Residual and Irreversible Effects of Prolonged Methamphetamine Use on Neural Progenitor Cells in the Human Brain

Abstract

Methamphetamine (METH) abuse has reached epidemic proportions, and it has become increasingly recognized that abusers suffer from a wide range of neurocognitive deficits. Much previous work has focused on the deleterious effects of METH on mature neurons, but little is known about the effects of METH on neural progenitor cells (NPCs). It is now well established that new neurons are continuously generated from NPCs in the adult hippocampus, and accumulating evidence suggests important roles for these neurons in hippocampal-dependent cognitive functions. In a rat hippocampal NPC culture system, we find that METH results in a dose-dependent reduction of NPC proliferation, and higher concentrations of METH impair NPC survival. NPC differentiation, however, is not affected by METH, suggesting cell-stage specificity of the effects of METH. We demonstrate that the effects of METH on NPCs are, in part, mediated through oxidative and nitrosative stress. Further, we identify seventeen NPC proteins that are post-translationally modified via 3-nitrotyrosination in response to METH, using mass spectrometric approaches. One such protein was pyruvate kinase isoform M2 (PKM2), an important mediator of cellular energetics and proliferation. We identify sites of PKM2 that undergo nitrotyrosination, and demonstrate that nitration of the protein impairs its activity. Thus, METH abuse may result in impaired adult hippocampal neurogenesis, and effects on NPCs may be mediated by protein nitration. Our study has implications for the development of novel therapeutic approaches for METH-abusing individuals with neurologic dysfunction and may be applicable to other neurodegenerative diseases in which hippocampal neurogenesis is impaired.

Table of Contents

Abstract2

Introduction4

Materials and methods6

Cell culture6

Immunocytochemistry and quantification of cell types6

Assays of cell viability and apoptosis7

Detection and inhibition of oxidative stress in NPCs8

Mass spectrometric analysis9

Nitrotyrosination and activity of native PKM2 protein11

Results11

Characterization of cultures of adult hippocampal progenitor cells11

METH impairs proliferation and survival of NPCs12

METH does not affect initial NPC differentiation14

METH induces oxidative and nitrosative stress in NPCs15

METH leads to nitrotyrosination and decreased activity of PKM2 in NPCs18

Discussion20

Summary22

End Notes24

Residual and Irreversible Effects of Prolonged Methamphetamine Use on Neural Progenitor Cells in the Human Brain

Introduction

Over 35 million people internationally abuse METH, and in the United States METH abuse has reached epidemic proportions. Through mechanisms that are not yet well understood, METH abusers suffer from a variety of neurocognitive deficits, including behavioral changes, executive dysfunction, deficits in perceptual speed and information manipulation, and impairment of verbal and spatial memory. Neurocognitive deficits may persist after cessation of METH abuse, are slow to improve, and may not completely reverse [1-5]. Although METH was initially thought to selectively damage monoaminergic nerve terminals, recent studies have consistently shown that widespread neuronal cell death results [6-10]. Cell death involves not only the striatum and cortex, but the hippocampus as well [7,11]. Although the molecular mechanisms underlying METH neurotoxicity are likely multifactorial, several key findings support a significant role for both oxidative and nitrosative stress. Mice overexpressing superoxide dismutase, an antioxidant, show markedly decreased METH-induced apoptosis [12]. Suppression of nitric oxide (NO) production, through both pharmacologic ...