COMPLEMENT 5A CONTROLS MOTILITY OF MURINE MICROGLIAL CELLS IN-VITRO VIA ACTIVATION OF AN INHIBITORY G-PROTEIN AND THE REARRANGEMENT OF THE ACTIN CYTOSKELETON

Microglial cells respond to most pathological events by rapid transfor mation from a quiescent to an activated phenotype characterized by inc reased cytotoxicity and motile activity. To investigate the regulation of microglial motility by different inflammatory mediators, we studie d cultured murine microglia by time-lapse video microscopy and a compu ter-based motility assay. Microglial cells exhibited a high resting mo tility. The acute application of complement 5a (C5a) immediately induc ed intense ruffling of microglial membranes followed by lamellipodia e xtension within few seconds, while formyl-Met-Leu-Phe-OH, bacterial en dotoxin (lipopolysaccharide) or inflammatory cytokines did not increas e motility. This process was accompanied by a rapid rearrangement of t he actin cytoskeleton as demonstrated by labelling with fluorescein is othiocyanate-phalloidin and could be inhibited by cytochalasin B. A GT P-binding protein was involved in the signal cascade, since pertussis toxin inhibited motility and actin assembly in response to C5a. Chemot actic migration in a gradient of C5a was also completely blocked by pe rtussis toxin and cytochalasin B. The C5a-induced motility reaction wa s accompanied by an increase in intracellular calcium ([Ca2+](i)) as m easured by a Fluo-3 based imaging system. Ca2+ transients were, howeve r, not a prerequisite for triggering the increase in motility; motilit y could be repeatedly evoked by C5a in nominally Ca2+-free solution, w hile Ca2+ signals occurred only upon the first stimulation. Moreover, conditions mimicking intracellular Ca2+ transients, like incubation wi th thapsigargin or Ca2+ ionophore A23187, were not able to induce any motility reaction, suggesting that Ca2+ transients are not necessary f or, but are associated with, microglial motility. Motile activity was shown to be restricted to a defined concentration range of [Ca2+](i) a s revealed by lowering [Ca2+](i) with BAPTA-AM or increasing [Ca2+](i) with A23187. Since complement factors are released at pathological si tes, this signal cascade could serve to increase motility and to direc t microglial cells to the lesioned or damaged area by means of a G-pro tein-dependent pathway and via the rearrangement of the actin cytoskel eton. Copyright (C) 1996 IBRO. Published by Elsevier Science Ltd.