Obtention and Characterization of Primary Astrocyte and Microglial Cultures From Adult Monkey Brains G. Guillemin, 1 F.D. Boussin, 1 * J. Croitoru, 1 M. Franck-Duchenne, 1 R. Le Grand, 1 F. Lazarini, 2 and D. Dormont 1 1 Service de Neurovirologie, CEA, DSV/DRM/SSA/IPSC, Fontenay-aux-Roses, France 2 Laboratoire de Neuropathologie Raymond Escourolle, Groupe Hospitalier Pitie ´-Salpe ˆtrie `re, Paris, France Simple methods for obtention of primary cultures of isolated astrocytes and microglia from adult simian brain have been developed. Characterization of these two glial cell populations were performed by morpho- logical observations and by immunocytochemistry. The astroglial cultures were obtained by an indirect method. After L-leucine methyl-ester treatment and trypsinizations, more than 99% of cells expressed glial fibrillary acidic protein (GFAP), whereas no macro- phages or microglia could be detected. Likely, the 1% remaining cells were immature astrocytes or cells that lost their GFAP expression. Cultured simian astro- cytes expressed vimentin, laminin, and fibronectin. We also found a constitutively low expression of major histocompatibility complex (MHC) class II by cul- tured astrocytes which was significantly enhanced by lipopolysaccharide (LPS), interferon gamma (IFN-), or tumor necrosis factor alpha (TNF-) treatments. Microglial cultures were obtained by a direct method of isolation using Percoll gradient separations and compared to simian monocyte-derived macrophages or alveolar macrophages. Microglial cells differed from macrophages by their proliferation upon granu- locyte-macrophage colony stimulating factor (GM- CSF) treatment and by their typical morphology when observed by scanning electron microscopy. As macro- phages, they expressed in vitro CD68, CD64, CD14, CD11b, MHC class II, and fibronectin. However, contrary to macrophages, simian cultured microglia expressed laminin. This observation suggests that microglia repre- sent a new potential source of this extracellular matrix protein in the brain. J. Neurosci. Res. 49:576–591, 1997.  1997 Wiley-Liss, Inc. Key words: astrocytes; microglia; macaque; primary cultures INTRODUCTION Glial cells allow more than brain cohesion (Vir- chow, 1846). Especially, astrocytes play important and multiple roles in brain development and functions which still remain poorly understood. Astrocytes represent the most abundant cell type in the central nervous system (CNS). They are highly heterogeneous upon brain local- ization in their profiles of enzymes, antigenic markers, transporters, receptors, and ion channels (Wilkin et al., 1990). Their shape is not static and can be modified upon physiologic and pathologic stimuli. They have a syncy- tium-like organization that is mediated by gap junctions allowing intense intercell communications (Giaume and McCarthy, 1996). Situated around neurons and their processes, they are implicated in neuronal guidance, neurite outgrowth, synaptogenesis, and synaptic plastic- ity (Muller et al., 1995). They provide energy, nutrient, and growth factor support for neurons (Tsacopoulos and Magistretti, 1996); regulate extracellular concentrations of ions (Sykova et al., 1992) and neurotransmitters (Martin, 1995); and have an important function in brain detoxification. Astrocytes are also involved in the mainte- nance of the blood-brain barrier (BBB) because they provide signals for the formation and the cohesion of the tight junctions between endothelial cells (Janzer and Raff, 1987; Giaume and McCarthy, 1996). One of the promi- nent features of astrocytes is their vigorous response following various neurologic insults (Hertz et al., 1990; Mucke and Eddelston, 1993). Reactive astrocytosis is characterized by an increase in number and size of cells expressing glial fibrillary acidic protein (GFAP), a spe- cific intermediate filament cytoskeletal protein (Bignami et al., 1972). Activated astrocytes also express a large number of various molecules normally not detected in resting astrocytes, allowing them to participate in many Contract grant sponsor: Agence Nationale de Recherches sur le SIDA. Contract grant sponsor: Conseil Re ´gional de Bourgogne. *Correspondence to: F.D. Boussin, Service de Neurovirologie, CEA, DSV/DRM/SSA/IPSC, 60-68, Avenue de la Division Leclerc, BP 6 92 265 Fontenay-aux-Roses Cedex, France. E-mail: boussin@dsvidf.cea.fr Received 5 November 1996; Revised 17 April 1997; Accepted 17 April 1997 Journal of Neuroscience Research 49:576–591 (1997)  1997 Wiley-Liss, Inc.