Journal zyxwvutsrqponmlkjihgfedcbaZYXWVUTSRQPONMLKJIHGFEDCBA of Hepatology zyxwvutsrqponmlkjihgfedcbaZYXWVUTSRQPONMLKJIHGFEDCBA 2000; 32:85-91 Printed in Denmark . All rights reserved Mtmksgaard . Copenhagen Copyright 0 European Association for the S&xiv of the Liver 2OfXJ zyxwvuts Journal of Hepatology ISSN 0168-8278 Increased GABAergic activity inhibits a-fetoprotein mRNA expression and the proliferativeactivity of the HepG2 human hepatocelhku carcinoma cell line Manna Zhang, Yeuwen Gong, Nimer Assy and Gerald Y. Minuk Liver Diseases Unit, Departments of Medicine & Pharmacology, University of Manitoba, W innipeg, Manitoba, Canada Background/Aims: Gamma aminobutyric acid (GABA) is a potent inhibitory neurotransmitter with growth regulatory properties Recent data indicate that increased GABAergic activity inhibits hepatocyte proliferation in regenerating livers. In the present study, we aimed to investigate whether GABA inhibits the growth of malignant hepatocytes. Methods: Increasing concentrations of muscimol (0.05-50 ,&l), a specific GABA* receptor agonist, were added to HepG2 human hepatocelhdar carci- noma cells and alpha-fetoprotein (AFP) and albumin mRNA expression were determined for varying periods of time (maximum 24 h) thereafter. Cell pro- liferation was also documented after 48 h of exposure to muscimol. Results: Muscimol significantly (j~O.0001) de- creased AFP mRNA expression (maximum decrease: 65% below baseline values) without affecting albumin mRNA expression. However, the effect on AFP mRNA was transient (maximum duration: 3-6 h) and not associated with changes in cell proliferation. Be- cause preliminary data indicate that GABA* receptor LTHOUGH originally identified as the principal in- zyxwvutsrqponmlkjihgfedcbaZYXWVUTSRQPONMLKJIHG A hibitory neurotransmitter in the mammalian brain, gamma aminobutyric acid (GABA) has since been demonstrated to be present and physiologically active in different tissues throughout the body (1). In the majority of these sites, GABA is thought to play a role in morphogenesis and maturation of the develop- Received 15 April; revised 22 June; accepted 23 June 1999 Correspondence: Gerald Y. Minuk, Liver Diseases Unit, GF407, Health Sciences Centre, 820 Sherbrook Street, Winnipeg, Manitoba, R3A lR9, Canada. Tel: 204 787 4662. Fax: 204 775 4255. e-mail: gminuk@cc.umanitoba.ca activity is markedly downregulated in malignant hepatocytes, transfection studies were performed wherein HepG2 cells were cotransfected with GABA* receptor /lt and y2 subunit genes in a pCDM8 express- ion vector or vector alone followed by re-exposure to either muscimol (5 PM) or saline. In this series of experiments, in addition to AFP mRNA inhibition being as extensive and more prolonged (maximum duration: 6-12 h) in muscimol-treated, GABA* recep- tor-transfected cells, proliferative activity was also significantly inhibited when compared to saline- treated GABA* receptor-transfected controls (p<O.Ol) and muscimol-treated cells transfected with vector alone QKO.005). Conclusion: The results of this study indicate that in- creased GABAergic activity inhibits AFP mRNA ex- pression and cell proliferation in this malignant he- patocyte cell line. Key words: Albumin; Alpha-fetoprotein, GABA; Hepatocelhrlar carcinoma; Muscimol; Regulation. ing tissue (2,3). GABA has also been reported to in- hibit squamous cell carcinoma growth in mice and GABA-like compounds enhance the differentiated state of erythroleukemic, ovarian and colon carcinoma cell lines (47). The mammalian liver contains both a sodium-de- pendent, bicuculine insensitive GABA transport sys- tem and a sodium-independent, bicuculine sensitive GABAA receptor system (8,9). The precise role of these carefully regulated and integrated systems has yet to be determined. Recently, our laboratory documented that increased GABAergic activity inhibits the growth of the liver following partial hepatectomy and during recovery from ethanol, galactosamine and carbon 85