Protein kinase C delta mediated cytotoxicity of 6-Hydroxydopamine via sustained extracellular signal-regulated kinase 1/2 activation in PC12 cells

Abstract

Objectives: The incidence of Parkinson's disease (PD) is increasing as the global population ages. 6-hydroxydopamine (6-OHDA) can induce PD-like neuropathology and biochemical changes in both in vitro and in vivo models. Therefore, clarification of the molecular mechanism of 6-OHDA-induced cell death might contribute to the understanding of the pathogenesis of PD.

Methods: With this goal in mind, we investigated the role of protein kinase C delta (PKC delta) in 6-OHDA-dependent death using the pheochromocytoma cell line, PC12. Cells were treated with 6-OHDA to induce toxicity with or without pretreatment using rottlerin (a PKC delta inhibitor), bisindolylmaleimide I (a general PKC inhibitor), Gö6976 (a PKC inhibitor selective for calcium-dependent PKC isoforms), or phorbol-12-myristate-13-acetate (PMA, a PKC activator).

Results: Phorbol-12-myristate-13-acetate decreased cell survival and increased the rate of apoptosis while rottlerin increased cell survival and decreased the rate of apoptosis. In contrast, neither bisindolylmaleimide I nor Gö6976 affected 6-OHDA-induced cell death. Western analysis demonstrated that phosphorylation of PKC delta on Thr 505 as well as extracellular signal-regulated kinase (ERK) phosphorylation increased after exposure to 6-OHDA. This increase in PKC delta phosphorylation was potentiated by PMA. However, rottlerin attenuated the 6-OHDA-stimulated increase in PKC delta and ERK phosphorylation.

Conclusion: These data suggest that PKC delta, rather than classic-type PKC (alpha, beta1, beta2, gamma), participates in 6-OHDA-induced neurotoxicity in PC12 cells, and PKC delta activity is required for subsequent ERK activation during cell death.

Keywords:

• 6-Hydroxydopamine
• Protein kinase C
• Subtype delta
• Extracellular signal-regulated kinases (ERK)
• Parkinson’s disease
• Apoptosis
• Rottlerin

This work was supported by the International Cooperation Project of Science and Technology Department of Heilongjiang Province, China (No.WB08B05), and the Science and Technology Foundation of Education Department of Heilongjiang Province, China (No.11521076). We are grateful to Prof. Hulun Li and Dr Bo Sun (Neurobiology Department of Harbin Medical University), Prof. Baoliang Guo (Surgery Department of the Second Affiliated Hospital of Harbin Medical University), Dr Ryun Huang and Dr Xiangning Meng (Genetic and Molecular Biology Department of the Harbin Medical University) for their kind assistance. We thank Medjaden Bioscience Limited for assisting in the preparation of this manuscript.