Ethanol induced changes in cyclin-dependent kinase-5 activity and its activators, P35, P67 (Munc-18) in rat brain Yadavalli Rajgopal, Mohan C. Vemuri 1, * School of Life Sciences, University of Hyderabad, Hyderabad - 500 046, India Received 14 March 2001; received in revised form 5 June 2001; accepted 8 June 2001 Abstract The expression of cyclin-dependent kinase-5 (Cdk5) and its regulators, p35 and p67 was investigated in adult rat cerebral cortex and cerebellum, using an experimental paradigm of in vivo chronic ethanol exposure. In parallel, the activity of Cdk5 kinase was measured using a speci®c substrate histone-H1 peptide. Western blot analysis revealed no appreciable change in the expression of Cdk5 protein levels while, its regulatory proteins, p35 and p67 showed decreased levels following chronic ethanol treatment. However, ethanol treatment resulted in increased Cdk5 activity in both cortex and cerebellum with relatively high activity in cortex. Given the abundant expression and functions of Cdk5 in neural cells, our data implies a regulatory role for Cdk5 in ethanol mediated cell injury and may contribute to impaired CNS development in brain atrophy associated with alcoholic neurodegeneration. q 2001 Published by Elsevier Science Ireland Ltd. Keywords: Ethanol; Cyclin-dependent kinase-5; p35; p67; Munc-18; Cerebral cortex; Cerebellum Cyclin-dependent kinase-5 (Cdk5) is a serine/threonine proline-directed kinase widely distributed in mammalian tissues with relatively high expression and kinase activity in postmitotic neurons [16±18]. Cdk5 activity depends on absolute requirement of brain-speci®c activators p35/p25, p39, and p67 (Munc-18) which bind to the catalytic cleft of Cdk5 interacting with N- and C- terminus [17]. Cdk5 and its regulatory protein p35, also known as neuronal Cdk5 activator (Nck5a) are involved in cytoskeletal structural dynamics, neuronal regeneration and pathological condi- tions such as Alzheimer's disease (AD) [8,13] due to tau hyperphosphorylation by which b-amyloid protein triggers neuronal death in AD. In addition, Cdk5 is associated with apoptotic cell death during development and tissue remodel- ing due to its capabilities to form multimeric complexes with different proteins in cells [21]. Recent gene knockout studies of Cdk5 2/2 and p35 2/2 emphasize a critical regula- tory role of Cdk5/p35 in the loss of speci®c subsets of corti- cal plate neurons [12]. Alcohol is a neuroactive addictive drug with profound effects on the signal transduction cascade involving protein kinase/phosphatase systems [4]. Protein kinases modulate cell growth, differentiation, cell death and are implicated in several neurological disorders [15]. Chronic alcohol abuse is associated with loss of neurons in speci®c regions of the brain, a pathological process known as alcohol related neuronal loss [5]. To our knowledge, the expression and activity of Cdk5 and the protein levels of p35, p67 have not been reported in chronic ethanol paradigm. In the present study, using a rat model of in vivo chronic ethanol exposure, we evaluated the role of Cdk5 and its regulators p35 and p67 (Munc-18) in the cerebral cortex and cerebel- lum of adult rat brain. Identi®cation of speci®c protein kinases that mediate alcohol effects on CNS provides a clue for the development of drug antagonists to treat alco- hol-related neurological disorders. Male Wistar strain rats (70 ^ 10 g, 1 month old) were maintained in the animal house facility at a temperature (258±288C) and light/dark cycle (12/12 h). Experimental protocols for the use of animals were followed as approved by the institutional as well as national ethical committee guidelines. The animals were divided into control, isocalo- ric and ethanol-treated groups. Each group (n  5 animals) had free access to standard commercial rat chow obtained from National Institute of Nutrition, Hyderabad, India. Control group of rats received basal diet and water while, Neuroscience Letters 308 (2001) 173±176 0304-3940/01/$ - see front matter q 2001 Published by Elsevier Science Ireland Ltd. PII: S0304-3940(01)02011-0 www.elsevier.com/locate/neulet * Corresponding author. Tel.: 11-215-503-7837; fax: 11-215- 923-3808. E-mail address: mohan.c.vemuri@lycos.com (M.C. Vemuri). 1 Present address: Department of Pathology, Anatomy and Cell Biology, Building: JAH Room # 511, Thomas Jefferson Univer- sity, 1020 Locust Street, Philadelphia, PA 19107, USA.