Effect of steroids on brain lipocortin immunoreactivity.

LCT-1, LCT-2 and LCT-5 were assessed in uninjured rats and rats subjected to a cortical freezing injury or middle cerebral artery (MCA) occlusion. Apart from animals receiving no treatment, other uninjured or injured animals received methylprednisolone (2 or 30 mg/kg) or the 21-aminosteroid U-74389F (10 mg/kg) one day and 2 hours before killing. The animals were killed by decapitation 1 hour after the freezing injury or the MCA occlusion and the area containing the lesion was removed and frozen in Freon. Frozen sections were treated with rabbit polyclonal anti-LCT antibody; binding of antibody was visualized by horseradish peroxidase-conjugated swine antirabbit antibody. Without steroid pretreatment, in the uninjured brain LCT immunoreactivity was absent in the greater part of the brain, except in sporadic microglia. In steroid-pretreated animals and in the freezing lesion of both pretreated and untreated animals there was extensive immunostaining; in the freezing lesion it may be due to passage of systemic LCT across the impaired blood-brain barrier in the lesion. The cellular elements showing immunostaining were meningeal cells, neurons, ependyma, choroid plexus, oligodendroglia and capillary endothelium. It implies that also in the brain the steroid effect is consistent with LCT formation.

The effect of steroid treatment on lipocortin immunoreactivity of rat brain.

Lipocortin-1, lipocortin-2 and lipocortin-5 were immunohistochemically assessed in rats. Apart from animals receiving no treatment, other animals received pretreatment with methylprednisolone, or the 21-aminosteroid U-74389F. Whereas Hpocortin immunoreactivity was absent in the greater part of the brain in animals not pretreated with steroid (except in sporadic microglial cells and choroid plexus), there was obvious immunostaining of parenchymatous elements in steroid pretreated animals. In the steroid pretreated animals lipocortin immunoreactivity of the brain tissue may indicate local formation of lipocortin under the influence of steroids that had entered the tissue. The cellular elements which showed immunostaining included meningeal cells, neurones, ependyma, oligodendroglia and capillary endotheHum.

Intracerebral and subcutaneous xenografts of human SCLC in the nude rat: comparison of monoclonal antibody localization and tumor infiltrating lymphocytes.

In the WAG/Rij nude rat, subcutaneous (s.c.) and intracerebral (i.c.) xenografts of the human SCLC cell line GLC-28 were evaluated for their growth behavior, in vivo monoclonal antibody binding and presence of tumor infiltrating lymphocytes. For the i.c. xenografts, two models of cerebral tumor growth were studied, one in the cerebral cortex and one in the lateral ventricle of the brain. In the s.c. and both i.c. xenografts models, in vivo localization of anti-carcinoma moab MOC-31 occurred within 4 hours after i.p. injection, with a maximal binding at 24 h after injection. A pronounced tumor infiltration of predominantly NK cells was observed for s.c. and intraventricular xenografts, but not for the GLC-28 tumors xenografted in the cerebral cortex. The presented nude rat/GLC-28 xenograft models may be used for the in vivo testing of experimental imaging techniques or alternative treatment strategies relevant to brain metastases of human SCLC.

Magnetite as a potent contrast-enhancing agent in magnetic resonance imaging to visualize blood-brain barrier disruption.

Imaging of blood-brain barrier damage by magnetic resonance was currently studied as to the potential of dextran-magnetite particles (DMP) for contrast enhancement. For that purpose, dextran-T10 (average molecular weight: 10.9 Kilodalton) was complexed with magnetite (Fe3O4) in ammonia. Experimental testing of the agent was made in vivo using Wistar rats with a freezing injury to the brain. DMP was i.v. injected at a dose of 90 microM Fe/kg b.w. followed by 2% Evans blue (0.6 ml). Control animals with trauma were studied without administration of DMP. Histochemical assessments were made to analyze the tissue distribution of DMP in the brain, kidney and liver after fixation in 4% formalin. MR imaging was conducted with 1.5 Tesla field strength with a circular coil 15 min after the freezing insult and administration of DMP. T1- and T2-weighted images were obtained using spin echo sequences among others. Regression analyses indicated a 50% reduction of T1 at a DMP concentration of 48 microM Fe, while for T2 only 4 microM/Fe(DMP) were sufficient for a 50% reduction. DMP was also accumulating in other organs, particularly in the Kupffer cells of the liver. Administration of DMP led to recognition of the freezing lesion as black area in agreement with macroscopical findings obtained by autopsy. In animals with a freezing lesion without administration of DMP, only T2-weighted images demonstrated a somewhat higher intensity attributable to the disruption of the blood-brain barrier. The present findings demonstrate the usefulness of DMP for contrast enhancement of lesions following disruption of the blood-brain barrier.

Dextran-magnetite particles: contrast-enhanced MRI of blood-brain barrier disruption in a rat model.

Dextran-magnetite particles (DMP) were studied for their use as a MR contrast agent to visualize lesions with a blood-brain barrier (bbb) disruption. A freezing injury to the rat cerebral cortex was used as a model of bbb disruption. The biodistribution of iv-injected DMP was studied using atomic absorption spectrophotometry, electron microscopy, and MRI. One hour after injection, focal accumulation of the particles in capillary endothelial cells could be demonstrated in the freezing lesion. Despite the observation that the relaxivity of DMP in vivo appears to be less well pronounced than that in vitro, the MR imaging studies show that DMP can be used to visualize bbb disruption with adequate contrast.

Qualitative and quantitative examination of rat and human fetal dopaminergic grafts.

This study was carried out as a prelude to possible implantations of cultured human fetal dopaminergic grafts in parkinsonian patients. Examination of fetal rat ventral mesencephalon tissue for morphology, viability, and dopamine content showed an optimal gestational age for neural grafting in rat experiments of approximately 17 days. Moreover, fetal rat ventral mesencephalon tissue was cultured, and neural dopaminergic cells were observed in cell culture in 4 (11%) out of 36 ventral mesencephalon specimens derived from 15- to 21-day-old rat fetuses. Human fetal donor material from elective abortions was examined for morphology and cell culture possibilities. In 48 curettements, fetal tissue was seen in 34 (71%) of these, resulting in 17 (50%) cultures containing dopaminergic cells. Both fetal rat and human cell cultures were continued for approximately 8 weeks and appeared to remain positive upon immunocytochemical dopamine staining.

An improved synthesis of carbon-11 labeled acetoacetic acid and an evaluation of its potential for the investigation of cerebral pathology by positron emission tomography.

1-11C-acetoacetic acid was synthesized by carboxylation of the acetone carbanion. Purification was carried out using HPLC. The product was obtained with a radiochemical yield of up to 58%, corrected for decay, in a total preparation time of 30 min. The distribution of 1-11C-acetoacetic acid after injection into adult Wistar rats and cats was investigated by PET. When the tracer was injected into cats, 3 weeks after inflicting a unilateral freezing lesion upon the brain, accumulation of 1-11C-acetoacetic acid in the ipsilateral brain hemisphere was observed.

Passage of DMP across a disrupted BBB in the context of antibody-mediated MR imaging of brain metastases.

To study the possible application of monoclonal antibody/dextran-magnetite conjugates in specific MR imaging of brain metastases, both components of these conjugates were tested for their ability to penetrate the endothelium during conditions of local blood-brain barrier (BBB) impairment. The passage of dextran-magnetite particles (DMP) across a disrupted blood-brain barrier was studied in a freezing lesion model using electron microscopy (EM) and MR imaging. One hour after i.v. injection, focal accumulation of DMP in capillary endothelial cells within the freezing lesion was shown by EM. In parallel with this, MR imaging indicated a strong contrast enhancement in the lesion. EM observations showed that the particles were still present in the endothelial cells four and eight hours after injection. The passage of an anti-small cell lung cancer (SCLC) monoclonal antibody across the endothelium of intracerebrally xenografted human SCLC was studied using immunohistological techniques. It was found that passage across endothelial cell occurred in the tumor within four hours after injection.

Positron emission tomographical studies of 1-11C-acetoacetate, 2-18F-fluoro-deoxy-D-glucose, and L-1-11C-tyrosine uptake by cat brain with an experimental lesion.

In cat brain with a freezing injury, the uptake of 1-11C-acetoacetate (11C-ACAC), 2-18F-fluorodeoxy-D-glucose (18FDG), and L-1-11C-tyrosine (11C-TYR) was monitored by positron emission tomography following intravenous administration of the tracers, at 1 day, and 1-3 weeks after the injury. The development and further course of the cold-induced oedema was monitored by magnetic resonance imaging. In the fresh (1 day old) lesion there was increased uptake of 11C-ACAC, probably due to release of the restrictive influence of the blood-brain barrier upon passage of the substance into brain. The uptake of 18FDG, which normally occurs by carrier-mediated transport at the barrier, was decreased in the fresh lesion, probably as a result of damage of the carrier mechanism. In the 3 week old lesion 18FDG uptake was still reduced, and 11C-ACAC uptake was still increased, although barrier function to Evans blue had recovered. It is suggested, that the increased 11C-ACAC uptake in the chronic lesion bears upon the proliferation of macrophages and reactive glial cells in the lesion. This is supported by the increased uptake of 11C-TYR in the 2 weeks old lesion, while in the fresh lesion 11C-TYR uptake was unchanged.

Cerebral cation shifts in hypoxic-ischemic brain damage are prevented by the sodium channel blocker tetrodotoxin.

We investigated the effect of the sodium channel blocker, tetrodotoxin, in two animal models of brain pathology. In the first, an acute model, we recorded the interstitial brain potential in the striatum of rats after cardiac arrest. The time of deflection of this potential, an indication of changes in cerebral cation concentrations, was determined in control rats, and in rats pretreated with intrastriatal tetrodotoxin. In control rats a deflection of the brain potential was noted 2 min after cardiac arrest; tetrodotoxin pretreatment delayed this deflection to about 5 min. The second, a survival model, was based on the Levine preparation in rats. A combination of ischemia and hypoxia produced unilateral, cerebral infarcts, which were characterized by a decrease of brain [K+], and by increases of [Ca2+] and [Na+] and thus of the Na+:K+ ratio. Data on the cation shifts, determined by chemical assay methods, were complemented by those of more conventional methods of assessment of brain damage, such as the determination of survival, of Evans blue staining, and of brain water content. Cation shifts could be prevented locally by tetrodotoxin. In conclusion, the drug can, at least partially, prevent the detrimental effects of an ischemic insult. In addition, our results showed that protective effects observed in the acute model may sometimes offer an indication of the effects to be expected in the survival model. Furthermore, the effect of tetrodotoxin on the brain potentials in the acute model showed that its protective action in the survival model may be brought about by delaying cell depolarization and by shortening the actual duration of the depolarized state. We conclude that Na+ influx and, consequently, neurotransmission may play a crucial role in the development of cerebral damage.

Changes of ventricular ependyma and choroid plexus in experimental hydrocephalus, as observed by scanning electron microscopy.

Hydrocephalus was induced in rats by the injection of silicone oil or kaolin suspension into the cisterna magna. One to 5 weeks later the walls of the lateral ventricles were studied with the scanning electron microscope after killing the animals by perfusion fixation. In contrast to controls, the hydrocephalic animals killed 1 or 2 weeks after injection showed degeneration of ependymal cilia and infestation of the ependymal and choroid plexus surface with reactive cells, which presumably may be identified as Kolmer phagocytic cells by their ultrastructural features as studied by the transmission electron microscope. A coating of debris on the surface of the choroid plexus in the hydrocephalic animals possibly bears upon the ciliary degeneration with consequent deficiency of the clearing effect of ciliary movement. In the longer surviving hydrocephalic animals regeneration of cilia seemed to have occurred.