Date Published: June 10, 2008
Publisher: Public Library of Science
Author(s): Chun-Ting Lee, Jia Chen, Teruo Hayashi, Shang-Yi Tsai, Joseph F Sanchez, Stacie L Errico, Rose Amable, Tsung-Ping Su, Ross H Lowe, Marilyn A Huestis, James Shen, Kevin G Becker, Herbert M Geller, William J Freed, Manuel Graeber
Abstract: BackgroundPrenatal exposure of the developing brain to cocaine causes morphological and behavioral abnormalities. Recent studies indicate that cocaine-induced proliferation inhibition and/or apoptosis in neural progenitor cells may play a pivotal role in causing these abnormalities. To understand the molecular mechanism through which cocaine inhibits cell proliferation in neural progenitors, we sought to identify the molecules that are responsible for mediating the effect of cocaine on cell cycle regulation.Methods and FindingsMicroarray analysis followed by quantitative real-time reverse transcription PCR was used to screen cocaine-responsive and cell cycle-related genes in a neural progenitor cell line where cocaine exposure caused a robust anti-proliferative effect by interfering with the G1-to-S transition. Cyclin A2, among genes related to the G1-to-S cell cycle transition, was most strongly down-regulated by cocaine. Down-regulation of cyclin A was also found in cocaine-treated human primary neural and A2B5+ progenitor cells, as well as in rat fetal brains exposed to cocaine in utero. Reversing cyclin A down-regulation by gene transfer counteracted the proliferation inhibition caused by cocaine. Further, we found that cocaine-induced accumulation of reactive oxygen species, which involves N-oxidation of cocaine via cytochrome P450, promotes cyclin A down-regulation by causing an endoplasmic reticulum (ER) stress response, as indicated by increased phosphorylation of eIF2α and expression of ATF4. In the developing rat brain, the P450 inhibitor cimetidine counteracted cocaine-induced inhibition of neural progenitor cell proliferation as well as down-regulation of cyclin A.ConclusionsOur results demonstrate that down-regulation of cyclin A underlies cocaine-induced proliferation inhibition in neural progenitors. The down-regulation of cyclin A is initiated by N-oxidative metabolism of cocaine and consequent ER stress. Inhibition of cocaine N-oxidative metabolism by P450 inhibitors may provide a preventive strategy for counteracting the adverse effects of cocaine on fetal brain development.
Partial Text: Abuse of cocaine during pregnancy exposes several hundred thousand infants per year to cocaine in the United States alone . A variety of disorders of central nervous system (CNS) development, e.g., intrauterine growth retardation , interference with neuronal migration and differentiation , and neurobehavioral deficits [4,5], have been associated with prenatal exposure to cocaine. Adverse effects of cocaine on brain development have also been demonstrated in nonhuman primates. Prenatal cocaine exposure results in neurobehavioral deficits in subhuman primate infants or adolescents, including deficits in attention and motor maturity . At the cellular level, cocaine exposure induces neocortical cytoarchitectural abnormalities including a decrease in the number of cortical neurons and abnormal positioning of cortical neurons in the primate embryonic cerebral wall [7,8]. Notably, these abnormalities are found only when cocaine is administered during the second trimester (E40–E102), the period when proliferation of neural progenitors is most active . The specific actions of cocaine in the second trimester and the decrease of neuron numbers in the cortex suggest that cocaine may affect important cellular functions of neural progenitor cells.
In the present study, we found that (1) cocaine causes proliferation inhibition and cyclin A down-regulation in neural progenitor cells both in vitro and in vivo; (2) restoring cyclin A reverses proliferation inhibition induced by cocaine; and (3) ROS-induced ER stress, activating the eIF2α-ATF4 pathway, is involved in cyclin A down-regulation induced by cocaine. Thus, this study identifies ES stress-induced cyclin A down-regulation as an important molecular event involved in cocaine-induced proliferation inhibition in neural progenitor cells. A diagram, illustrating this pathway is shown in Figure 8.