Ž . Brain Research 832 1999 84–96 Research report Peripheral injections of Freund’s adjuvant in mice provoke leakage of serum proteins through the blood–brain barrier without inducing reactive gliosis Alexander G. Rabchevsky a, ) , Jean-Denis Degos b , Patrick A. Dreyfus a a INSERM U421, IM3, Faculte de Medecine, 94010 Creteil, France ´ ´ ´ b ( ) SerÕice de Neurologie AP r HP , CHU Henri-Mondor, 94010 Creteil, France ´ Accepted 6 April 1999 Abstract Ž . Breakdown of the blood–brain barrier BBB and ensuing gliosis are common events following physical trauma to the central nervous Ž . Ž . system CNS or during autoimmune diseases such as experimental allergic encephalomyelitis EAE . Some studies of EAE in rodents Ž . report that peripheral injections of complete Freund’s adjuvant CFA , which contains heat-inactivated Mycobacterium to provoke peripheral inflammation without adversely affecting the CNS, can itself lead to increased BBB permeability to small tracer molecules and certain serum proteins. To study the equivocal relationship between serum protein extravasation and reactive gliosis, we injected C57BLr6 mice with CFA and histologically assessed the permeability of various serum proteins and the reactivity of proximal microglia and astrocytes in the uninjured brainstem and spinal cord enlargements after 1–4 weeks. Our results confirm that CFA injections induce progressive increases in the perivascular extravasation of serum IgG, albumin, IgM, and exogenous horseradish peroxidase, all to varying degrees, most prominently in the brainstem and cervical spinal cord after 2–3 weeks. More importantly, neither microglial cells nor astrocytes in regions of focal serum protein leakage appeared morphologically reactive based on immunoreactivity for CR3 receptors Ž . Ž . Mac-1 or glial fibrillary acidic protein GFAP , respectively. Because we found no evidence of T cell infiltration accompanying the exudates, our results indicate that in the absence of physical trauma or inflammatory cells resident CNS neuroglia remain quiescent upon exposure to extravasated serum proteins. q 1999 Elsevier Science B.V. All rights reserved. Keywords: Astrocyte; Complete Freund’s adjuvant; Extravasation; Glial fibrillary acidic protein; IgG; Mac-1; Microglia 1. Introduction Ž . The central nervous system CNS has long been con- sidered an ‘immunologically-privileged’ site based, in part, Ž . on the existence of a functional blood–brain barrier BBB constituted by endothelial cells which limit the passage of solutes and cells from the blood. In this regard, the critical steps in the initiation of inflammatory processes in the CNS parenchyma are perturbations in the structural in- tegrity of the BBB and the recruitment of blood-borne w x cells, most notably T cells 23 . A common feature of CNS inflammation induced by either physical trauma, neurode- generation, or direct injection of proinflammatory agents is the systematic association of serum protein extravasation through the BBB in the affected regions with the activation of resident neuroglial cells leading to reactive gliosis ) Corresponding author. 229 Sanders-Brown Center on Aging, Univer- sity of Kentucky, Lexington, KY 40536-0230, USA. Fax: q1-606-323- 2866; E-mail: agrab@pop.uky.edu w x 1–4,14,26,40,50,55 . Although these events appear to oc- cur concomitantly, the cellular consequences of exposure to extravasated serum proteins in vivo remains unclear due to the myriad of secondary effects stemming from the pathology of CNS insults. Ž . Experimental allergic encephalomyelitis EAE is a cell-mediated autoimmune disease of the rodent CNS that is associated with breakdown of the BBB, inflammatory w x cell recruitment and reactive gliosis 15,17,27,28,33 . The disease is generally induced by peripheral injections of CNS tissue homogenized with complete Freund’s adjuvant Ž . CFA which contains heat-inactivated Mycobacterium tu- w x berculosis 43 . It is common practice to use CFA injec- tions alone as a control for EAE because this leads to peripheral inflammation without the development of clini- cal signs or cellular inflammatory responses in the CNS w x 12,35,53 . Nevertheless, studies have shown that rodents immunized with CFA alone display increased BBB perme- ability to blood-borne tracers and certain serum proteins w x 39,60 , indicating that peripheral inflammation can pro- 0006-8993r99r$ - see front matter q 1999 Elsevier Science B.V. All rights reserved. Ž . PII: S0006-8993 99 01479-1