis. Additional studies in primary mesencephalic neuronal cultures confirmed endosulfan's effect on autophagy and neuronal degeneration. Collectively, our results demonstrate that a functional interplay between autophagy and apoptosis dictate pesticide-induced neurodegenerative processes in dopaminergic neuronal cells. Our study provides insight into cell death mechanisms in environmentally linked neurodegenerative diseases.

Keywords: PKCδ; Parkinson’s disease; apoptosis; autophagy; neurotoxicity; pesticides.