Evans Blue Injection Research Articles

Recombinant or plasma-derived antisecretory factor inhibits cholera toxin-induced increase in Evans blue permeation of rat intestinal capillaries.

The effect of cholera toxin on small intestinal capillary function, utilizing the Evans blue dye method, was analyzed. The modulatory influence of plasma-derived or recombinant human antisecretory factor on this variable was also investigated. Male Sprague-Dawley rats were briefly anesthetized with ether, and a jejunal loop was constructed that was challenged for 90 min with phosphate-buffered saline or cholera toxin. Five minutes prior to death, the rats received an intravenous injection of Evans blue. The tissue content of dye in the loop was quantitated spectrophotometrically or demonstrated histochemically. Cholera toxin increased the recovery of Evans blue; extravasation of the dye was prominent in the top of the villi, while the crypts were spared. It is suggested that the toxin caused increased transcapillary permeation of albumin in a heterogenous fashion in the gut wall. This effect of the toxin was prevented by pretreatment with the antisecretory factor.

Increased gastric bicarbonate secretion in portal hypertensive anesthetized rats: role of prostaglandins and nitric oxide.

Gastric bicarbonate secretion might be modified in portal hypertension as a consequence of the intramucosal increase in prostaglandins and nitric oxide content. Therefore, we studied gastric bicarbonate secretion in control and portal hypertensive rats and investigated the role of prostaglandins and nitric oxide. Basal gastric bicarbonate secretion was studied in rats, using a gastric pH back-titration technique, two weeks after partial portal vein ligation or a sham operation. The effects of the following drugs were investigated: the prostaglandin synthase inhibitor indomethacin (5 mg/kg intravenously), prostaglandin (PGE2) (1 mg/kg intravenously), the nitric oxide synthase inhibitors N(G)-nitro-L-arginine methyl ester (L-NAME, 5 mg/kg intravenously) and N(G)-monomethyl-L-arginine (L-NMMA, 50 mg/kg intravenously), and the nitric oxide donor nitroprusside (5 mmol/liter in the gastric perfusate). Plasma leakage in the gastric wall was also measured after Evans blue dye injection in portal hypertensive and sham-operated rats pretreated by indomethacin (5 mg/kg, intravenously) and L-NAME (5 mg/kg, intravenously). Basal bicarbonate secretion was significantly increased in portal hypertensive rats as compared to controls. After indomethacin, the bicarbonate secretion was significantly reduced to a similar level in both groups. PGE2 increased bicarbonate secretion significantly more in portal hypertensive rats than in sham-operated rats. The NO synthase inhibitor L-NMMA significantly increased bicarbonate secretion in portal hypertensive rats only, while the other inhibitor, L-NAME, increased it significantly more in portal hypertensive than in the sham-operated rats. Plasma leakage in portal hypertensive rats, which was increased in the basal condition as compared to control, was further enhanced by indomethacin but not by L-NAME pretreatment. The nitric oxide donor significantly reduced bicarbonate secretion in portal hypertensive rats to reach a similar level as in sham-operated rats. Basal gastric bicarbonate secretion is increased in portal hypertensive rats. This could be due to an enhanced prostaglandin mucosal level. Nitric oxide, which reduces bicarbonate secretion, may contribute to limiting prostaglandin-induced bicarbonate overproduction.

Brain mast cell degranulation regulates blood-brain barrier.

Mast cells synthesize vasoactive agents and a number of neurotransmitters. They are particularly numerous in the medial habenular region of the epithalamus, the attachment site of the choroid plexus. The present study examined whether degranulation of brain mast cells alters the permeability of the blood-brain barrier (BBB). To this end, doves were injected intramuscularly with the mast cell degranulator, compound 48/80 (C40/80), followed by i.v. injection of Evans blue. The distribution of the dye in the parenchyma was examined using digital imaging. Three brain areas were analyzed: the medial habenula (which also contains mast cells), the paraventricular nucleus (PVN, which abuts the third ventricle, but has no mast cells), and the lateral septal organ (LSO, a circumventricular organ with fenestrated capillaries). Significantly more Evans blue tracer and fewer toluidine blue-positive mast cells were detected in the medial habenula of subjects treated with C48/80 compared to saline controls. Evans blue did not enter the PVN in either the experimental or control group, while it entered the LSO equally in both. Degranulation of mast cells after C48/80 treatment was confirmed histochemically and ultrastructurally. The results support the hypothesis that brain mast cell degranulation locally alters BBB permeability. Activation of brain mast cells may provide a mechanism for regulated opening of the BBB.

Accuracy of an experimental stereotactic system for MRI-based gamma knife irradiation in the rat.

Stereotactic devices for experimental Gamma Knife irradiation and magnetic resonance imaging (MRI) have recently been developed for experimental studies using rats [6,7]. The present study examined the accuracy of these devices using the following two approaches. In the first approach, Gamma Knife irradiation was performed using the stereotactic device with targets based on a standard stereotactic atlas. Gadolinium-enhanced T1-weighted magnetic resonance imaging was performed using the MRI stereotactic device. Animals were then sacrificed after Evans blue injection, and the rat brain was sliced using an attachment to the stereotactic device. The center coordinates of the gadolinium-enhanced area from the MRI and Evans blue-stained area from the tissue sections were obtained using a computer-assisted image analysis system. These coordinates were compared with the target coordinates planned from the stereotactic atlas. In the second approach, a thermoluminescence dosimeter was implanted in the rat brain. Stereotactic MRI was performed using the stereotactic MRI device, and the coordinates of the implant were obtained. Gamma Knife irradiation was then performed at this target using the stereotactic device. The absorbed dose was measured and compared with the planning dose. These experiments demonstrated a spatial error of 0.6 mm (standard error +/- 0.07) between Gamma Knife irradiation based on a comparison of the atlas coordinates and the lesion, and a spatial error of 1.0 mm (standard error +/- 0.13) based on a comparison of the stereotactic MR images and the lesion. Gamma Knife irradiation based on MR images using the stereotactic device demonstrated a maximum error of 10% in absorbed dose at the target center. Together, the stereotactic devices for Gamma Knife irradiation and magnetic resonance imaging provide useful tools for Gamma Knife research in an animal model.

Hepatic local disposition of a drug with high protein binding and high hepatic clearance using BOF-4272 as a model drug.

The local disposition of a drug, which is efficiently eliminated by the liver in spite of very high binding with serum albumin, was investigated using BOF-4272 as a model drug in the hepatic perfusion system. Bovine serum albumin (BSA) labeled with Evans Blue was used as the marker of hepatic blood space. The perfusion experiments were carried out at 37 degrees C and at 4 degrees C to evaluate the local disposition in active and inactive livers, respectively. The perfusates included 0, 0.25, 0.5, 1.0 and 4.0% of BSA in the injection of BOF-4272, and 0 and 1.0% of BSA in the injection of Evans Blue. After an instantaneous injection of BOF-4272 or Evans Blue (labeling reagent of BSA), the outflow time profile from the liver was analyzed by curve-fitting based on two-compartment dispersion model. All estimated parameters of BSA were almost the same between 37 degrees C and 4 degrees C, which showed that the blood space in the liver was unaffected by perfusate temperature. The recovery ratio (FH) and the mean transit time (FH) of BSA were 100% and about 0.1 min, respectively, both with BSA (1.0%) and without BSA in the perfusate. FH of BOF-4272 increased from 20% to 50% with an increase in the perfusate BSA at 37 degrees C, whereas FH was almost constant (90%) regardless of BSA concentration at 4 degrees C. FH of BOF-4272 was almost 0.1 min regardless of BSA concentration at 37 degrees C, whereas FH decreased from 0.29 min to 0.17 min at 4 degrees C with an increase in the perfusate BSA. A large elimination (10%) of BOF-4272 was noticed even at 4 degrees C in the presence of perfusate BSA, which demonstrates that the passive transfer of BOF-4272 from BSA to the hepatic tissues is considerably rapid. This efficient transfer is responsible for the large hepatic clearance of BOF-4272 with very high binding with BSA.

Contribution of Peripheral Macrophages and Microglia to the Cellular Reaction after Mechanical or Neurotoxin-Induced Lesions of the Rat Brain

Lesions of the central nervous system result in invasion of peripheral phagocytes and/or in situ activation and proliferation of microglia, depending on the direct or indirect nature of the injury. Neurotoxins which are widely used to induce neurodegeneration have been reported to elicit a pure microglial reaction when administered intraventricularly. However, the mechanical lesion at the injection site, although remote from the lesioned area, could give access to blood-derived cells. Therefore, this study compares the phagocytic reaction occurring after a traumatic lesion of the brain causing a breakdown of the blood-brain barrier (BBB), or after degeneration of the inferior olivary neurons induced by intraperitoneal administration of 3-acetylpyridine, a process that respects the integrity of the BBB as suggested by the results of intravascular injection of Evans blue. The identification of the macrophages at the lesion site used specific binding of the B 4 isolectin from Griffonia simplicifolia, preloading of the peripheral monocytes/macrophages with trypan blue, and characteristic morphological features. In traumatically lesioned rats, the lectin-labeled macrophages were mainly large rounded peripheral cells recruited 1-3 days postlesion, whereas in chemically lesioned rats, the cellular reaction appeared 24-36 h postinjection and peaked between 3 and 12 days before undergoing a slow decline. Lectin binding and morphological characteristics indicated that these small cells were reactive microglia. These results confirm that a brain injury leaving the BBB intact involves essentially the recruitment and/or the proliferation of microglia.

Suppressive effect of human blood coagulation factor XIII on the vascular permeability induced by anti-guinea pig endothelial cell antiserum in guinea pigs

We investigated the effect of blood coagulation factor XIII(FXIII) on enhanced permeability induced by anti-endothelial cell antiserum, that was produced by the immunization of guinea pig endothelial cells with adjuvant into rabbits repeatedly. We have found that this antiserum reacts to human and guinea pig endothelial cells but not guinea pig fibroblast cells. The permeability was enhanced by intradermal injection of 400-fold dilution of this antiserum into dorsal skin of guinea pigs. The mixture of equal volume of antiserum and FXIII was intradermally injected into dorsal skin of guinea pig after Evans blue injection, and 15 minutes later the quantity of Evans blue at the each injection site was determined. We recognized the suppressive effect of FXIII on the dye leakage. We also studied the suppressive effect on swelling induced by the antiserum. After the subcutaneous injection of the mixture of antiserum and FXIII into the back of guinea pigs, we measured the thickness of skins at the injection site after day 1, 2 and 3. As a result, FXIII significantly suppressed the swelling. We found that FXIII suppresses the acute and subacute permeability enhancement. These results suggest that FXIII plays an important role on an inflammatory site and that it may exert as an anti-inflammatory protein.

Correlative Study of the Morphology and Central Connections of Ipsilaterally Projecting Retinal Ganglion Cells in the Albino Rat

Injection of Evans Blue (EB) and 4′,6-diamidino-2-phenylindole dihydrochloride (DAPI), fluorescent dyes, into the dorsal nucleus of the lateral geniculate body (dLGN) and the ipsilateral superior colliculus (SC) of albino rats, retrogradely labelled retinal ganglion cells (RGCs). We classified these cells into three categories: (1) double-labelled, geniculate and collicular-projecting RGCs; (2) single-labelled, geniculate-projecting RGCs; and (3) single-labelled, collicular-projecting RGCs. Cell soma sizes were measured and the soma size histogram of each category was compared with the others as well as with that of the corresponding category in the contralateral retina. The mean soma size of the ipsilaterally projecting double-labelled RGCs was larger, and their soma size histogram was significantly different in comparison with those of the other two categories. On the contralateral side, the mean soma size of the RGCs labelled only from the SC was smaller, and also their soma size histogram was significantly different from those of the other two categories. On comparing both sides, only the ipsilaterally-projecting double-labelled cells had a significantly different soma size histogram and larger soma size.

Effects of topical FK506 on experimental allergic conjunctivitis

In vitro studies of FK506 demonstrated that the agent inhibited the T-cell receptormediated signal transduction that results in the transcription of interleukin 2. Recent in vitro studies have demonstrated an identical spectrum of activities of FK506 on IgE receptor-mediated signal transduction that results in exocytosis of secretory granules from basophils and mast cells. The effects of topical FK506 on allergic conjunctivitis were therefore studied in the rat. Passive anaphylaxis was induced in Wistar rats by injecting anti-ovalbumin IgE at the conjunctiva followed by intravenous injection of antigen (ovalbumin) and Evans blue. Topical FK506 (0.03-0.1%) significantly suppressed the passive anaphylaxis in the rat conjunctiva when given at 15 and five minutes, or six, four and two hours before the antigen challenge. The efficacy was more intense than that of 0.1% betamethasone or 2% disodium cromoglycate. These data thus suggest that topical FK506 might be beneficial for the therapy of severe allergic diseases in the eye.

Microendoscopic observations of the hemodynamics in the rabbit ciliary processes.

The in vivo hemodynamics of the ciliary process vasculature in the albino rabbit were studied with an improved technique of intraocular microendoscopy. The spreading pattern and velocity of the dye front has been observed in defined vessel segments following intracarotid injection of Evans Blue. In the anterior portion of the ciliary processes a "thoroughfare channel" (bypass) was found leading from the arteriolar tree directly into the marginal venule bypassing the capillary network of the ciliary processes. This bypass is characterized by a higher blood flow velocity (1.5-3mm/sec) than was found for the capillary network (0.8-1mm/sec). A stoppage of blood flow was observed in the capillary network at 45-50mmHg after step-wise elevation of intraocular pressure (IOP): blood flow was not stopped in the thoroughfare channel until IOP values of 55-60mmHg were reached. Intraarterial administration of vasoconstrictive agents could lead to a complete stoppage of blood flow in the capillary net whereas the marginal route often remained patent.

Intravenous injection of evans blue labels magnocellular neuroendocrine cells of the rat supraoptic nucleus in situ and after dissociation

Previous work has demonstrated that intravenous injection of neuronal tracers, e.g. horseradish peroxidase or Fast Blue, can retrogradely label neurons in brain areas that project outside the blood-brain barrier, e.g. magnocellular neuroendocrine neurons of the hypothalamus. Here we have shown that 24 h after intravenous injection of the fluorescent retrograde tracer Evans Blue, the same population of magnocellular neuroendocrine neurons is labeled in the paraventricular, supraoptic and accessory magnocellular nuclei. Parvicellular neuroendocrine cells in the paraventricular nuclei are also labeled. Most Evans Blue-labeled magnocellular neuroendocrine cells in the supraoptic nucleus could be stained immunocytochemically for neurophysins, suggesting that these neurons continue to produce their peptide hormones after taking up the fluorescent dye. Ultrastructural observation of supraoptic cells retrogradely labeled with Evans Blue shows that 95% of the neurons appeared healthy. There was no ultrastructural evidence of degeneration, hyperstimulation, or interruption of the axoplasmic flow. Labeling the neuroendocrine cells with Evans Blue did not alter the size of magnocellular cells, the animal's fluid balance or ingestive behavior. Following enzymatic/mechanical dissociation of the supraoptic nucleus from animals that had been injected with Evans Blue 24 h previously, phase-bright neurons that often contained fluorescent material were observed, thus identifying these neurons as neuroendocrine. Recording from identified neuroendocrine cells showed that these neurons generated spontaneous or currentevoked overshooting action potentials with an afterhyperpolarization and had negative resting membrane potentials. Action potential broadening, a feature of magnocellular neurons, was observed during bursts of action potentials elicited by depolarizing current injection. Taken together, this work would suggest that Evans Blue is non-toxic at the doses used and that it provides a method to identify single neuroendocrine cells in primary cell cultures made from adult hypothalamus for voltage-clamp recordings.

Effects of glucocorticoid and chemotherapy on the peritumoral edema and astrocytic reaction in experimental brain tumor.

To study the effects of glucocorticoids and chemotherapeutic agents on the pathophysiology of the tumor-induced brain edema, the site of Evans blue-albumin extravasation, the distribution of extravasated serum albumin, and the extent of local astrocytic reaction were examined in a rat model of implanted brain tumor. Experimental brain tumors were produced by implanting small pellets of Walker 256 carcinosarcoma into the cerebral cortex of Wistar rats. In the steroid group, rats were administered with intraperitoneal methylprednisolone succinate (15 mg/kg) daily on and after the 6th day postimplantation, and sacrificed on the 14th day. In the chemotherapy group, rats were given an intravenous injection of cyclophosphamide (30 mg/kg) on the 14th day, and sacrificed on the 21st day. Rats in the untreated group were sacrificed on the 14th day without any therapy. Each animal was sacrificed by the transcardiac perfusion with paraformaldehyde 30 min after intravenous injection of Evans blue. Firstly, coronal blocks of the brain were examined for Evans blue staining macroscopically. Paraffin embedded sections were studied for the Evans blue fluorescence and for the immunohistochemical reaction to serum albumin and GFAP. The examination of Evans blue demonstrated that the origin of extravasation of serum albumin was the tumor and the adjacent brain with dense tumor cell infiltration in any group of rats. The extravasated serum albumin distributed widely and the astrocytic reaction was prominent in the brain of the untreated group. A positive correlation was observed between the intensity of albumin immunoreaction and the degree of astrocytic proliferation. Chemotherapy effectively decreased the size of tumor and reduced the extravasation of serum albumin. The astrocytic reaction was however, not reduced.(ABSTRACT TRUNCATED AT 250 WORDS)

実験的脳梗塞に対するbifemelane hydrochlorideの効果

Bifemelane hydrochloride is a new substance which exhibits antianoxic and memory retrieval effects in cases of ischemic cerebral damage. Recent clinical trials have also revealed favorable effects on vascular dementia.This study was designed to comfirm neuropathologically the effects of this substance on ischemic changes in the brains of Mongolian gerbils caused by ligated left carotid arteries. Two-hundred 8-week-old Mongolian gerbils were divided into 4 groups of 50 animals each. Bifemelane hydrochloride was administered intraperitoneally to the animals in 1.0% physiological saline at doses of 25mg/kg body weight. Bifemelane hydrochloride was given to group II 30 minutes before, and twice to group III 30 minutes before and after the ligation procedure. Group I served as control. This group was administered intraperitoneally the same volume of physiological saline as bifemelane hydrochloride solution used for animals with the same body weight. Group IV was prepared for an observation of vascular permeability after administering bifemelane hydrochloride by intravenous injection of Evans blue. All the animals were sacrificed 3 hours after the ligation procedure by perfusion fixation and were observed with a microscope.The survival rates and the neurological deficits did not vary significantly between groups. The incidence of prominent neurological changes in the brains of group I, II and III were 62, 54 and 54% respectively. Group I displayed a generalized reduction of stainability and widely dispersed severe ischemic lesions with a spongy state and edema, especially, in white matter and the caudate nucleus. These morphological changes expanded out from periventricular areas, and were demarcated from deeply stained normal tissues.Hippocampus samples from group I subjects showed decreased neuronal cells, vacuolar degeneration of these cells and irregular dendritic trees. Electron microscopic observations of group I specimens revealed degeneration of endothelial cells, intracellular edema, and swelling of neuronal mitochondria. Histopathological changes of group II and III subjects administered bifemelane hydrochloride were generally not serious, and boundary areas of the ischemic lesions of these subjects appeared to be intermingled with normal histological figures. Electron microscopic examination of group II and III subjects revealed that a considerable amount of organelles, such as mitochondria, microtubules, synaptic vesicles, and membranes of neuronal cells, kept their ordinal structures in the intermingling state. Evans blue transdated slightly to ischemic lesions of the animals administered bifemelane hydrochloride. Neuropathological changes observed in group I were somewhat similar to those in the brains of patients with vascular dementia; therefore, slight changes manifested in groups II and III should be regarded as evidence suggesting the utility of bifemelane hydrochloride in treating vascular dementia.

Blood flow and blood volume determinations in aorta and in coronary circulation by density dilution.

Continuous blood mass-density measurements were performed in anesthetized dogs and injections of 0.7-1.4 ml/kg isotonic saline solution were applied. The resulting density dilution curves were used to compute blood volume, total flow in the aorta and local flow in the coronary circulation. Blood volume calculations were compared with blood volume determined by Evans blue injections and a close agreement was found. Blood flow determined by density dilution was independent from the investigated sites of injection or sampling. We conclude from these results that small volume injections of isotonic saline solution can be used to determine blood volume and flow by density dilution. In addition to these findings, a marked retention of the injected fluid was observed. Possible mechanisms to explain this retention include albumin deposition in the endothelial pores and/or variations of blood viscosity and capillary pressure.

Cerebral microangiopathy in stroke-prone spontaneously hypertensive rats. An immunohistochemical and ultrastructural study.

The morphology of cerebral microvessels was studied immunohistochemically and ultrastructurally in 6- to 9-month-old normotensive Wistar-Kyoto rats (WKY), spontaneously hypertensive rats (SHR), and stroke-prone SHR (SHRSP) with a systolic blood pressure of 138 +/- 15 mm Hg, 189 +/- 9 mm Hg, and 258 +/- 30 mm Hg respectively. Regions with major opening of the blood-brain barrier (BBB) were revealed by an i.v. injection of Evans Blue. Multifocal BBB opening with massive leakage of plasma constituents rich in fibrinogen-fibrin-related antigen occurred in SHRSP with a blood pressure above 210-220 mm Hg. BBB-leakage sites were found in the cerebral cortex and the basal ganglia, most frequently in the arterial border zones. The perivascular tissue spaces were dilated within the BBB-leakage sites, in particular around arterioles. Damaged endothelial and smooth muscle cells were replaced by fibrin-like material, multiple layers of basement membranes and bundles of collagen fibrils surrounded by proliferated fibroblasts. The degenerative-infiltrative-proliferative disease process transformed short segments of single arterioles into severely thickened, tortuous and stenotic vessels. Fibrinoid degeneration, formation of microaneurysms and fibrin-rich vascular occlusions were observed. In contrast, only minor or no vascular alterations were seen in regions with preserved BBB in SHRSP and SHR. A severely increased intraluminal pressure load appears to be of major pathogenetic importance for breakdown of the BBB and initiation of the vascular disease process in SHRSP. However, since only short segments of a limited number of widely separated vessels are severely affected, and the number of affected vessels increase towards arterial end and border zones, additional predisposing and aggravating factors may play significant roles in the development of fibrinoid vascular lesions in arterial hypertension.

NMR Imaging and Spectroscopy of the Mammalian Central Nervous System after Heavy Ion Radiation

NMR imaging, NMR spectroscopy, and histopathologic techniques were used to study the proton relaxation time and related biochemical changes in the rodent brain after in vivo helium beam irradiation with single doses of 10, 20, 30, and 50 Gy. Two-dimensional Fourier transform spin-echo imaging and saturation recovery with projection reconstruction were used to measure the NMR relaxation parameters. These parameters were correlated with proton spectroscopy and histopathology. Additional high resolution in vitro proton spectroscopy was performed on brain extracts to observe chemical changes that could not be seen in vivo. The major findings from these experiments were that at 4-14 days postirradiation, image intensity and T1 relaxation time decreased on the irradiated side and increased on the nonirradiated side relative to nonirradiated control animals. In vivo surface coil proton spectroscopy methods demonstrated changes in lipid and phosphatidylcholine (p-choline) peaks. In vitro studies of the aqueous fraction of brain extracts showed radiation-induced changes in lactate, 4-aminobutyric acid, and p-choline peak areas. In the organic fraction, radiation-induced changes were observed in phosphatidylcholine, phosphatidylethanolamine, and phosphatidylserine. With histology and Evans blue injections, blood-brain barrier alterations were seen as early as 4 days after a dose of 50 Gy.

FC fragments of human IGG may influence allergic reactions

The antiallergic effect of Fc fragments prepared from human polyclonal IgG was investigated in several experimental models. In an in vitro assay, isolated ileum of cynomolgus monkeys was sensitized with serum from atopic patients. In six of fifteen monkeys the subsequent addition of specific allergens reproducibly resulted in an ileum contraction measured in the Schultz - Dale apparatus. In all six positive monkeys pre-treatment of the isolated ileum with Fc (papain) before sensitization inhibited ileum contraction. Fc (plasmin) or Fc (pepsin) was less or not effective, aggregated IgG, monomeric IgG, sulfonated IgG, the Fc-free F(ab') 2 moiety, or albumin had no significant or no reproducible inhibitory effect. In an in vivo assay passive cutaneous anaphylaxis was studied in 28 cynomolgus monkeys. Different dilutions in PBS of the particular specific allergens were subsequently administered to sensitized skin sites, simultaneously to i.v. injection of Evans blue. The degree of the local allergic (passive cutaneous anaphylactia) reaction was evaluated by measuring the diameter of the blue area at the dermal injection sites. About 50% of sera induced positive reaction in monkeys. Compared to the control (application of PBS), the degree of the positive reactions could be reduced by about 50% if Fc (papain) (optimal dose 25 mg/kg body weight) was injected i.v. either 2 h before or after local sensitization. No significant inhibitory effect could be found with the Fab moiety of IgG. Follow-up studies showed that the inhibitory effect of Fc lasted for about 10 days. The pharmacological basis of the observed antiallergic action of Fc is not yet understood.

Does neurogenic inflammation alter the sensitivity of unmyelinated nociceptors in the rat?

This study on neurogenic inflammation aimed at comparing the effects of antidromic nerve stimulation and of cutaneous application of mustard oil on the mechanical and thermal sensitivity of cutaneous C-fibres in the rat saphenous nerve. They were tested with von Frey hairs and series of radiant heat stimuli in 5-min intervals before and after one of the above treatments. Antidromic electrical stimulation was effective in evoking a plasma extravasation in the saphenous region as revealed by prior intravenous injection of Evans blue. However, it did not evoke spontaneous activity in the polymodal C-fibres tested nor did it markedly affect their mechanical and thermal excitability in periods of up to 1 h after stimulation. Mustard oil, topically applied to the receptive fields, also evoked localized Evans blue extravasation and strongly affected the C-units. They responded to the application of the irritant with sustained firing; most units showed an increased thermal sensitivity after the treatment. This involved a lowering of the threshold and an increase of the suprathreshold discharge. Even low threshold mechanosensitive C-fibres transiently developed heat sensitivity. In some polymodal units a transient sensitization was followed by a persistent desensitization which abolished their thermal and mechanical sensitivity. Mustard oil application and antidromic nerve stimulation seem to be similar in evoking cutaneous vasodilatation and plasma extravasation but different in inducing nociceptor sensitization.

The effects of hyperthermia on vascular permeability in experimental liver metastasis.

The effects of hyperthermia on vascular permeability in Walker carcinosarcomas and host liver tissue were studied in Sprague-Dawley rats. A quantitative Evans blue technique was used to measure permeability. With tumors heated to 40 degrees C, a nontherapeutic level, no changes in tumor vascular permeability as compared to control levels were noted. However, with tumors heated to 43 degrees C, within the therapeutic range of hyperthermia, significant rises in tumor vascular permeability occurred. Permeability was increased at both time periods studied, 30 minutes and 6 hours after hyperthermia and injection of Evans blue. These changes are similar to those seen after physical damage from freeze-thaw. It is likely that alterations in tumor microcirculation play a role in the therapeutic effect of intense hyperthermia.

1359 BILIRUBIN (BR) FLUX AND DISTRIBUTION IN THE BRAIN

Carotid artery injection of Evans Blue or albumin-bound bilirubin (ALB+BR) for 60 sec. in adult rats produced no cortical staining with an intact blood-brain barrier(BBB) and non-uniform “tufted” perivascular staining after barrier opening to ALB by 1.8 Molar Urea × 30 sec. Unbound BR given for 10-60 sec. to 25 rats without barrier opening, followed by carotid perfusion with saline, produced uniform staining, and the uptake of BR measured by chloroform extraction was dose-dependent (r=0.641, p<0.001). Retained BR was 11.6±6.2% (MEAN±S.D.) of the dose injected, and the permeability-surface area (P.S) product for BR was 0.13±0.05 ml·min−1 · g−1. The estimated volume of distribution for unbound BR at 60 sec. was 195±72 μl/g. When injection of unbound BR was followed by re-infusion with 5% ALB, residual brain BR decreased as shown:Bound BR crossing a disrupted BBB and unbound BR crossing an intact barrier appear to have different extravascular patterns of distribution, consistent with the greater lipid solubility and capillary permeability of unbound BR, versus the tendency of ALB + BR to cross the barrier only at osmotically opened capillaries. Because ALB decreases residual brain BR, we speculate that BR is initially distributed in a “shallow” compartment and is not immediately strongly bound or irreversibly precipitated in the brain.