Immunohistochemical demonstration of increased prostaglandin F2α levels in the rat hippocampus following kainic acid-induced seizures.

Prostaglandin (PG) F(2α) is one of the major prostanoids biosynthesized by cyclooxygenases (COXs) from arachidonic acid. Although it has been reported that there is a selective surge in PGF(2α) production in the hippocampus during kainic acid (KA)-induced seizure activity, the precise intra-hippocampal distribution of PGF(2α) has not been elucidated due to the paucity of effective histological techniques for detecting PGs in tissues. We investigated the tissue distribution of PGF(2α) in the rat hippocampus 30 min after KA injection by developing fixation and immunohistological-staining methods. To detect PGF(2α) directly on histological sections, we used systemic perfusion fixation with water-soluble carbodiimide fixative, followed by immersion of the brains in Zamboni's fixative. We then performed immunofluorescence staining with anti-PGF(2α) antibody, with negative control experiments used to confirm the staining specificity. Definitive immunolabeling for PGF(2α) was evident most markedly in pyramidal cells of the hippocampal cornu Ammonis (CA) 3 sector and neurons of the hilus in KA-treated rats. Immunolabeling for PGF(2α) was also evident in granule cells of the dentate gyrus. Double immunfluorescence staining revealed that PGF(2α)-immunopositive neurons expressed cytosolic phospholipases A(2), COX-2, and FP receptor. These results suggest that the major source of PGF(2α) production immediately after KA injection was neurons of the hippocampal CA3 sector, hilus and dentate gyrus. These neurons exert PGF(2α)-mediated functions via FP receptors in an autocrine/paracrine manner and may play pathophysiological roles in the acute phase (30 min) of excitotoxicity.

Immunohistochemical localization of aggresomal proteins in glial cytoplasmic inclusions in multiple system atrophy.

AIMS: Multiple system atrophy (MSA) is pathologically characterized by the formation of α-synuclein-containing glial cytoplasmic inclusions (GCIs) in oligodendrocytes. However, the mechanisms of GCI formation are not fully understood. Cellular machinery for the formation of aggresomes has been linked to the biogenesis of the Lewy body, a characteristic α-synuclein-containing inclusion of Parkinson's disease and dementia with Lewy bodies. Here, we examined whether GCIs contain the components of aggresomes by immunohistochemistry. METHODS: Sections from five patients with MSA were stained immunohistochemically with antibodies against aggresome-related proteins and analysed in comparison with sections from five patients with no neurological disease. We evaluated the presence or absence of aggresome-related proteins in GCIs by double immunofluorescence and immunoelectron microscopy. RESULTS: GCIs were clearly immunolabelled with antibodies against aggresome-related proteins, such as γ-tubulin, histone deacetylase 6 (HDAC6) and 20S proteasome subunits. Neuronal cytoplasmic inclusions (NCIs) were also immunopositive for these aggresome-related proteins. Double immunofluorescence staining and quantitative analysis demonstrated that the majority of GCIs contained these proteins, as well as other aggresome-related proteins, such as Hsp70, Hsp90 and 62-kDa protein/sequestosome 1 (p62/SQSTM1). Immunoelectron microscopy demonstrated immunoreactivities for γ-tubulin and HDAC6 along the fibrils comprising GCIs. CONCLUSIONS: Our results indicate that GCIs, and probably NCIs, share at least some characteristics with aggresomes in terms of their protein components. Therefore, GCIs and NCIs may be another manifestation of aggresome-related inclusion bodies observed in neurodegenerative diseases.