Irena Westergren

Intracerebral dialysis and the blood-brain barrier.

Abstract: The aim of the study was to evaluate how implantation of a dialysis probe influences the blood‐brain barrier. Leakage of endogenous serum albumin was evaluated by Evans blue/albumin staining and by immunohistochemistry. The passage from blood to dialysate of two substances that normally do not pass into the brain, [3H]inulin and glutamate, was studied 3 and 24 h after insertion of a dialysis probe. Evans blue, given 20 min before rats were killed, was observed around the probe and surrounding brain tissue. Albumin immunoreactivity was seen at considerable distance from the probe with larger spread at 24 h than at 3 h after probe insertion. Glutamate and [3H]inulin were detected in the dialysate with no significant further increase of radioactivity after intracarotid infusion of protamine sulfate that enhances the permeability over the blood‐brain barrier. When protamine was followed by infusion of glutamate, the concentrations of taurine increased in the dialysate in four of eight rats. That plasma constituents have access to the brain around the dialysis probe is essential to consider, particularly in studies using substances and drugs that do not pass an intact blood‐brain barrier.

Concentrations of amino acids in extracellular fluid after opening of the blood-brain barrier by intracarotid infusion of protamine sulfate

Abstract: This article evaluates the influence of an opening of the blood‐brain barrier (BBB) on compounds in brain extracellular fluid. The concentrations of amino acids and some other primary amines were determined in dialysates sampled from the right parietal cortex of rats before and after an intracarotid infusion of protamine sulfate. Extravasated plasma proteins were visualized by Evans blue/albumin and immunohistochemistry. CSF albumin— an indicator of blood‐CSF barrier opening—was quantified with immunoelectrophoresis. The brains were macroscopically edematous after 10 mg but not after 5 mg of protamine sulfate. The higher dose led to a 50% death rate. The concentrations of amino acids did not change 10 min after the BBB opening. No significant alterations in the amino acid concentrations were observed after the lower dose. The concentrations of glutamate, aspartate, GABA, glycine, taurine, and phosphoethanolamine increased significantly within 50–80 min after the infusion of 10 mg of protamine sulfate. CSF albumin levels were significantly increased 1 h after infusion. We conclude that a dysfunction of the BBB, of a degree known to induce brain edema (10 mg of protamine sulfate), significantly increases the extracellular concentration of excitatory amino acids, GABA, taurine, and phosphoethanolamine in the extracellular space.

NBQX, an AMPA antagonist, reduces glutamate-mediated brain edema.

Glutamate (2.5 mg) was administered after the blood-brain barrier had been opened by a unilateral intracarotid infusion of 5 mg protamine sulfate in rats. Whereas the brain specific gravity, measured 24 h later, did not differ between the injected and non-injected side after protamine alone, glutamate significantly reduced the specific gravity in the right hemisphere indicating brain edema (P less than 0.01). NBQX, a potent AMPA receptor antagonist, significantly reduced the edema (P less than 0.01) and completely inhibited the glutamate mediated increase in albumin content in cerebrospinal fluid (P less than 0.01).

Blood-brain barrier leakage and brain edema in stroke-prone spontaneously hypertensive rats. Effect of chronic sympathectomy and low protein/high salt diet

SummaryBrain edema associated with severe chronic hypertension was studied in stroke-prone spontaneously neously hypertensive rats (SHRSP), 5 to 9 months of age. Blood-brain barrier (BBB) leakage sites and intracerebral spreading pathways for plasma proteins were delineated by an intravenously (i.v.) injected exogenous dye tracer (Evans blue), known to form a complex with albumin in blood, and by immunohistochemical visualization of extravasated endogenous plasma proteins. The tissue content of edema fluid was estimated by measuring the specific gravity of selected brain regions, stained or unstained by the tracer dye, on a bromobenzene-kerosene gradient column. Multifocal BBB leakage sites were macroscopically detected within the cerebral cortex and the deep gray matter after i. v. circulation of Evans blue-albumin for 30 min. After 24 h of i.v. circulation the dye tracer had spread not only locally in the gray matter but also into the adjacent white matter, where it was widely distributed. Immunohistochemically visualized plasma proteins showed similar distribution. Unilateral superior cervical ganglionectomy performed at 4 weeks of age neither increased the incidence of major BBB opening to Evans blue-albumin nor altered the specific gravity of the ipsilateral cerebral hemisphere in grown-up SHRSP, furthermore, the blood pressure remained unchanged. The lack of significant effect on BBB function may possibly be attributed to the extensive reinnervation of the cerebral arteries, verified in the grown-up SHRSP using the Falck-Hillarp fluorescence method for visualization catecholaminergic nerve fibers. In SHRSP raised on a low-protein and high-salt diet the mean arterial blood pressure was 212 mm Hg compared to 195 mm Hg in controls (P<0.05) and the incidence of BBB opening was 72% compared to 25% in controls (P<0.05). After 24 h of i.v. circulation of Evans blue-albumin, brain regions stained by the dye tracer showed significantly reduced specific gravity (P<0.001), while unstained regions had normal values. Thus the brain edema fluid spread, as revealed by specific gravity measurements, corresponded to the intracerebral distribution of extravasated plasma proteins.

Neuropeptides in Cerebrospinal Fluid in Normal-Pressure Hydrocephalus and Dementia

Delta-sleep-inducing peptide (DSIP), vasoactive intestinal peptide (VIP), peptide YY (PYY) and somatostatin (SOM) were assayed with specific radioimmunological methods in cerebrospinal fluid (CSF) of healthy volunteers, 12 patients with Alzheimers disease (AD), 11 patients with multi-infarct dementia (MID) and 10 patients with normal-pressure hydrocephalus (NPH). Patients with NPH were reinvestigated 3 months after a ventriculoperitoneal shunt operation. DSIP, PYY and SOM levels in CSF were decreased in patients with NPH compared to controls. The CSF concentration of SOM was also significantly reduced in patients with AD. No correlations were found between the degree of dementia in any of the illnesses and the CSF concentrations of the peptides. The concentration of DSIP, VIP and SOM increased significantly in parallel to the clinical improvement after the shunt operation in NPH patients.

Changes in physiological parameters of rat cerebrospinal fluid during chronic sampling: Evaluation of two sampling methods

Although rat cerebrospinal fluid (CSF) is increasingly being used in pharmacological and biochemical research, methodological studies on basic physiological data are lacking. We have determined the albumin content and number of erythrocytes and leukocytes in CSF obtained by two different methods of sampling from cisterna magna-repeated sampling from an implanted cannula and by repeated punctures. In the initial samples the albumin content was 0.08 +/- 0.03 micrograms/microliters. Chronic cannulation of the cisterna magna resulted in a meningeal reaction with increased cell and albumin content: a reaction that could be reduced but not prevented by using a sterile cannula. The number of leukocytes but not erythrocytes was highly correlated to the albumin content. Repeated sampling in the absence of a permanent cannula did not significantly increase albumin content but carried a higher risk for erythrocyte contamination.

Blockade of AMPA Receptors Reduces Brain Edema following Opening of the Blood—Brain Barrier

The aim of our study was to evaluate whether blockade of α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) receptors could reduce brain edema in two experimental models of edema following opening of the blood–brain barrier (BBB). The brain specific gravity was determined 2 h after opening the BBB by a 30-s infusion of protamine sulfate (10 mg in 200 μl 0.9% NaCl) or arabinose (1.5 or 1.8 mol/L, 0.06 ml · s−1) into the right internal carotid artery. Cisternal CSF was withdrawn for albumin determination before the carotid infusion and before killing 2 h later. After infusion of protamine sulfate or arabinose, CSF albumin increased in all groups. The brain specific gravity was significantly lower in the right than in the left (control) frontal, parietal, and occipital cortex and striatum. NBQX (2,3-dihydroxy-6-nitro-7-sulfamoylbenxo(F)quinoxaline), an AMPA receptor antagonist, given intravenously 10 min after opening the BBB (5 mg/kg), significantly increased the specific gravity in the treated rats (p < 0.01 for the difference from control rats) without reducing CSF albumin or albumin extravasation in the brain as evaluated with Evans blue. We hypothesize that intracerebral (glial?) AMPA receptors may play a role in brain edema following opening of the BBB.

Albumin content in brain and CSF after intracarotid infusion of protamine sulfate: A longitudinal study

The endogenous serum albumin content was determined by immunoelectrophoresis in brain and cisternal CSF 1, 24, and 72 h after a transient opening of the blood-brain barrier. Protamine sulfate, 5 mg in 100 microliters 0.9% NaCl, was infused during 30 s into the internal carotid artery via a catheter in the external carotid artery in conscious rats. The albumin content in CSF was 0.08 +/- 0.03 g/liter before protamine infusion and 0.09 +/- 0.02 g/liter in rats infused with saline only. The levels were significantly increased one and 24 h after protamine infusion (0.37 +/- 0.19 and 0.23 +/- 0.09 g/liter, P less than 0.001) but not at 72 h (0.14 +/- 0.05 g/liter). The albumin content in the right (injected) hemisphere decreased with time but was significantly higher than that in the left hemisphere at all times (P less than 0.001 1 and 24 h after protamine; P less than 0.01 at 72 h). There was no correlation between the albumin contents in brain and CSF. Pretreatment with dixyrazine, a phenothiazine derivate, significantly reduced the protamine-induced leakage of endogenous serum albumin into brain and CSF.

Glutamate enhances brain damage and albumin content in cerebrospinal fluid after intracarotid protamine infusion.

SummaryThe blood-brain barrier was opened by intracarotid infusion of 5 mg protamine sulfate in 100 μl 0.9% NaCl over a period of 30 s either alone or followed by infusion of 10 mg L-glutamate in 0.9% NaCl. Glutamate alone was infused in four control rats. Cisternal cerebrospinal fluid (CSF) was withdrawn before protamine administration and before the brains were fixed in situ 1, 24 or 72 h later. The albumin extravasation and glial reactivity was evaluated by immunohistochemistry on paraffin sections. The CSF albumin was significantly increased in both the protamine and protamine/glutamate groups but remained high at 24 and 72 h in the protamine/glutamate group only. Spongiotic lesions with shrunken nerve cells were observed 1 and 24 h after infusion of protamine alone or in combination with glutamate. Whereas such changes were not seen 72 h after protamine infusion, they remained and had progressed at 72 h in the protamine/glutamate group, indicating that glutamate induces delayed cellular damage when given access to the brain through an altered blood-brain barrier.

Tirilazad Reduces Brain Edema After Middle Cerebral Artery Ligation in Hypertensive Rats

Tirilazad (3 mg/kg i.v.) or vehicle was given 10 min, 3 and 12 hours after ligation of the right middle cerebral artery in male spontaneously hypertensive rats (n = 12 in each group). Brain specific gravity was determined in 23 predetermined cortical regions covering the core and penumbra areas 24 hours after the occlusion. The specific gravity was significantly higher in tirilazad-treated rats than in controls (p < 0.0001). When individual regions of the two groups were compared, the difference was significant in 9 of the 23 samples predominantly, but not exclusively, in the penumbra zone.