Infusion of hypertonic solutions into the carotid artery is one method by which the blood-brain barrier (BBB) can be opened transiently in experimental animals. This technique has also been tried in clinical situations in which an enhanced uptake of intravenously injected chemotherapeutic drugs into the brain is desired. We have previously found that infusion of hypertonic mannitol or urea into the carotid artery of the rat, leading to a BBB opening, is associated with light microscopic signs of cellular damage in the brain parenchyma. An electron microscopic study has now been made to obtain more detailed information about the events taking place in the rat brain 1 to 72 h after an intracarotid infusion of hyperosmolar solution of mannitol. Toluidine blue-stained semithin epon sections were also available for high-resolution light microscopy of brain samples from urea-infused animals. Intravenously injected Evan's blue dye was used to confirm that BBB opening had occurred as a consequence of the carotid infusions. The infused hemispheres had numerous structural changes. The dominating light microscopic alteration was the presence of multifocal lesions in the gray or the white matter with closely packed microvacuoles causing status spongiosus. Ultrastructurally the microvacuoles corresponded to very pronounced watery swelling of astrocytic processes and to a minor degree to expansion of dendrites and axons. There was also a light or moderate perivascular astrocytic swelling. In the "spongy" lesions as well as occasionally in non-vacuolated parts of the cerebral cortex, there were collapsed electron-dense neurons with pronounced mitochondrial alterations such as severe swelling associated with rupture of christae.(ABSTRACT TRUNCATED AT 250 WORDS)
Structural changes in the rat brain after carotid infusions of hyperosmolar solutions: a light microscopic immunohistochemical study A solution of mannitol or urea was infused into the carotid artery of rats to open the blood-brain barrier (BBB) and to find out if such a procedure results in brain injury, Paraformaldehyde-fixed, paraffin-embedded material was available to determine the localization and extent of albumin extravasation by immunochemistry. Other light microscopic and immunocytochemical techniques were applied on consecutive sections to find out if structural damage had occurred.The cerebral cortex, the hippocampus and the basal ganglia of the infused brain hemisphere contained within regions of albumin extravasation scattered, collapsed, acidophilic neurons. In addition, there were multifocal lesions with marked sponginess of the neuropil which contained numerous shrunken, acidophilic neurons and a perifocal astrocytic gliosis. A moderate macrophage infiltration was present in rats with 72 h survival.In conclusion, infusion of hypertonic mannitol or urea into the carotid artery of the rat may result in structural brain damage within regions showing BBB injury. The presence of acidophilic neurons and the macrophage response indicate that some of the brain changes are irreversible.
An immunohistochemical study was carried out on rat brain to determine if a transient opening of the blood-brain barrier (BBB), leading to extravasation of serum albumin, is also associated with exudation and cellular uptake of fibronectin and fibrinogen. Both of them might exert important biological effects provided that they pass the BBB and come into contact with cells of the brain parenchyma. Hyperosmolar solutions of urea or mannitol were infused in the carotid artery for 30 s to open the BBB and the animals were killed at various time intervals thereafter. Formaldehyde-fixed, paraffin-embedded material was used for immunohistochemical demonstration of extravasated proteins by an avidin-biotin peroxidase technique. Multifocal, often confluent areas of widely different sizes with signs of albumin extravasation were observed both in the grey and the white matter of the cerebral hemispheres exposed to the hyperosmolar solutions. Much less pronounced changes were observed in rats given an intracarotid saline infusion alone. Immunoreactive material indicating extravasation of fibronectin and fibrinogen was present in the infused cerebral hemispheres but albumin immunoreactivity was much more widespread. Reaction product was observed in vascular walls, presumably in extracellular spaces and in nerve cells. Immunoreactivity in the perikaryon of neurons formed different patterns in various cells. A granular type most probably represents accumulation of the proteins in lysosomal organelles after pinocytotic uptake into the neuron. The second so-called diffuse variety is presumably the result of a severe nerve cell injury with an uncontrolled leakage of proteins into the cytoplasm. Our results indicate that vascular walls, extracellular spaces, glial cells and neurons will be exposed to extravasated fibronectin and fibrinogen as well as to albumin and that antigenic sites in such compounds remain for a long period after the BBB opening. In addition, there are indications that carotid infusions of hyperosmolar solutions may cause nerve cell injuries in regions with BBB opening. These findings have obvious clinical and experimental significance.
The presence of plasma proteins in the spinal nerve roots of normal rats was investigated using an avidin‐biotin peroxidase technique on formaldehyde‐fixed, paraffin‐embedded material. Sections from the roots, exposed to a rabbit‐anti rat albumin antiserum showed widespread, intense immunoreactivity which filled the spaces between the nerve fibers. The reaction product usually ended at the junction between the roots and the spinal cord. The sheath enclosing the roots showed the same strong immuno‐reaction. There was also a marked reaction in the dorsal root ganglia and peripheral nerve. Spinal cord sections, however, showed no extracellular reactivity, but many motor neurons of the ventral horn were distinctly positive, presumably the result of a normally occurring retrograde axonal transport from the periphery. Parallel sections from the roots exposed to rabbit anti rat IgG antiserum, rabbit anti rat IgM antiserum, rabbit anti human fibrinogen antiserum and rabbit anti human fibronectin antiserum revealed no positive immunoreaction. Thus, rat spinal nerve roots normally contain material with albumin antigenic properties. This would indicate that albumin is present in the extracellular fluid of the roots in the same way as in the endoneurium of peripheral nerves. The fluid microenvironment of the roots, therefore, appears to be different from that in the CNS which lacks extracellular albumin due to the impermeability of the blood‐brain barrier.
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