Microglial NO induces delayed neuronal death following acute injury in the striatum.
We have established a novel injury model in the central nervous system by a stereotaxic injection of ethanol into rat striatum to induce necrosis. With this model, we clarify a function of inducible nitric oxide synthase (iNOS) in a healing mechanism around a necrotic lesion. A semiquantitative reverse transcriptase-polymerase chain reaction (RT-PCR) revealed that the iNOS mRNA arose at 6 h, peaked at 24 h, and declined to a lower level 48 h after an intrastriatal 5-microL ethanol injection. From in situ hybridization, this iNOS mRNA was expressed in the area surrounding the injury. By immunohistochemistry, mononuclear cells at this boundary area of necrosis were stained with anti-iNOS antibody on the first day after the injury. These cells turned out to be reactive microglia from the positive staining of GSA-I-B4, ED-1 and OX-42. Haematoxylin-eosin (HE) staining showed that neurons in this boundary area gradually disappear up to 5 days after the injury with an increment of microglial cells, and this area became cavernous. Nuclei of neurons in this area were stained positive by the terminal deoxynucleotidyl-transferase-mediated dUTP-biotin nick end-labelling (TUNEL) assay on the first day after the injury. These TUNEL-positive neurons gradually disappeared toward the third day, while microglial cells increased. L-Ng-nitro-arginine methylester (L-NAME), a competitive NOS inhibitor, administration diminished the elimination of neurons by microglia in this boundary area surrounding necrosis. Microglial NO may act as a neurotoxic agent to eliminate damaged neurons near the necrosis in the form of delayed neuronal death, and may reintegrate the neuronal circuits with functionally intact neurons.
Takeuchi
A
A
Department of Immunology, Nagoya University School of Medicine, Aichi, Japan.
Isobe
K I
KI
Miyaishi
O
O
Sawada
M
M
Fan
Z H
ZH
Nakashima
I
I
Kiuchi
K
K
D000818:Animals; D016923:Cell Death / Q000187:drug effects; D003342:Corpus Striatum / Q000187:drug effects* / Q000293:injuries / Q000473:pathology; D053938:DNA Fragmentation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D000431:Ethanol / Q000633:toxicity*; D017628:Microglia / Q000187:drug effects*; D019331:NG-Nitroarginine Methyl Ester / Q000494:pharmacology; D009336:Necrosis; D009474:Neurons / Q000187:drug effects* / Q000473:pathology; D009569:Nitric Oxide / Q000502:physiology*; D019001:Nitric Oxide Synthase / Q000032:analysis / Q000037:antagonists & inhibitors / Q000235:genetics; D052247:Nitric Oxide Synthase Type II; D016133:Polymerase Chain Reaction / Q000379:methods; D051381:Rats; D013238:Stereotaxic Techniques; D014158:Transcription, Genetic
D004791:Enzyme Inhibitors; D009569:Nitric Oxide; D000431:Ethanol; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; C496318:Nos2 protein, rat; D019331:NG-Nitroarginine Methyl Ester