A modified silver technique (de Olmos stain) for assessment of neuronal and axonal degeneration.
ABSTRACT Silver impregnation histological techniques yield excellent visualization of degenerating neurons and their processes in animal models of neurological diseases. These methods also provide a particularly valuable complement to current immunocytochemical techniques for recognition of axon injury in the setting of brain or spinal cord trauma, ischemia, or neurodegenerative diseases. Despite their utility, silver methods are not commonly used because of complex preparation requirements and inconsistent results obtained by inexperienced histologists. This chapter details a modification of the de Olmos amino-cupric-silver protocol, which has been adapted for efficient processing of large numbers of mouse or rat brains. One author (T.I.T.) has used this method for several years to identify degenerating neurons in adult and neonatal rodent brains. A detailed protocol is provided, with attention to the most critical variables in tissue fixation and solution preparation. Examples are shown of axon injury in the rat brain after focal ischemia.
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ABSTRACT: A comprehensive experimental model of controlled overpressure brain impact in rats is presented. Repeatable blast signatures of controlled duration, peak pressure and transmitted impulse enable to reproduce blast impact in laboratory animals in accurate fashion. Animal survival, brain pathomorphology and the expression of GFAP and CNPase after head-directed nonpenetrating blast of 358 kPa magnitude at surface for approximately 10 msec. The high speed imaging demonstrated strong head acceleration/jolting accompanied by typical intracranial hematomas and brain swelling. The microscopic injury was revealed by prominent silver staining in the deep brain areas including nucleus subthalamicus zone suggesting both diffused and focal neurodegeneration. GFAP and CNPase, markers of astroglia and oligodendroglia, respectively, accumulated substantially in hippocampus 24 h after blast and persisted for 30 days post-blast.We propose that following blast overpressure brain impact, critical pathological signatures of blast brain injury may be triggered by complex cerebrovascular responses, including blood brain barrier disruption, glia activation and neuronal alterations.05/2009;
Article: Morphologic and biochemical characterization of brain injury in a model of controlled blast overpressure exposure.[show abstract] [hide abstract]
ABSTRACT: Existing experimental approaches for studies of blast impact in small animals are insufficient and lacking consistency. Here, we present a comprehensive model, with repeatable blast signatures of controlled duration, peak pressure, and transmitted impulse, accurately reproducing blast impact in laboratory animals. Rat survival, brain pathomorphology, and levels of putative biomarkers of brain injury glial fibrillary acid protein (GFAP), neuron-specific enolase, and ubiquitin C-terminal hydrolase (UCH)-L1 were examined in brain, cerebrospinal fluid (CSF), and blood after 10 msec of 358 kPa peak overpressure blast exposure. The high-speed imaging demonstrated a strong head acceleration/jolting accompanied by typical intracranial hematomas and brain swelling. Microscopic injury was revealed by prominent silver staining in deep brain areas, including the nucleus subthalamicus zone, suggesting both diffused and focal neurodegeneration. GFAP and 2',3'-cyclic nucleotide 3'-phosphodiesterase (CNPase), markers of astroglia and oligodendroglia, accumulated substantially in the hippocampus 24 hours after blast and persisted for 30 days postblast. However, GFAP content in the blood significantly increased 24 hours after injury, followed by a decline and subsequent accumulation in CSF in a time-dependent fashion. A similar profile is shown for UCH-L1 increase in blood, whereas increased CSF levels of UCH-L1 persisted throughout 14 days after blast and varied significantly in individual rats. Neuron-specific enolase levels in blood were significantly elevated within 24 hours and 48 hours postblast. The proposed model of controlled nonpenetrating blast in rats demonstrates the critical pathologic and biochemical signatures of blast brain injury that may be triggered by cerebrovascular responses, including blood-brain barrier disruption, glia responses, and neuroglial alterations.The Journal of trauma 03/2010; 69(4):795-804. · 2.48 Impact Factor