Maria Pannell, Petya Georgieva, Songbo Guo, Susanne Wolf, Vitali Matyash
Communication between neurons is mediated by neurotransmitters and neurohormones and their respective receptors. In the last years it has become evident that microglial cells, astrocytes and oligodendrocytes also express such receptors and are part of that communication system in the CNS.
In this study, we present evidence that microglial cells express serotonin receptors and studied the impact of serotonin receptor activation on distinct microglial properties. First, we tested the impact of serotonin on the microglial response to an insult caused by a laser lesion in acute slices. In the presence of serotonin the microglial processes moved more rapidly towards the laser lesion which is considered to be a chemotactic response to ATP. Similarly the chemotactic response of cultured microglia to ATP was also enhanced by serotonin in a Boyden chamber assay. Quantification of phagocytic activity by determining the uptake of microspheres showed that the amoeboid microglia in slices from early postnatal animals or microglia in culture respond to serotonin application with a decreased phagocytic activity whereas we could not detect any significant change in ramified microglia in situ. The presence of microglial serotonin receptors was confirmed by patch-clamp experiments in culture and amoeboid microglia and by qPCR analysis of RNA isolated from primary cultured and acutely isolated adult microglia. These data suggest that microglia express functional serotonin receptors linked to distinct microglial properties. (funded by Deutsche Forschungsgemeinschaft).
Microglial phagocytic activity in vitro and in situ can be analyzed by uptake of microspheres.
Representative confocal pictures of the uptake of micropsheres (green) by microglial cells (labeled red) in control conditions (left) and upon serotonin application (middle) are displayed for neonatal cultured microglia (A), neonatal amoeboid microglia in situ (B) and ramified adult microglia in mouse coronal brain slices (C). Microglial cells in (A) and (B) were stained with tomatolectin whereas microglia in (C) were visualized by Iba-1 immunohistochemistry. Right panel displays representative orthogonal views of microsphere engulfment in the different preparations. Scale bar: 10 µm.
Krabbe G, Matyash V, Pannasch U, Mamer L, Boddeke HW, Kettenmann H. Activation of serotonin receptors promotes microglial injury-induced motility but attenuates phagocytic activity. Brain Behav Immun. 2012 Mar;26(3):419-28. Epub 2011 Dec 17
In another study, we investigated how micrioglial expression of neurotransmitter and neurohormone receptors is linked to Ca2+ signalling. Visualisation of the calcium change in cultured cells is carried out through the use of fluorescent dyes such as fluo-4. Since these dyes cannot be taken up by microglia in brain slices, we developed a technique to retrovirally insert a calcium indicator into microglia through a stab wound injury in the cortex of a mouse. Microglia proliferating around the injury site were then able to take up the calcium indicator, and the calcium change could be observed upon application of endothelin, histamine, substance P and serotonin. Around 50% of microglia responded to endothelin, histamine and serotonin 6 days after injury, while 100% of microglia responded to substance P. At day 42 this population was reduced to 55%. This response is reflected in cultured microglia activated by LPS. Untreated cultured microglia showed a low response to all substances. These data indicate a possible upregulation in neurotransmitter/-hormone receptors on microglia after brain injury, resulting in beneficial or detrimental changes in function which could be targeted for therapeutic intervention.
Microglia/brain macrophages respond to substance P with an increase in intracellular calcium. (A) Fluorescence images before (A1) and during (A2) application of 4μM substance P and before (A3) and during (A4) application of 1mM ATP. (A5) Tomato lectin staining. (A6) Overlay of the fluorescence of the calcium sensor during application of substance P (A2) and tomato lectin (A5). (B) Changes in the relative fluorescence intensity obtained from a microglial cell at day 6 after stab wound injury during application of substance P and ATP.