COCAINE-RESPONSIVE GENE EXPRESSION CHANGES IN RAT HIPPOCAMPUS W. M. FREEMAN, K. BREBNER, W. J. LYNCH, D. J. ROBERTSON, D. C. S. ROBERTS and K. E. VRANA* Center for the Neurobiological Investigation of Drug Abuse, Department of Physiology and Pharmacology, Wake Forest University School of Medicine, Medical Center Boulevard, Winston-Salem, NC 27157-1083, USA AbstractöChronic cocaine use is known to elicit changes in the pattern of gene expression within the brain. The hippocampus plays a critical role in learning and memory and may also play a role in mediating behaviors associated with cocaine abuse. To pro¢le the gene expression response of the hippocampus to chronic cocaine treatment, cDNA hybridization arrays were used to illuminate cocaine-regulated genes in rats treated non-contingently with a binge model of cocaine (45 mg/kg/day, i.p.) for 14 days. Validation of mRNA changes illuminated by hybridization array analysis was accomplished by measuring immunoreactive protein (via speci¢c immunoblots). The induction of protein kinase CK, potassium channel 1.1, and metabotropic glutamate receptor 5 seen by hybridization arrays was con¢rmed at the level of protein. Immunoblot screening of previously described cocaine-responsive genes demonstrated increased levels of protein tyrosine kinase 2, L-catenin, and protein kinase CO. While some of these changes exist in previously described cocaine- responsive models, others are novel to any model of cocaine use. The inductions of potassium channel 1.1, protein tyrosine kinase 2 and metabotropic glutamate receptor 5 are novel ¢ndings to hippocampal cocaine-responsive gene expression. These proteins have been shown to subserve learning and memory and/or long-term potentiation functions within the hippocampus. Additionally, these genes are known to interact with one another, forming a more complex pattern of gene expression changes. The ¢ndings suggest altered expression of genes with a number of di¡erent functions in the rat hippocampus after a `binge' style of non-contingent cocaine administration. These changes in gene expression may play roles in neuronal plasticity and the behavioral phenomena associated with cocaine abuse. ß 2001 IBRO. Published by Elsevier Science Ltd. All rights reserved. Key words: cDNA array, protein tyrosine kinase 2, protein kinase C, L-catenin, metabotropic glutamate receptor, potassium channel Kv1.1. Chronic psychomotor stimulant use is well known to alter neuronal gene expression (for review see: White and Kalivas, 1998; Nestler, 2001). Speci¢cally, cocaine administration has been shown to a¡ect the expression of a number of genes in di¡erent brain regions. These genes encode a wide variety of molecular functions, ranging from transcription factors, to neurotransmitter receptors and transporters, and signal transduction mechanisms. Some of these changes in gene expression have been shown to subserve the behavioral e¡ects of cocaine (Carlezon et al., 1998; Kelz et al., 1999). While the iden- ti¢cation of this gene set represents a signi¢cant step forward in understanding cocaine's neurobiological actions, it is likely that there are many other cocaine- induced changes in gene expression that have yet to be discovered. Since the dopamine (DA) transporter has been shown to be the site of action involved in mediating cocaine's reinforcing e¡ects, much attention has focused on areas of the brain that receive a dense DA innervation, such as the dorsal and ventral striatum. These regions have been shown to be key mediators of cocaine's reinforcing e¡ects (Roberts et al., 1977; Koob and Le Moal, 1997). However, other areas of the brain interconnected with the mesolimbic DA system are also involved in pro- cessing information related to cocaine use. Hence, increased attention is now being directed towards the involvement of a number of limbic system structures, such as the hippocampus, and amygdala as well as cog- nition-related regions such as prefrontal cortex. In par- ticular, the hippocampus is known to subserve functions 371 *Corresponding author. Tel. : +1-336-716-8620 ; fax : +1-336-716- 8501. E-mail address : kvrana@wfubmc.edu (K. E. Vrana). Abbreviations : AP-1, activator protein complex 1; Cdk5, cyclin- dependent kinase 5; DA, dopamine; EDTA, ethylenediamine- tetraacetic acid ; ELK, ets oncogene proteins ; Gi K1 , G-protein inhibiting K 1; HEPES, N-(2-hydroxyethyl)piperazine-NP-(2-etha- nesulfonic acid) ; Kv1.1, shaker-related voltage-gated potassium channel member ; LTP, long-term potentiation ; MEK1, mitogen- activated protein kinase kinase 1; mGluR5, metabotropic gluta- mate receptor 5; NAcc, nucleus accumbens; OATP2, organic anion transporter polypeptide 2 ; PAGE, polyacrylamide gel elec- trophoresis; PKA cK , protein kinase A K catalytic subunit ; PKCK, protein kinase C K ; PKCO, protein kinase C O ; PP2A, protein phosphatase 2A K catalytic subunit ; PYK2, protein tyrosine ki- nase 2; Ras-GAP, Ras-GTPase-activating protein; SDS, sodium dodecyl sulfate; SMN, survival of motor neuron; SSC, saline sodium citrate. NSC 5273 28-11-01 Cyaan Magenta Geel Zwart www.neuroscience-ibro.com Neuroscience Vol. 108, No. 3, pp. 371^380, 2001 ß 2001 IBRO. Published by Elsevier Science Ltd All rights reserved. Printed in Great Britain PII:S0306-4522(01)00432-8 0306-4522 / 01 $20.00+0.00