Accumulating microglia phagocytose injured neurons in hippocampal slice cultures: involvement of p38 MAP kinase

In this study, microglial migration and phagocytosis were examined in mouse organotypic hippocampal slice cultures, which were treated with N-methyl-D-aspartate (NMDA) to selectively injure neuronal cells. Microglial cells were visualized by the expression of enhanced green fluorescent protein. Dail...

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Bibliographic Details
Published in:PloS one 2012-07, Vol.7 (7), p.e40813-e40813
Main Authors: Katayama, Takahiro, Kobayashi, Hayato, Okamura, Toshiyuki, Yamasaki-Katayama, Yuko, Kibayashi, Tatsuya, Kimura, Hiroshi, Ohsawa, Keiko, Kohsaka, Shinichi, Minami, Masabumi
Format: Article
Language:English
Subjects:
Accumulation
Animals
Aspartate
Attenuation
Biology
Bisphosphonates
Brain
Brain slice preparation
c-Jun protein
Cell Movement - drug effects
Cell Proliferation - drug effects
Clodronic acid
DNA-Binding Proteins
Enzyme inhibitors
Extracellular signal-regulated kinase
Female
Fluorescence
Glutamate receptors
Glutamic acid receptors
Green fluorescent protein
Hippocampus
Hippocampus - pathology
Inhibitors
Injuries
Iodides
JNK protein
Kinases
Male
MAP kinase
Medicine
Mice
Mice, Inbred C57BL
Mice, Transgenic
Microglia
Microglia - drug effects
Microglia - metabolism
Microglia - pathology
Microglial cells
Mitogens
Motility
N-methyl-D-aspartate
N-Methyl-D-aspartic acid
N-Methylaspartate - pharmacology
Nerve Tissue Proteins - metabolism
Neurochemistry
Neurons
Neurons - drug effects
Neurons - enzymology
Neurons - pathology
Neurosciences
Nuclear Proteins - metabolism
p38 Mitogen-Activated Protein Kinases - metabolism
Phagocytosis
Phagocytosis - drug effects
Pharmaceutical sciences
Pharmacology
Propidium - metabolism
Propidium iodide
Protein Kinase Inhibitors - pharmacology
Proteins
Pyramidal Cells - drug effects
Pyramidal Cells - metabolism
Pyramidal Cells - pathology
Rodents
Stratum radiatum
Time Factors
Time-Lapse Imaging
Tissue Culture Techniques
Transcription factors
Transgenic animals
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Summary:In this study, microglial migration and phagocytosis were examined in mouse organotypic hippocampal slice cultures, which were treated with N-methyl-D-aspartate (NMDA) to selectively injure neuronal cells. Microglial cells were visualized by the expression of enhanced green fluorescent protein. Daily observation revealed microglial accumulation in the pyramidal cell layer, which peaked 5 to 6 days after NMDA treatment. Time-lapse imaging showed that microglia migrated to the pyramidal cell layer from adjacent and/or remote areas. There was no difference in the number of proliferating microglia between control and NMDA-treated slices in both the pyramidal cell layer and stratum radiatum, suggesting that microglial accumulation in the injured areas is mainly due to microglial migration, not to proliferation. Time-lapse imaging also showed that the injured neurons, which were visualized by propidium iodide (PI), disappeared just after being surrounded by microglia. Daily observation revealed that the intensity of PI fluorescence gradually attenuated, and this attenuation was suppressed by pretreatment with clodronate, a microglia toxin. These findings suggest that accumulating microglia phagocytosed injured neurons, and that PI fluorescence could be a useful indicator for microglial phagocytosis. Using this advantage to examine microglial phagocytosis in living slice cultures, we investigated the involvements of mitogen-activated protein (MAP) kinases in microglial accumulation and phagocytosis. p38 MAP kinase inhibitor SB203580, but not MAP kinase/extracellular signal-regulated kinase inhibitor PD98059 or c-Jun N-terminal kinase inhibitor SP600125, suppressed the attenuation of PI fluorescence. On the other hand, microglial accumulation in the injured areas was not inhibited by any of these inhibitors. These data suggest that p38 MAP kinase plays an important role in microglial phagocytosis of injured neurons.
ISSN:1932-6203
1932-6203
DOI:10.1371/journal.pone.0040813