Simultaneous imaging of cortical blood flow and haemoglobin concentration with LASCA and RGB reflectometry.

We demonstrate a system for the simultaneous imaging of cortical blood flow and haemoglobin oxygenation by laser speckle contrast analysis (LASCA) and RGB reflectometry. The sensitivity of the system was tested by observing changes of haemoglobin oxygenation and blood flow in rats in response to ischaemic stroke, hypercapnia, hyperoxia, hypoxia, cortical spreading depression and cortical activation following forepaw stimulation.

Towards noninvasive molecular fluorescence imaging of the human brain.

Fluorescence molecular brain imaging is a new modality allowing the detection of specific contrast agents down to very low concentration ranges (picomolar) in disease models. Here we demonstrate a first noninvasive application of fluorescence imaging in the human brain, where concentrations down to about 100 nM of a nonspecific dye were detected. We argue that due to its high sensitivity, optical molecular imaging of the brain is feasible, which - together with its bedside applicability - makes it a promising technique for use in patients.

Blockade of nitric oxide synthesis in rats strongly attenuates the CBF response to extracellular acidosis.

We tested the hypothesis that the CBF response to extracellular acidosis is mediated by nitric oxide (NO). A closed cranial window, superfused with artificial CSF (aCSF), was implanted over the parietal cortex in anesthetized and ventilated Wistar rats. Regional cerebral blood flow (rCBF) was measured continuously with laser-Doppler flowmetry (LDF). The reaction of rCBF to hypercapnia (PaCO2 from 30.5 +/- 1.8 to 61.3 +/- 5.8 mm Hg by adding CO2 to the inspiratory gas) was 2.9 +/- 1.4%/mm Hg, and the reaction of rCBF to H+ (superfusion of acidic aCSF, pH 7.07 +/- 0.05) was 101.7 +/- 24.7%/pH unit. The regional NO synthase (NOS) activity was blocked by superfusing aCSF containing 10(-3) M N omega-nitro-L-arginine (L-NA, n = 10). After 30 min of L-NA superfusion, rCBF was reduced to 80.1 +/- 6.5% of baseline, and the rCBF responses to hypercapnia (PaCO2 from 30.9 +/- 2.9 to 58.8 +/- 7.7 mm Hg) and extracellular acidosis (aCSF pH 7.08 +/- 0.06) were reduced to 0.8 +/- 1.1%/mm Hg and 10.1 +/- 23.0%/pH unit, respectively (both p < 0.001). This effect was stereospecific since aCSF containing 10(-3) M N omega-nitro-D-arginine affected neither baseline rCBF nor the response to H+ (n = 5). The NOS blockade did not affect the vasodilatation by the NO donor sodium nitroprusside (n = 5, 114.3 +/- 25.1% before vs. 130.2 +/- 24.7% after NOS blockade). The results confirm the involvement of NO in the CBF reaction to hypercapnia and demonstrate for the first time that NOS blockade also strongly attenuates the H+ response of the cerebral vasculature.(ABSTRACT TRUNCATED AT 250 WORDS)

Increased formation of reactive oxygen species after permanent and reversible middle cerebral artery occlusion in the rat.

In barbiturate-anesthetized rats, we induced 3 hours of permanent middle cerebral artery occlusion (MCAO) by an intraluminal thread (n = 6), or 1 hour MCAO followed by 2 hours of reperfusion (n = 6). Through a closed cranial window over the parietal cortex, the production of reactive oxygen species (ROS) was measured in the infarct border using online in vivo chemiluminescence (CL) while monitoring the appearance of peri-infarct depolarizations (PID). The borderzone localization of the ROS and direct current (DC) potential measurements was confirmed in additional experiments using laser-Doppler scanning, mapping regional CBF changes through the cranial window after permanent (n = 5) or reversible (n = 5) MCAO. CL measurements revealed a short period (10 to 30 minutes) of reduced ROS formation after vessel occlusion, followed by a significant increase (to 162 +/- 51%; baseline = 100%; P < .05) from 100 minutes of permanent MCAO onward. Reperfusion after a 1-hour period of MCAO led to a burst-like pattern of ROS production (peak: 489 +/- 330%; P < .05). When the experiments were terminated 3 hours after induction of MCAO, CL was still significantly increased above baseline after permanent and reversible MCAO (to 190 +/- 67% and 211 +/- 64%, respectively; P < .05). Simultaneous DC potential recordings detected 6.4 +/- 2.7 PID in the first, 4.7 +/- 2.3 in the second, and 2.8 +/- 2.0 in the third hour after permanent MCAO. In animals with reversible MCAO, PID were abolished from 15-minutes recirculation onward. There was no temporal relationship between ROS production and peri-infarct DC potential shifts. In conclusion, using a high temporal resolution ROS detection technique (CL), we found that permanent MCAO (after an initial decrease) was accompanied by a steady increase of ROS production during the 3-hour observation period, while reperfusion after 1 hour of MCAO produced a burst in ROS formation. Both patterns of ROS production were not related to the occurrence of PID.

Noninvasive near infrared spectroscopy monitoring of regional cerebral blood oxygenation changes during peri-infarct depolarizations in focal cerebral ischemia in the rat.

Intermittent peri-infarct depolarizations (PID), which spread from the vicinity of the infarction over the cortex, have been reported in focal ischemia. These depolarizations resemble cortical spreading depression except that they damage the cortex and enlarge the infarct volume possibly because of compromised oxygen delivery. The main purpose of this study was to evaluate the noninvasive technique of near-infrared spectroscopy (NIRS) for the identification of PID and to evaluate its capability for further pathophysiological studies. We used male barbiturate-anesthetized Wistar rats (n = 10) in which middle cerebral artery occlusion had been performed with a surgical thread. Middle cerebral artery occlusion resulted in a drop in parietally measured regional cerebral blood flow (laser Doppler flowmetry) to 31 +/- 8% of baseline flow. Six +/- 4 minutes after the induction of focal ischemia, 5 +/- 2 direct current deflections were recorded during a one-hour measurement period which may be regarded as PID. Measuring regional cerebral blood oxygenation changes with a NIRO 500 revealed dynamic concentration changes in the three chromophores oxyhemoglobin [HbO2], deoxyhemoglobin [Hb], and the oxidized form of cytochrome aa3 [CytO] during PID. Typically, an initial slight decrease of [HbO2] (-6.1 +/- 1.7 arbitrary units [AU] and an increase of [Hb] (+11.5 +/- 7.7 AU) were followed by an increase of [HbO2] (+10.8 +/- 4.7 AU) and a decrease of [Hb] (-4.7 +/- 5.5 AU); [CytO] decreased during the depolarizations (-2.0 +/- 1.2 AU). We conclude that NIRS can detect typical PID-associated changes in blood oxygenation. We hypothesize that during the course of PID, unlike "normal" spreading depression, hypoxygenation precedes hyperoxygenation of the microcirculation in a given cortex volume as the depolarization wave propagates through hemodynamically compromised to intact tissue. This would accord with the known damaging effect of PID. The NIRS "fingerprint" of PID encourages the search for PID during early stroke in patients.

Nitric oxide modulates the CBF response to increased extracellular potassium.

The response of the regional cerebral blood flow (rCBF) to brain topical superfusion of 20 mM K+ was characterized in a closed cranial window preparation in barbiturate anesthetized and ventilated rats: Increasing K+ in the artificial cerebrospinal fluid (ACSF) induced a rCBF elevation (measured by laser-Doppler flowmetry) of +85 +/- 37% above baseline (n = 19). This elevation was stable for > 3 h with continuous superfusion of increased K+ (n = 5) and partially reversible to a level of +18 +/- 19% above baseline when returning to a physiological K+ concentration. Nitric oxide synthase (NOS) inhibition by brain topical superfusion with N omega-nitro-L-arginine (L-NA) revealed (a) Addition of L-NA to high-potassium ACSF reduced the rCBF increase from +94 +/- 36% to +21 +/- 18% (p < or = 0.01, n = 7). (b) When L-NA was superfused for 60 min before increasing K+, rCBF decreased to -17 +/- 7% below baseline. Subsequent coapplication of L-NA and increased K+ induced only an elevation of +7 +/- 4% above baseline (n = 4). (c) When the NO donor S-nitroso-N-acetylpenicillamine (SNAP) was added during NOS inhibition to restore basal tissue NO levels, the resultant level of rCBF was +28 +/- 54% above baseline. Subsequent increase of K+ in the presence of NOS inhibition and SNAP elevated rCBF to +137 +/- 89% above baseline (n = 4).(ABSTRACT TRUNCATED AT 250 WORDS)

Systemic nitric oxide synthase inhibition does not affect brain oxygenation during cortical spreading depression in rats: a noninvasive near-infrared spectroscopy and laser-Doppler flowmetry study.

Cortical spreading depression (CSD) has been implicated in the migraine aura and in stroke. This study demonstrates near-infrared spectroscopy (NIRS) for the first time as capable of noninvasive on-line detection of CSD in the pentobarbital-anesthetized rat. CSD was accompanied by a brief and rapid increase of regional CBF (by laser-Doppler flowmetry) to 200-400% baseline. NIRS demonstrates that this hyperperfusion is associated with concentration increases of oxyhemoglobin, while deoxyhemoglobin decreases. Simultaneously, oxygen partial pressure, measured on the brain surface with a solid-state polarographic probe, was shown to be raised by at least 14 mm Hg during CSD. Oxygen-dependent phosphorescence life-time quenching measurements confirmed this finding. NIRS data on cytochrome aa3, however, showed a CSD-related shift toward a more reduced state, despite raised blood oxygenation. This may suggest either limited O2 transport from the blood to mitochondria or decreased oxygen utilization during CSD as supposed by theories about compartmentalization of energy metabolism favoring glycolytic rather than aerobic energy supply during CSD. However, the data on cytochrome aa3 warrant caution and are discussed critically. Nitric oxide synthase inhibition by systemic application of N'-nitro-L-arginine had no significant effect on the perfusion response or the tissue PO2 during CSD. During most CSD episodes, a brief decrease in MABP by 4-8 mm Hg was noted that might be caused by functional decortication during CSD.

Role of nitric oxide synthase inhibition in leukocyte-endothelium interaction in the rat pial microvasculature.

We investigated the role of nitric oxide (NO) in leukocyte-endothelium interaction, blood-brain barrier (BBB) function and oxygen free-radical production in the rat pial microcirculation. In a closed cranial window preparation (dura removed) over the parietal cortex of pentobarbital-anesthetized Wistar rats, NO synthase (NOS) was inhibited by systemic and/or topical application of N omega-nitro-L-arginine (L-NNA) under physiological conditions and during leukotriene B4 (LTB4) activation. Circulating leukocytes were labeled by intravenous injection of rhodamine 6G. We used a confocal laser scanning microscope (CLSM) and studied leukocyte rolling and sticking in pial veins and arteries before and after NOS inhibition. At the end of the experiments, sodium-fluorescein was injected intravenously to test BBB integrity. Brain cortex oxygen free-radical production was investigated in the cranial window preparation using lucigenin-enhanced chemiluminescence (CL). L-NNA application did not lead to significant changes in leukocyte-endothelium interaction, BBB function, and oxygen free-radical production under physiological conditions [leukocyte-endothelium interaction: control (n = 5), L-NNA systemically (n = 5), L-NNA topically (n = 5): at baseline rollers/100 microns: 0.76 +/- 0.55, 0.64 +/- 0.94, 0.44 +/- 0.55 and stickers/100 microns: 0.90 +/- 0.28, 0.76 +/- 0.24, 0.84 +/- 0.42; at 60 min rollers/100 microns: 1.49 +/- 0.66, 1.21 +/- 0.99, 0.67 +/- 0.66 and stickers/100 microns: 1.04 +/- 0.20, 1.19 +/- 0.23, 1.21 +/- 0.54; oxygen free-radical production (n = 4): CL count before L-NNA application 35 +/- 17 cps, after 1 h of topical superfusion of L-NNA 38 +/- 14 cps; p < 0.05]. In contrast to the results achieved under physiological conditions, a significant further increase of rolling leukocytes and BBB permeability occurred due to NOS inhibition under LTB4-activated conditions [76 +/- 47% significant (p < or = 0.01, n = 7) further increase of rollers/100 microns due to 60 min L-NNA application following the activation period of 120 min LTB4 superfusion]. Our results support a modulatory role for NO in leukocyte-endothelium interaction and BBB permeability in the pial microcirculation when this interaction is increased.

Global cerebral ischemia in the rat: online monitoring of oxygen free radical production using chemiluminescence in vivo.

Using online in vivo chemiluminescence (CL), we studied for the first time continuously the production of reactive oxygen species (ROS) after global cerebral ischemia and the relationship of ROS production to CBF. In anesthetized rats equipped with a closed cranial window, the CL enhancer, lucigenin (1 mM), was superfused onto the brain topically. CL was measured through the cranial window with a cooled photomultiplier, and CBF was measured simultaneously with laser-Doppler flowmetry. Reperfusion after 10 min (n = 8) of global cerebral ischemia led to a CL peak to 188 +/- 77% (baseline = 100%) within 10 +/- 4 min. After 2 h of reperfusion, CL had returned to 102 +/- 28%. Reperfusion after 20 min (n = 8) of ischemia increased CL to 225 +/- 48% within 12 +/- 3 min. After 2 h, CL was still increased (150 +/- 44%, p < 0.05 compared with 10 min of ischemia). CL after 10 min of ischemia was neither affected by brain topical free CuZn-superoxide dismutase (SOD) (100 U/ml, n = 3) nor by i.v. administration of free CuZn-SOD (104 U/kg, followed by 104 U/kg/h, n = 3). The CBF hyperfusion peak on reperfusion preceded the CL peak in all experiments by several minutes. In additional in vitro experiments we investigated the source of CL: Intracellular loading of lucigenin was demonstrated in cultured CNS cells, and a very similar pattern of CL as in the in vivo preparation after ischemia developed in rat brain slices after 15 min of hypoxia, which was unaffected by free CuZn-SOD (100 U/ml) but strongly attenuated by liposome-entrapped CuZn-SOD. We conclude that lucigenin-enhanced CL is a promising tool to study ROS production continuously from the in vivo brain of experimental animals and brain slices, and that the CL signal most likely derives from the intracellular production of superoxide. The production of ROS is preceded by reperfusion, is burst-like, and is dependent on the duration of the ischemic interval.

Nitric oxide scavenging by hemoglobin or nitric oxide synthase inhibition by N-nitro-L-arginine induces cortical spreading ischemia when K+ is increased in the subarachnoid space.

We investigated the combined effect of increased brain topical K+ concentration and reduction of the nitric oxide (NO.) level caused by nitric oxide scavenging or nitric oxide synthase (NOS) inhibition on regional cerebral blood flow and subarachnoid direct current (DC) potential. Using thiopental-anesthetized male Wistar rats with a closed cranial window preparation, brain topical superfusion of a combination of the NO. scavenger hemoglobin (Hb; 2 mmol/L) and increased K+ concentration in the artificial cerebrospinal fluid ([K+]ACSF) at 35 mmol/L led to sudden spontaneous transient ischemic events with a decrease of CBF to 14+/-7% (n=4) compared with the baseline (100%). The ischemic events lasted for 53+/-17 minutes and were associated with a negative subarachnoid DC shift of -7.3+/-0.6 mV of 49+/-12 minutes' duration. The combination of the NOS inhibitor N-nitro-L-arginine (L-NA, 1 mmol/L) with [K+]ACSF at 35 mmol/L caused similar spontaneous transient ischemic events in 13 rats. When cortical spreading depression was induced by KCl at a 5-mm distance, a typical cortical spreading hyperemia (CSH) and negative DC shift were measured at the closed cranial window during brain topical superfusion with either physiologic artificial CSF (n=5), or artificial CSF containing increased [K+]ACSF at 20 mmol/L (n=4), [K+]ACSF at 3 mmol/L combined with L-NA (n=10), [K+]ACSF at 10 mmol/L combined with L-NA (five of six animals) or [K+]ACSF at 3 mmol/L combined with Hb (three of four animals). Cortical spreading depression induced longlasting transient ischemia instead of CSH, when brain was superfused with either [K+]ACSF at 20 mmol/L combined with Hb (CBF decrease to 20+/-20% duration 25+/-21 minutes, n=4), or [K+]ACSF at 20 mmol/L combined with L-NA (n=19). Transient ischemia induced by NOS inhibition and [K],ACSF at 20 mmol/L propagated at a speed of 3.4+/-0.6 mm/min, indicating cortical spreading ischemia (CSI). Although CSH did not change oxygen free radical production, as measured on-line by in vivo lucigenin-enhanced chemiluminescence, CSI resulted in the typical radical production pattern of ischemia and reperfusion suggestive of brain damage (n=4). Nimodipine (2 microg/kg body weight/min intravenously) transformed CSI back to CSH (n=4). Vehicle had no effect on CSI (n=4). Our data suggest that the combination of decreased NO. levels and increased subarachnoid K+ levels induces spreading depression with acute ischemic CBF response. Thus, a disturbed coupling of metabolism and CBF can cause ischemia. We speculate that CSI may be related to delayed ischemic deficits after subarachnoid hemorrhage, a clinical condition in which the release of Hb and K+ from erythrocytes creates a microenvironment similar to the one investigated here.

Heparin inhibits leukocyte rolling in pial vessels and attenuates inflammatory changes in a rat model of experimental bacterial meningitis.

Heparin is a natural proteoglycan that was first described in 1916. In addition to its well characterized effect on blood coagulation, it is becoming clear that heparin also modulates inflammatory processes on several levels, including the interference with leukocyte-endothelium interaction. Anecdotal observations suggest a better clinical outcome of heparin-treated patients with bacterial meningitis. The authors demonstrate that heparin, a glycosaminoglycan, inhibits significantly in the early phase of experimental pneumococcal meningitis the increase of 1) regional cerebral blood flow (125 +/- 18 versus 247 +/- 42%), 2) intracranial pressure (4.5 +/- 2.0 versus 12.1 +/- 2.2 mm Hg), 3) brain edema (brain water content: 78.23 +/- 0.33 versus 79.49 +/- 0.46%), and 4) influx of leukocytes (571 +/- 397 versus 2400 +/- 875 cells/microL) to the cerebrospinal fluid compared with untreated rats. To elucidate the possible mechanism of this observation, the authors investigated for the first time leukocyte rolling in an inflammatory model in brain venules by confocal laser scanning microscopy in vivo. Heparin significantly attenuates leukocyte rolling at 2, 3, and 4 hours (2.8 +/- 1.3 versus 7.9 +/- 3.2/100 microm/min), as well as leukocyte sticking at 4 hours (2.1 +/- 0.4 versus 3.5 +/- 1.0/100 microm/min) after meningitis induction compared with untreated animals. The authors conclude that heparin can modulate acute central nervous system inflammation and, in particular, leukocyte-endothelium interaction, a key process in the cascade of injury in bacterial meningitis. They propose to evaluate further the potential of heparin in central nervous system inflammation in basic and clinical studies.

Respiratory chain inhibition induces tolerance to focal cerebral ischemia.

The authors show that the inhibitor of the succinate dehydrogenase, 3-nitroproprionic acid (3-NPA), which in high doses and with chronic administration is a neurotoxin, can induce profound tolerance to focal cerebral ischemia in the rat when administered in a single dose (20 mg/kg) 3 days before ischemia. Infarcts were approximately 70% and 35% smaller in the 3-NPA preconditioned groups of permanent and transient focal cerebral ischemia, respectively. This regimen of 3-NPA preconditioning neither induced necrosis, apoptosis, or any other histologically detectable damage to the brain, nor did it affect behavior of the animals. 3-NPA led to an immediate (1-hour) and long-lasting (3-day) decrease in succinate dehydrogenase activity (30% reduction) throughout the brain, whereas only a short metabolic impairment occurred (ATP decrease of 35% within 30 minutes, recovery within 2 hours). The authors found that 3-NPA induces a burst of reactive oxygen species and the free radical scavenger dimethylthiourea, when administered shortly before the 3-NPA stimulus, completely blocked preconditioning. Inhibition of protein synthesis with cycloheximide given at the time of 3-NPA administration completely inhibited preconditioning. The authors were unsuccessful in showing upregulation of mRNA for the manganese superoxide dismutase, and did not detect increased activities of the copper-zinc and manganese superoxide dismutases, prototypical oxygen free radicals scavenging enzymes, after 3-NPA preconditioning. The authors conclude that it is possible to pharmacologically precondition the brain against focal cerebral ischemia, a strategy that may in principal have clinical relevance. The data show the relevance of protein synthesis for tolerance, and suggests that oxygen free radicals may be critical signals in preconditioning.

Excessive oxygen or glucose supply does not alter the blood flow response to somatosensory stimulation or spreading depression in rats.

We investigated the influence of hyperoxia (arterial pO2 446 +/- 43 mmHg) and hyperglycemia (blood glucose 19.4 mmol/l) on somatosensory stimulation (whisker deflection) employing laser Doppler flowmetry (LDF). Our aim was to test the hypothesis that a possible substrate-sensing mechanism for glucose and oxygen contributes to the coupling between cortical activity and regional cerebral blood flow (rCBF) in order to match increased demand with substrates. In addition, we looked at the influence of hyperglycemia (blood glucose 17.9 mmol/l) and hypercapnia (arterial pCO2 62 mmHg) on rCBF (LDF) and regional cerebral blood oxygenation changes (rCBO) in the even stronger metabolic stimulus of cortical spreading depression (CSD). For the latter we employed the new non-invasive technique of near infrared spectroscopy (NIRS). All experiments were done using chloralose/urethane-anesthetized rats. Somatosensory stimulation increased rCBF by about 20% of baseline, in the case of both norm- and hyperoxia as well as both normo- and hyperglycemia. The blood-flow response to CSD consisted of a temporary sharp increase in rCBF to more than 400%. At the same time, the concentration of oxyhemoglobin [HbO2] increased, while deoxyhemoglobin [Hb] decreased, indicating excessive oxygenation. Hyperglycemia altered neither the rCBF nor the rCBO response. Preexisting hypercapnia, however, produced reductions in both hyperperfusion (rCBF) and hyperoxygenation (rCBO) during CSD. We found that, for experimental hyperglycemia, i.v. may be superior to i.p. application of glucose because of the latter's side effects in connection with blood flow. Our findings cannot support the hypothesis of a substrate sensing mechanism in coupling.

Role of nitric oxide in the coupling of cerebral blood flow to neuronal activation in rats.

We tested the hypothesis that nitric oxide (NO) is a mediator in the coupling of cerebral blood flow to neuronal activation. The production of NO was blocked in anesthetized rats with the NO-synthase inhibitor N omega-nitro-L-arginine (L-NA). In controls, vibrissae stimulation for 60 s led to a fast (< or = 2 s), 17% increase in regional cerebral blood flow (rCBF) in the contralateral somatosensory cortex. Systemical (10 mg/kg) as well as topical (10(-3) M) application of L-NA reduced the response to stimulation by approximately 50%. Systemical application primarily attenuated the early component of the response, whereas topical application led to an attenuation throughout the whole 60-s stimulation interval. We conclude that NO is involved in rCBF coupling to neuronal activation.

Nitric oxide synthase inhibition does not affect somatosensory evoked potentials in the rat.

Nitric oxide synthase (NOS) inhibition reduces the regional cerebral blood flow (rCBF) responses to somatosensory stimulation. It is controversial whether this is caused by a signalling role of nitric oxide (NO) between neurons and vascular smooth muscle, or by effects of NOS inhibition on neuronal activity. We here report that more than 85% inhibition of NOS activity by topical application of the NOS inhibitor N omega-nitro-L-arginine (L-NNA) for 2 h does not affect somatosensory evoked potentials (SEPs) elicited by vibrissal deflection or electrical forepaw stimulation in choloralose anaesthetised rats equipped with a closed cranial window, whereas cerebral blood flow (CBF) responses due to these stimulation paradigms are reduced by approximately 60%. We conclude that the decrease of the regional vascular response to increased neuronal activity during NOS inhibition is not caused by a suppression of neuronal activity.

Histamine (H1) receptor antagonist inhibits leukocyte rolling in pial vessels in the early phase of bacterial meningitis in rats.

The present study tested the hypothesis whether a histamine dependent pathway is involved in leukocyte-endothel interaction in the early phase of bacterial meningitis. Using confocal laser scanning microscopy we investigated leukocyte rolling in brain venules in vivo during 4 h in experimental pneumococcal meningitis in the rat. Leukocyte rolling, but not firm adhesion induced by intracisternally (i.c.) injected pneumococcal cell wall components, was temporarily inhibited (2 h, 5.6 +/- 1.9 vs. 2.3 +/- 0.9; 3 h, 7.4 +/- 2.7 vs. 3.1 +/- 1.3/100 microm/min) by diphenhydramine, a histamine H1 receptor antagonist. Histamine, possibly released by activated mast cells, is known to initiate P-selectin upregulation and subsequent leukocyte rolling. This data suggest that histamine is a mediator of leukocyte rolling in the early phase of bacterial meningitis.

The cerebrovascular response to elevated potassium--role of nitric oxide in the in vitro model of isolated rat middle cerebral arteries.

We investigated the role of nitric oxide (NO) in the vascular response to high extraluminal K(+)-concentrations in the in vitro model of isolated rat middle cerebral arteries (MCA). Under control conditions, rat MCA dilated at 20, 30, 40 and 60 mM K(+). At 80 mM K(+), a slight vasoconstriction occurred. The unspecific NO synthase (NOS)-inhibitor L(omega)-nitro-L-arginine (L-NNA) increased the resting tone at 3 mM K(+) by 31+/-5% (P<0.01). While the vasodilatative effect of 20 mM K(+) was unaffected by L-NNA, NOS-inhibition resulted in vasoconstriction at > or = 40 mM K(+) (P<0.01). In presence of L-NNA, the basal vessel diameter was restored by either the NO-donor S-nitroso-N-acetylpenicillamine (SNAP) or the cell-permeable guanosine-3',5'-cyclic monophosphate (cGMP) analogue 8-Br-cGMP. Co-application of L-NNA with either SNAP or 8-Br-cGMP resulted in partial restitution of the vasodilatative effect of 40 mM K(+), respectively. In presence of the soluble guanylyl cyclase inhibitor 1 H-[l,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ), the vascular response to 40 mM K(+) was abolished. Our findings together with findings from the literature indicate a modulator role of NO at K(+) > or = 40 mM K(+), involving a cGMP-dependent mechanism.

Products of hemolysis in the subarachnoid space inducing spreading ischemia in the cortex and focal necrosis in rats: a model for delayed ischemic neurological deficits after subarachnoid hemorrhage?

OBJECT: The pathogenesis of delayed ischemic neurological deficits after subarachnoid hemorrhage has been related to products of hemolysis. Topical brain superfusion of artificial cerebrospinal fluid (ACSF) containing the hemolysis products K+ and hemoglobin (Hb) was previously shown to induce ischemia in rats. Superimposed on a slow vasospastic reaction, the ischemic events represent spreading depolarizations of the neuronal-glial network that trigger acute vasoconstriction. The purpose of the present study was to investigate whether such spreading ischemias in the cortex lead to brain damage. METHODS: A cranial window was implanted in 31 rats. Cerebral blood flow (CBF) was measured using laser Doppler flowmetry, and direct current (DC) potentials were recorded. The ACSF was superfused topically over the brain. Rats were assigned to five groups representing different ACSF compositions. Analyses included classic histochemical and immunohistochemical studies (glial fibrillary acidic protein and ionized calcium binding adaptor molecule) as well as a terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling assay. Superfusion of ACSF containing Hb combined with either a high concentration of K+ (35 mmol/L, 16 animals) or a low concentration of glucose (0.8 mmol/L, four animals) reduced CBF gradually. Spreading ischemia in the cortex appeared when CBF reached 40 to 70% compared with baseline (which was deemed 100%). This spreading ischemia was characterized by a sharp negative shift in DC, which preceded a steep CBF decrease that was followed by a slow recovery (average duration 60 minutes). In 12 of the surviving 14 animals widespread cortical infarction was observed at the site of the cranial window and neighboring areas in contrast to findings in the three control groups (11 animals). CONCLUSIONS: The authors conclude that subarachnoid Hb combined with either a high K+ or a low glucose concentration leads to widespread necrosis of the cortex.

Physical model for the spectroscopic analysis of cortical intrinsic optical signals.

We used Monte Carlo simulations and the diffusion approximation to estimate correction terms for the analysis of reflectance spectra of cortical intrinsic optical signals. These corrections depend on scattering and absorption properties, i.e. they are dependent on assumptions on the tissue blood content and oxygen saturation. The analysis was applied to reflectance spectra acquired during whisker barrel stimulation in the rat where attenuation spectra were converted to changes in oxygenated and deoxygenated haemoglobin concentration. The description of the experimental data as judged by the residual and sensitivity to variations of wavelength was considerably improved when the correction terms were included. Inclusion of the correction does have a considerable impact on the time course of deoxyhaemoglobin concentration changes. In contrast to the calculation without correction terms, there is no indication for an early increase in deoxyhaemoglobin ('early dip'). This finding might further current interpretation of the coupling between neuronal activation and oxygen extraction and supply.