Antioxidants attenuate microvascular changes in the early phase of experimental pneumococcal meningitis in rats.

BACKGROUND AND PURPOSE: We tested in a rat meningitis model 1) whether pneumococcal cell wall components are capable of producing changes in regional cerebral blood flow, brain water content, and intracranial pressure similar to those we have already observed after intracisternal inoculation of live pneumococci and 2) whether antioxidants would modulate these alterations in the early phase of meningitis. METHODS: Regional cerebral blood flow as measured by laser Doppler flowmetry and intracranial pressure were monitored continuously for 4 hours after intracisternal challenge. Brain edema formation was assessed by brain water content determinations. We investigated the following groups: rats challenged intracisternally with the whole intact pneumococcal cell wall (n = 7) or the pneumococcal cell wall hydrolyzed by the M1-muramidase (n = 7); rats injected intracisternally with phosphate-buffered saline (n = 6); rats pretreated intravenously with superoxide dismutase conjugated with polyethylene glycol (10,000 units/kg) and injected intracisternally with cell wall components (n = 5) or phosphate-buffered saline (n = 6); rats injected intracisternally with phosphate-buffered saline and pretreated intravenously with polyethylene glycol (10% solution, 1.2 ml/kg, n = 5) or continuously treated with intravenous free superoxide dismutase (22,000 units/kg per hour, n = 6); and rats continuously treated intravenously with deferoxamine mesylate (10 mg/kg per hour) and injected intracisternally with cell wall components (n = 6) or phosphate-buffered saline (n = 7). RESULTS: Both pneumococcal cell wall preparations produced a significant increase in regional cerebral blood flow, intracranial pressure, and brain water content. Conjugated superoxide dismutase as well as deferoxamine prevented the increase in intracranial pressure and brain water content. In addition, the increase in regional cerebral blood flow as observed in untreated, cell wall-challenged rats (baseline, 100%; 183.1 +/- 12.3% after 4 hours, mean +/- SEM) was significantly attenuated by administration of both conjugated superoxide dismutase (136.6 +/- 14.1%) and deferoxamine (149.8 +/- 8.2%) (p < 0.05). Polyethylene glycol-conjugated superoxide dismutase alone produced an increase in regional cerebral blood flow (125.6 +/- 8.7% after 4 hours). We found that polyethylene glycol per se accounts for this action. CONCLUSIONS: These data show that pneumococcal cell wall components containing teichoic acid produce changes in regional cerebral blood flow, intracranial pressure, and brain water content and that oxygen radicals contribute to these pathophysiological alterations in the early phase of experimental pneumococcal meningitis.     

Endothelin inhibition improves cerebral blood flow and is neuroprotective in pneumococcal meningitis

By using an infant rat model of pneumococcal meningitis, we determined whether endothelins contribute to neuronal damage in this disease. Cerebrospinal fluid analysis demonstrated a significant increase of endothelin-1 in infected animals compared with uninfected controls. Histopathological examination 24 hours after infection showed brain damage in animals treated with ceftriaxone alone (median, 9.2% of cortex; range, 0-49.1%). In infected animals treated intraperitoneally with the endothelin antagonist bosentan (30 mg/kg, every 12 hours) also, injury was reduced to 0.5% (range, 0-8.6%) of cortex. Cerebral blood flow was reduced in infected animals (6.5 +/- 4.0 ml/min/100 g of brain vs 14.9 +/- 9.1 ml/min/100 g in controls. Treatment with bosentan restored cerebral blood flow to levels similar to controls (12.8 +/- 5.3 ml/min/100 g). Improved blood flow was not mediated by nitric oxide production, because bosentan had no effect on cerebrospinal fluid or plasma nitrite/nitrate concentrations at 6, 12, or 18 hours. These data indicate that endothelins contribute to neuronal injury in this model of pneumococcal meningitis by causing cerebral ischemia.     

Anti ICAM-1 (CD 54) monoclonal antibody reduces inflammatory changes in experimental bacterial meningitis

We investigated whether monoclonal antibodies directed against intracellular adhesion molecule 1 (ICAM-1 mAb) inhibit brain edema, increase of intracranial pressure (ICP), regional cerebral blood flow (rCBF) and recruitment of white blood cells (WBC) into the cerebrospinal fluid (CSF) in the rat model of the early phase of bacterial meningitis. Brain edema was assessed by brain water content determinations. rCBF measured by laser Doppler flowmetry and ICP were recorded continuously for 6 h after intracisternal challenge. Meningitis was induced with pneumococcal cell walls (PCW). Increase of ICP and brain water content were significantly inhibited (P < 0.05) by intravenous treatment with ICAM-1 mAb (TM-8, 1 mg/kg). Furthermore, ICAM-1 mAb treatment profoundly attenuated (P < 0.05) rCBF increase and WBC invasion into the CSF. These results suggest that the ICAM-1 pathway is critically involved in the early phase of bacterial meningitis.     

Triptans reduce the inflammatory response in bacterial meningitis.

Severe headache and meningism provide clear evidence for the activation of trigeminal neurotransmission in meningitis. The authors assessed the antiinflammatory potential of 5HT1B/D/F receptor agonists (triptans), which inhibit the release of proinflammatory neuropeptides from perivascular nerve fibers. In a 6-hour rat model of pneumococcal meningitis, zolmitriptan and naratriptan reduced the influx of leukocytes into the cerebrospinal fluid, and attenuated the increase of regional cerebral blood flow. Elevated intracranial pressure as well as the brain water content at 6 hours was reduced by triptans. These effects were partially reversed by a specific 5HT1D as well as by a specific 5HT1B receptor antagonist. Meningitis caused a depletion of calcitonin gene-related peptide (CGRP) and substance P from meningeal nerve fibers, which was prevented by zolmitriptan and naratriptan. In line with these findings, patients with bacterial meningitis had significantly elevated CGRP levels in the cerebrospinal fluid. In a mouse model of pneumococcal meningitis, survival and clinical score at 24 hours were significantly improved by triptan treatment. The findings suggest that, besides mediating meningeal nociception, meningeal nerve fibers contribute to the inflammatory cascade in the early phase of bacterial meningitis. Adjunctive treatment with triptans may open a new therapeutic approach in the acute phase of bacterial meningitis.     

Induced hypothermia in experimental pneumococcal meningitis.

Pneumococcal meningitis resulting from Streptococcus pneumoniae has a death rate of 28% in adults. In severe head injury and stroke, inflammatory changes and intracranial hypertension are improved by induced hypothermia, which also is neuroprotective. We hypothesized that moderate hypothermia ameliorates inflammatory changes in experimental pneumococcal meningitis. Wistar rats were cooled systemically, and meningitis was induced by pneumococcal cell wall components. The increase of regional cerebral blood flow in the meningitis animals was blocked by hypothermia at 6 hours. The reduction of intracranial pressure correlated with temperature. The influx of leukocytes into the cerebrospinal fluid and levels of tumor necrosis factor alpha in the cerebrospinal fluid were decreased. Cooling the animals 2 hours after meningitis induction to 30.5 degrees C was also protective. We conclude that hypothermia is a new adjuvant approach to reduce meningitis-induced changes, in particular intracranial pressure, in the early phase of the disease.     

Differential regulation of blood-brain barrier permeability in brain trauma and pneumococcal meningitis-role of Src kinases

Increased vascular permeability causing vasogenic brain edema is characteristic for many acute neurological diseases such as stroke, brain trauma, and meningitis. Src family kinases, especially c-Src, play an important role in regulating blood-brain barrier permeability in response to VEGF, but also mediate leukocyte function and cytokine signalling. Here we demonstrate that pharmacological inhibition of Src or c-Src deficiency does not influence cerebrospinal fluid (CSF) pleocytosis, brain edema formation, and bacterial outgrowth during experimental pneumococcal meningitis despite the increased cerebral expression of inflammatory chemokines, such as IL-6, CCL-9, CXCL-1, CXCL-2 and G-CSF as determined by protein array analysis. In contrast, inhibition of Src significantly reduced brain edema formation, lesion volume, and clinical worsening in cold-induced brain injury without decreasing cytokine/chemokine expression. While brain trauma was associated with increased cerebral VEGF formation, VEGF levels significantly declined during pneumococcal meningitis. Therefore, we conclude that in brain trauma blood-brain barrier tightness is regulated by the VEGF/Src pathway whereas c-Src does not influence brain edema formation and leukocyte function during bacterial meningitis.     

Increased cerebral blood volume in benign intracranial hypertension.

In two patients with benign intracranial hypertension, the regional cerebral blood volume was markedly elevated (mean of 85 percent) while regional cerebral blood flow was slightly reduced (mean of 10 percent). Reduction of cerebrospinal fluid pressure by removal of cerebrospinal fluid reduced the mean values of regional cerebral blood volume by 13 percent without significant change in regional cerebral blood flow. The abnormal regional volume and regional flow returned to normal concurrent with the clinical improvement. Vinous engorgement and increased intracranial blood volume appear to play an important part in the pathophysiology of increased intracranial pressure in benign intracranial hypertension. A unified concept of the pathogenesis of benign intracranial hypertension is proposed.     

Hypothermia as an adjunctive treatment for severe bacterial meningitis

Brain injury due to bacterial meningitis results in a high mortality rate and significant neurologic sequelae in survivors. The objective of this study was to determine if the application of moderate hypothermia shortly after the administration of antibiotics would attenuate the inflammatory response and increase in intracranial pressure that occurs in meningitis. For this study we used a rabbit model of severe Group B streptococcal meningitis. The first component of this study evaluated the effects of hypothermia on blood-brain barrier function and markers of inflammation in meningitic animals. The second part of the study evaluated the effects of hypothermia on intracranial pressure, cerebral perfusion pressure and brain edema. This study demonstrates that the use of hypothermia preserves CSF/serum glucose ratio, decreases CSF protein and nitric oxide and attenuates myeloperoxidase activity in brain tissue. In the second part of this study we show a decrease in intracranial pressure, an improvement in cerebral perfusion pressure and a decrease in cerebral edema in hypothermic meningitic animals. We conclude that in the treatment of severe bacterial meningitis, the application of moderate hypothermia initiated shortly after antibiotic therapy improves short-term physiologic measures associated with brain injury.     

Production of nitrite by primary rat astrocytes in response to pneumococci

Recent studies using a rat model of pneumococcal meningitis have shown that nitric oxide synthase (NOS) inhibitors greatly attenuated microvascular changes and brain edema formation. The site of NO production during bacterial meningitis is unknown. In this study we tested whether primary astrocyte cultures from neonatal rat cortex can be induced to release NO upon stimulation with pneumococci. NO production was assessed by measuring nitrite in the cell culture supernatant using the Griess reaction. Stimulation with heat-killed unencapsulated pneumococci (HKP) increased nitrite concentrations in astrocyte culture supernatants in a dose-dependent fashion. Administration of AT-nitro-L-arginine (L-NA), aminoguanidine, L-canavanine, cycloheximide, and dexamethasone prevented the increase in nitrite concentrations. Addition of L-arginine, but not of o-arginine, partially reversed the inhibitory effect of L-NA. Administration of SOD increased nitrite accumulation. Moreover, at 72 h after stimulation with heat-killed pneumococci (10⁷ cfu/ml) astrocytes showed an inducible NOS-like immunoreactivity. Accumulation of nitrite was also observed when rat cerebellar neurons and microglia were stimulated with HKP, whereas there was only a slight increase of nitrite in media of rat C6 glioma cells, but no increase of nitrite when the human glioblastoma cell line LN-229 was stimulated with HKP. There was a stronger increase in nitrite levels when astrocytes from Lewis rats were used compared to that from Wistar rats. In conclusion, our study indicates that astrocytes, neurons and microglia are inducible for NO production upon stimulation with pneumococci.     

Transcranial Doppler sonography and intracranial pressure monitoring in children and juveniles with acute brain injuries or hydrocephalus

Transcranial Doppler sonography is a noninvasive method of obtaining information about changes in cerebral hemodynamics and intracranial pressure. After severe head injuries the development of brain swelling and brain edema can be assessed and the efficacy of treatment monitored. Development of severe brain edema accompanied by a rapid increase in intracranial pressure can be recognized by a decrease in blood flow velocity and rise in the pulsatility index. In hydrocephalic children the behavior of the cerebral blood flow velocity and the pulsatility index will warn of an increase of the ventricular fluid pressure or a shunt insufficiency.Presented at the 12th European Congress of Paediatric Neurosurgery, Warsaw 1990     

Microvascular changes during the early phase of experimental bacterial meningitis

We investigated the temporal profile of the changes in regional CBF (rCBF) and intracranial pressure (ICP) during the early phase of pneumococcal meningitis in the rat. rCBF, as measured by laser-Doppler flowmetry, and ICP were continuously monitored during 6 h post infection (p.i.). Brain edema formation was assessed by brain water content determinations. Meningitis was induced by intracisternal injection of 75 microliters of 10(7) colony-forming units/ml pneumococci (n = 7). In control animals (n = 6), saline was injected. There was no change in the rCBF or ICP of controls throughout the experiment. However, there was a dramatic increase in rCBF and ICP associated with brain edema formation in untreated meningitis animals. rCBF increased to 135.3 +/- 33.8% (mean +/- SD) in the untreated animals at 1 h p.i. and reached 211.1 +/- 40.5% at 6 h p.i. (p less than 0.05 compared with controls). ICP increased from 2.9 +/- 1.4 to 10.4 +/- 4.7 mm Hg at 6 h p.i. (p less than 0.05 compared with controls). Brain water content was significantly elevated (79.69 +/- 0.24 compared with 78.94 +/- 0.16% in the control group, p less than 0.05). We investigated the effect of dexamethasone (3 mg/kg i.p.), which was given prior to the induction of meningitis (n = 3) or at 2 h after pneumococcal injection (n = 5), indomethacin (10 mg/kg i.v., n = 5), and superoxide dismutase (SOD; 132,000 U/kg i.v. per 6 h, n = 6).(ABSTRACT TRUNCATED AT 250 WORDS)     

Intracranial venous pressures,hydrocephalus and effects of cerebrospinal fluid shunts

Data concerning venous anatomy, interstitial fluid pressure and cerebral blood flow indicate that obstruction of cerebral venous outflow (as a whole or involving the deep venous system alone) is the essential cause of hydrocephalus. Choroidal and ventricular veins both belong to the deep system. Choroidal venous pressure determines cerebrospinal fluid pressure; pressure in the ventricular veins determines interstitial fluid pressure in the paraventricular white matter. A decrease in deep cerebral blood flow causes paraventricular atrophy. CSFP is higher than interstitial fluid pressure, normally and in venous obstruction. Thus, CSFP prevents venocongestive edema (but not inflammatory edema) of the brain. Collateral venous pathways are described. Venous obstruction causes hydrocephalus only when it leads to insufficient blood flow. Cerebrospinal fluid shunting causes increased CBF as essential therapeutic effect in hydrocephalus, but also causes venocongestive brain edema, which explains the decrease in ventricle size and the side effects of shunting.Presented at the XVI Annual Meeting of the International Society for Pediatric Neurosurgery, Rome 1988     

Role of Caspase-1 in experimental pneumococcal meningitis: Evidence from pharmacologic Caspase inhibition and Caspase-1-deficient mice

Caspase 1 plays a pivotal role in generating mature cytokine interleukin-1beta. Interleukin-1beta is implicated as a mediator of pneumococcal meningitis, both in experimental models and in humans. We demonstrated here that (1) Caspase 1 mRNA and protein expression is upregulated in the brain during experimental pneumococcal meningitis, and (2) Caspase 1 levels are elevated in the cerebrospinal fluid of patients with acute bacterial meningitis. The upregulation/activation of Caspase 1 was associated with increased levels of interleukin-1beta. Depletion of the Caspase 1 gene and pharmacologic blockade of Caspase 1 significantly attenuated the meningitis-induced increase in interleukin-1beta. This was paralleled by a significantly diminished inflammatory host response to pneumococci. The antiinflammatory effect of Caspase 1 depletion or blockade was associated with a marked reduction of meningitis-induced intracranial complications, thus leading to an improved clinical status. In humans, cerebrospinal fluid Caspase 1 levels correlated with the clinical outcome. Thus, pharmacologic inhibition may provide an efficient adjuvant therapeutic strategy in this disease.     

Experimental meningitis in the rabbit. II. Cerebral energy metabolism in relation to increased cerebrospinal fluid concentrations of lactate

We have analyzed cerebral energy metabolism in rabbits with Streptococcus pneumoniae or Escherichia coli meningitis aiming at an increased understanding of the cerebrospinal fluid (CSF) lactacidosis observed in this disease. After intracisternal inoculation of bacteria the lactate concentration in the CSF increased to 9.7 +/- 0.7 (mean +/- SE) mmol/l compared to control values of 3.2 +/- 0.2 mmol/l. Simultaneously sampled brain tissue from parietal cortex, caudate nucleus, and thalamus showed no increase in lactate concentrations. The high-energy phosphate content decreased only marginally, phosphocreatine levels by 11-17% in the cortex and in the caudate nucleus, and adenosine triphosphate concentrations by 15%, but only in the caudate nucleus. Our results indicate that the CSF lactate increase in bacterial meningitis is not primarily linked to cerebral lactacidosis. The decreased concentrations of high-energy phosphates in diseased animals need further study but may be due to increased intracranial pressure and reduced capillary blood flow.     

Lack of endothelial nitric oxide synthase aggravates murine pneumococcal meningitis.

Nitric oxide (NO) plays a central role in the pathogenesis of bacterial meningitis. However, the role of NO produced by endothelial NO synthase (eNOS) in meningitis is still unclear. We investigated the influence of eNOS depletion on the inflammatory host response, intracranial complications, and outcome in experimental pneumococcal meningitis. Leukocyte accumulation in the cerebrospinal fluid was more pronounced in infected eNOS-deficient mice than in infected wild type mice. This effect could be attributed to an increased expression of P-selectin, macrophage inflammatory protein-2, keratinocyte-derived cytokine, and interleukin (IL)-1beta in the brain of infected eNOS-deficient mice. However, no differences in the cerebral expression of intercellular adhesion molecule-1, tumor necrosis factor-alpha, and IL-6 as well as of neuronal NOS and inducible NOS could be detected between infected wild type and mutant mice. In addition to enhanced leukocyte infiltration into the CSF, meningitis-associated intracranial complications including blood-brain barrier disruption and the rise in intracranial pressure were significantly augmented in infected eNOS-deficient mice. The aggravation of intracranial complications was paralleled by a worsening of the disease, as evidenced by a more pronounced hypothermia, an enhanced weight reduction, and an increased death rate. The current data indicate that eNOS deficiency is detrimental in bacterial meningitis. This effect seems to be related to an increased expression of (certain) cytokines/chemokines and adhesion molecules; thus leading to increased meningeal inflammation and, subsequently, to aggravated intracranial complications.     

Is nitric oxide involved as a mediator of cerebrovascular changes in the early phase of experimental pneumococcal meningitis?

Abstract

Nitric oxide (NO) is a mediator of haemodynamic changes and cytotoxicity in in vivo models of inflammation such as endotoxaemic shock. The purpose of this study was to investigate whether NO may be involved in the increase of cerebral blood flow (CBF), intracranial pressure (ICP) and brain water content, known to occur in the early phase of pneumococcal meningitis. Rats injected intracisternally with live pneumococci were either untreated or received 5 mg kg^–1 i.v. N^G-nitro-L-arginine methyl ester (L-NAME), an inhibitor of NO synthase. Pretreatment with L-NAME prevented the increase in CBF, ICP and brain water content, as seen in untreated animals. CBF tended to return towards baseline when rats were treated with L-NAME 2 h after pneumococcal injection. Whereas none of the untreated and L-NAME-pretreated animals died during the 6 h observation period 3 out of 9 rats treated with L-NAME and 7 out of 9 rats with simultaneous i.v. injection of L-NAME and L-arginine died. Our results provide preliminary evidence that NO may be involved as a mediator of CBF changes and oedema formation in the early phase of pneumococcal meningitis in the rat. NO inhibition, however, may have detrimental effects of still unidentified cause, as indicated by the increased mortality in treated animals. Further studies with analysis of the causes of mortality, structurally different NO synthase inhibitors and direct evaluation of NO synthase induction are needed to further support this hypothesis. [Neurol Res 1994; 16: 108-112]     

N(omega) -nitro-L-arginine methyl ester (L-NAME) attenuates the acute inflammatory responses and brain injury during the early phase of experimental Escherichia coli meningitis in the newborn piglet.

We evaluated the anti-inflammatory and neuroprotective effect of nonselective NOS inhibitor, N(omega)-nitro-L-arginine methyl ester (L-NAME), in experimental bacterial meningitis in the newborn piglet. Meningitis was induced by intracisternal injection of 10(8) colony forming units of Escherichia coli. L-NAME 10 mg kg(-1) was given intravenously 30 min before induction of meningitis. L-NAME significantly attenuated the increase in intracranial pressure and decrease in cerebrospinal fluid glucose concentration observed in the meningitis group. Systemic and cerebral perfusion pressure were even higher compared to the control and meningitis groups. However, the meningitis-induced increase in tumor necrosis factor-alpha level, leukocyte numbers and lactate level in the cerebrospinal fluid was not significantly attenuated with L-NAME administration. Reduced cerebral cortical cell membrane Na+, K+ -ATPase activity and increased lipid peroxidation products, indicative of meningitis-induced brain cell membrane dysfunction, were significantly improved with L-NAME treatment. Decreased brain glucose and ATP levels were also significantly improved with L-NAME treatment. These findings suggest that L-NAME was effective in attenuating the acute inflammatory responses and brain injury in neonatal bacterial meningitis.     

Endothelium-derived nitric oxide synthase inhibition. Effects on cerebral blood flow, pial artery diameter, and vascular morphology in rats.

BACKGROUND AND PURPOSE: We determined the effects of inhibiting the production of cerebral endothelium-derived nitric oxide on pial artery diameter, cortical blood flow, and vascular morphology. METHODS: An inhibitor of endothelium-derived nitric oxide synthesis, NG-nitro-L-arginine methyl ester hydrochloride (L-NAME), or an equivalent volume of 0.9% saline was infused into rats intra-arterially in a retrograde fashion via the right external carotid artery at a rate of 3 mg/kg/min to a total dose of 190 mg/kg or intravenously at 1 mg/kg/min to a total dose of 15 mg/kg. Large pial arteries were continuously visualized through an operating microscope, and cortical cerebral blood flow was monitored by laser-Doppler flowmetry. To localize areas of morphological interest, the protein tracer horseradish peroxidase was injected 15 minutes before termination of the L-NAME infusion and the rats were perfusion-fixed 15 minutes later for light and electron microscopic analysis. RESULTS: Infusion of L-NAME significantly raised arterial blood pressure at both doses (for 190 mg/kg, from 103.2 +/- 3.4 to 135 +/- 3.4 mm Hg; for 15 mg/kg, from 125 +/- 2.8 to 144.4 +/- 4.0 mm Hg). Pial arteries constricted within 10 minutes after the start of the intracarotid infusion to 40% of the preinfusion diameter, while cortical cerebral blood flow decreased to an average of 72.5% of that at baseline. Morphological abnormalities in the experimental rats included microvascular stasis and focal areas of blood-brain barrier disruption to protein. Ultrastructural examination of cortical leaky sites revealed constricted arterioles with many endothelial pinocytotic vesicles and microvilli. CONCLUSIONS: These observations suggest that inhibition of endothelium-derived nitric oxide synthesis affects the relation between cerebral arterial diameter and cerebral blood flow and can lead to subtle cerebral vascular pathological changes consistent with focal brain ischemia.     

Acute intracranial hypertension

Acute intracranial hypertension is a syndrome with multiple etiologies. Diagnosis and treatment must be performed urgently to save the patient's life and prevent the development of significant disabilities. The appearance of this syndrome is due to intracraincreased volumes and -in turn- the pressure of the intracranial contents, either through an increase in the physiological components (blood, cerebrospinal fluid and brain parenchyma), or through the appearance of a volume in the form of added mass. The underlying brain edema in this condition may be of several types: cytotoxic, vasogenic, interstitial, or hydrostatic. Increased intracranial pressure decreases cerebral perfusion pressure, creating a vicious cycle because of the resulting cerebral ischemia, which progressively increases cerebral blood volume by decreasing resistance and further increases intracranial pressure. Treatment depends on the etiology and will generally require medical and surgical care. Patient management is usually carried out in neurocritical units and involves intracranial pressure monitoring to guide treatment. Correction of all hemostasis disorders is also crucial to patient survival.     

Nitric oxide donor-induced increase in permeability of the blood-brain barrier

The goal of the present study was to determine the effect of nitric oxide (NO) donors on the permeability of the blood-brain barrier in vivo. We examined the pial microcirculation in rats using intravital fluorescence microscopy. Permeability of the blood-brain barrier was quantitated by calculating the clearance of fluorescent-labeled dextran (M(w)=10000 Da; FITC-dextran-10K) during suffusion with vehicle, S-nitroso-N-acetylpenicillamine (SNAP; 100 microM) and 3-morpholinosydnonimin (SIN-1; 100 microM). In addition, we examined changes in arteriolar diameter during suffusion with vehicle, SNAP and SIN-1. During suffusion with vehicle, clearance of FITC-dextran-10K from pial vessels and diameter of pial arterioles remained relatively constant during the experimental period. In contrast, suffusion with SNAP or SIN-1 markedly increased clearance of FITC-dextran-10K from the cerebral microcirculation and produced a rapid, sustained dilatation of pial arterioles. Thus, NO donors increase the permeability of the blood-brain barrier and produce pronounced dilatation of cerebral arterioles. In light of evidence suggesting that NO donors may produce their effect by the simultaneous release of NO and superoxide anion to form peroxynitrite, we elected to examine the role of superoxide anion in increases in permeability of the blood-brain barrier in response to SNAP and SIN-1. We found that suffusion with tiron (1 mM) did not alter basal permeability of the blood-brain barrier, but significantly inhibited increases in permeability of the blood-brain barrier in response to SNAP and SIN-1. In addition, tiron did not alter baseline diameter of cerebral arterioles, or SNAP- and SIN-1-induced cerebrovasodilatation. The findings of the present study suggest that NO donors produce an increase in permeability of the blood-brain barrier which appears to be related to the presence of NO and superoxide anion, to presumably form peroxynitrite. We suggest that increases in NO formation observed during brain trauma may contribute to disruption of the blood-brain barrier.