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Induction of NADPH-diaphorase activity in the rat forebrain after middle cerebral artery occlusion
Z C Peng1, C Pietra, A Sbarbati
1Institute of Anatomy and Histology, University of Verona, Italy.
Abstract:
Induction of NADPH-diaphorase (NDP) following ischemic infarction was studied by means of histochemistry in the rat cerebral cortex 1,2,7, and 14 days after distal occlusion of the right middle cerebral artery (MCA). The fine structure of cells in the penumbra region of the necrotic center was also investigated. MCA distal occlusion resulted in ischemic lesion of the frontoparietal cortex of variable extent; NDP induction was detected in neurons, astrocytes, macrophages, and endothelial cells, with regional specificity and a temporal gradient. One, two, and seven days after MCA occlusion, weak NDP positivity was consistently induced in some pyramidal neurons in cortical areas neighboring the necrotic area; NDP induction was also seen in pyramidal neurons of the ipsilateral anterior cingulate and infralimbic cortices and in the tenia tecta. In addition, numerous NDP-positive pyramidal neurons were detected in the contralateral frontoparietal cortex after relatively large ischemic lesions. Two weeks after MCA occlusion, NDP induction in neurons was only evident in the deep cortical layers near the lesion. NDP histochemistry combined with glial fibrillary acidic protein immunofluorescence, performed 7 days after MCA occlusion, indicated that the astrocytes at the periphery of the necrotic area were hypertrophic and some of them were also NDP-positive. One and two days after MCA occlusion, numerous macrophages displaying NDP positivity of variable intensity were seen at the periphery of the necrotic area and in the external capsule of the ischemic cerebral hemisphere. Many endothelial cells in the cortex and subcortical white matter were consistently NDP-positive in all rats. Electron microscopic studies indicated that the area adjacent to the necrotic center was composed of fibrous astrocytes, with the morphological characteristics of proliferation, and numerous lysosome-filled macrophages. Altogether the present results suggest that focal cerebral ischemia may induce in different cell types nitric oxide synthase, which is equivalent to NDP in fixed tissue. The induction of nitric oxide synthase may be related to (1) blood-flow regulation at relatively early postischemic stages, which may decline when collateral circulation is established, and/or (2) cytotoxic or neuroprotective mechanisms.
Insights
Focal cerebral ischemia induces NADPH-diaphorase (NDP) in various brain cells, including neurons and astrocytes. This induction may play roles in regulating blood flow or mediating cytotoxic/neuroprotective mechanisms post-stroke.
Area of Science:
- Neuroscience
- Cerebrovascular Biology
- Histochemistry
Background:
- Ischemic stroke leads to brain tissue damage and cellular changes.
- NADPH-diaphorase (NDP) activity is a marker for nitric oxide synthase (NOS) in the brain.
- Understanding cellular responses to ischemia is crucial for developing treatments.
Purpose of the Study:
- To investigate the induction and cellular localization of NADPH-diaphorase (NDP) in the rat cerebral cortex following focal cerebral ischemia.
- To examine the fine structure of cells in the penumbra region of ischemic lesions.
- To explore the potential roles of NDP induction in post-ischemic brain injury.
Main Methods:
- Histochemical staining for NADPH-diaphorase (NDP) in rat cerebral cortex.
- Induction of focal cerebral ischemia via middle cerebral artery (MCA) occlusion.
- Immunofluorescence for glial fibrillary acidic protein (GFAP) to identify astrocytes.
- Electron microscopy to study cellular ultrastructure in the penumbra region.
Main Results:
- NDP induction was observed in neurons, astrocytes, macrophages, and endothelial cells at various time points (1, 2, 7, 14 days) after MCA occlusion.
- NDP-positive neurons were found in cortical areas adjacent to the lesion, ipsilateral cingulate/infralimbic cortices, and contralateral cortex.
- Hypertrophic, NDP-positive astrocytes and numerous NDP-positive macrophages were present at the lesion periphery; endothelial cells were consistently NDP-positive.
Conclusions:
- Focal cerebral ischemia induces nitric oxide synthase (NOS), equivalent to NDP in fixed tissue, in diverse cell types.
- NDP induction may be linked to early post-ischemic blood-flow regulation or to cytotoxic/neuroprotective mechanisms.
- Cellular responses involving NDP are spatially and temporally regulated following ischemic infarction.