Review Pathophysiological roles of extracellular nucleotides in glial cells: differential expression of purinergic receptors in resting and activated microglia Fabio Bianco a,b,1 , Marta Fumagalli a,c,1 , Elena Pravettoni a,b , Nadia D’Ambrosi d , Cinzia Volonte d , Michela Matteoli a,b , Maria P. Abbracchio c, * ,1 , Claudia Verderio a,b, * ,1 a Department of Medical Pharmacology, CNR Institute of Neuroscience, Cellular and Molecular Pharmacology, Milan, Italy b Center of Excellence on Neurodegenerative Diseases, University of Milan, Milan, Italy c Department of Pharmacological Sciences, University of Milan, Milan, Italy d CNR, Fondazione Santa Lucia, Rome, Italy Accepted 9 December 2004 Available online 22 January 2005 Abstract Microglial cells are the major cellular elements with immune function inside the CNS and play important roles in orchestrating inflammatory brain response to hypoxia and trauma. Although a complete knowledge of the endogenous factors leading to a prompt activation of microglia is not yet available, activation of P2 purinoreceptors by extracellular ATP has been indicated as a primary factor in microglial response. A still unresolved question, however, is which subtype(s) of P2 receptors mediate(s) the response to ATP. By a combination of RT-PCR, Western blotting, and single-cell calcium imaging, we assessed the presence and the activity of P2 receptor subtypes in the mouse microglial cell line N9. All members of the P2 receptor family, including the recently reported receptor for sugar nucleotides (P2Y 14 ), were found to be present in these cells at mRNA and/or protein level. The functionality of the receptors was assessed by analysis of the calcium responses evoked by specific agonists both in N9 cells and in primary microglia from rat brain. Interestingly, a different functional profile of P2 receptors was observed in resting or in LPS-activated N9 cells. Overnight exposure to LPS increased P2Y 6 and P2Y 14 , decreased P2X 7 , and left unchanged P2Y 1 and P2Y 2,4 receptor activity. The change in the P2 receptor profile in activated cells suggests selective roles for specific P2 receptor subtypes in microglial activation triggered by LPS. We speculate that modulation of microglial cell function via subtype-selective P2 receptor ligands may open up new strategies in the therapeutic management of inflammatory neurological diseases characterized by abnormal microglia response. D 2004 Elsevier B.V. All rights reserved. Theme: Cellular and molecular biology Topic: Neuroglia and myelin Keywords: Microglia; Calcium signaling; LPS activation; Adenine and uridine nucleotides; UDP–glucose; P2X and P2Y receptors Contents 1. Introduction ........................................................... 145 2. Materials and methods ...................................................... 146 2.1. Cell cultures ....................................................... 146 0165-0173/$ - see front matter D 2004 Elsevier B.V. All rights reserved. doi:10.1016/j.brainresrev.2004.12.004 * Corresponding authors. Maria P. Abbracchio is to be contacted at Department of Pharmacological Sciences, University of Milan Via Balzaretti 9, 20133 Milan, Italy. Fax: +39 02 503 18284. Claudia Verderio, CNR-Institute of Neuroscience, Via Vanvitelli 32, 20129 Milan, Italy. Fax: +39 02 749 0574. E-mail addresses: mariapia.abbracchio@unimi.it (M.P. Abbracchio)8 c.verderio@in.cnr.it (C. Verderio). 1 These authors equally contributed to the work. Brain Research Reviews 48 (2005) 144 – 156 www.elsevier.com/locate/brainresrev