2,3-Dihydroxy-6-nitro-7-sulfamoyl-benzo(F)quinoxaline: a neuroprotectant for cerebral ischemia.

2,3-Dihydroxy-6-nitro-7-sulfamoyl-benzo(F)quinoxaline (NBQX) is an analog of the quinoxalinedione antagonists to the non-N-methyl-D-aspartate (non-NMDA) glutamate receptor. NBQX is a potent and selective inhibitor of binding to the quisqualate subtype of the glutamate receptor, with no activity at the NMDA and glycine sites. NBQX protects against global ischemia, even when administered 2 hours after an ischemic challenge.

A task failure has no effect on the electromechanical delay of the peroneus longus.

OBJECTIVE: Ankle inversion injuries represent the most common trauma sustained by athletes. Muscle fatigue from activity may contribute to a delay in the response of the ankle proprioceptors and dynamic restraints during unexpected inversion. The purpose of this investigation was to determine if the electromechanical delay (EMD) of the peroneus longus is influenced by a task failure exercise. SUBJECTS: Sixteen subjects (age 20 +/- 1.1 y; mass 71.6 +/- 12.5 kg; height 173.0 +/- 8.7 cm; 9 male, 1 female) with no lower extremity injuries reported for data collection. MEASUREMENTS: Data were collected from each subject's dominant leg using surface electromyography (EMG). Electrodes were applied over the peroneus longus (PL) using a standard protocol. A stimulating electrode was applied to the common peroneal nerve. Subjects were placed in a monopedal stance on a force platform. A low amplitude, short duration stimulus was applied to the common peroneal nerve. The EMG was used to determine timing of the M wave and the force platform was used to determine the onset of foot pronation. Once 6 trials were recorded, subjects completed 2 sets of an isotonic activity that isolated the peroneals. The task was completed to failure for each set. Immediately following the task failure exercise, subjects returned to the force platform for 6 additional trials recorded as before. Analysis of data was performed by determining the onset of the M wave as the beginning of positive EMG activity following the end of the imposed stimulus response. This point was superimposed on the force platform curve and the point at which a 10 N.m force change occurred was used to calculate the EMD (time difference between the force platform indicator and the M wave indicator). RESULTS: Average EMD prior to the task failure exercise was 13.35 +/- 3.47 ms. Following the task failure exercise, the average EMD was 12.67 +/- 3.86 ms. A paired samples t test revealed no significant differences with regard to EMD between pre- and post-task failure exercise for the PL (p = 0.448). CONCLUSION: We concluded that the task failure exercise did not affect the electromechanical delay of the PL.

Contaminating viral sequences in high-throughput sequencing viromics: a linkage study of 700 sequencing libraries.

OBJECTIVES: Sample preparation for high-throughput sequencing (HTS) includes treatment with various laboratory components, potentially carrying viral nucleic acids, the extent of which has not been thoroughly investigated. Our aim was to systematically examine a diverse repertoire of laboratory components used to prepare samples for HTS in order to identify contaminating viral sequences. METHODS: A total of 322 samples of mainly human origin were analysed using eight protocols, applying a wide variety of laboratory components. Several samples (60% of human specimens) were processed using different protocols. In total, 712 sequencing libraries were investigated for viral sequence contamination. RESULTS: Among sequences showing similarity to viruses, 493 were significantly associated with the use of laboratory components. Each of these viral sequences had sporadic appearance, only being identified in a subset of the samples treated with the linked laboratory component, and some were not identified in the non-template control samples. Remarkably, more than 65% of all viral sequences identified were within viral clusters linked to the use of laboratory components. CONCLUSIONS: We show that high prevalence of contaminating viral sequences can be expected in HTS-based virome data and provide an extensive list of novel contaminating viral sequences that can be used for evaluation of viral findings in future virome and metagenome studies. Moreover, we show that detection can be problematic due to stochastic appearance and limited non-template controls. Although the exact origin of these viral sequences requires further research, our results support laboratory-component-linked viral sequence contamination of both biological and synthetic origin.

Gap-junction-mediated propagation and amplification of cell injury.

Gap junctions are conductive channels that connect the interiors of coupled cells. We determined whether gap junctions propagate transcellular signals during metabolic stress and whether such signaling exacerbates cell injury. Although overexpression of the human proto-oncogene bcl2 in C6 glioma cells normally increased their resistance to injury, the relative resistance of bcl2+ cells to calcium overload, oxidative stress and metabolic inhibition was compromised when they formed gap junctions with more vulnerable cells. The likelihood of death was in direct proportion to the number and density of gap junctions with their less resistant neighbors. Thus, dying glia killed neighboring cells that would otherwise have escaped injury. This process of glial 'fratricide' may provide a basis for the secondary propagation of brain injury in cerebral ischemia.

Hand movements in laparoscopic suturing: a simple vector analysis.

BACKGROUND: Laparoscopic suturing is a complex task that is vital to the performance of many advanced laparoscopic procedures. Mastery can be difficult and problematic for surgical trainees. METHODS: We present a description of hand movements in laparoscopic suturing. Complex maneuvers are simplified into linear motions using vectors. The analysis is intended to be a tool for training in the art of laparoscopic surgery. RESULTS: Linear hand movements in the x and y axes produce opposite motions at the instrument tip. Position along the z axis influences the extent of hand movement relative to the instrument tip. Rotational movements of the hand produce an equal rotation of the instrument tip. Revolution is a complex motion that combines movements in x and y axes. Vector analysis reveals that the arc of revolution must be reversed to produce the desired needle motion. CONCLUSIONS: A conceptual understanding of hand-movement vectors facilitates the efficient mastery of the complex skills required for laparoscopic suturing.

Effects of a task failure exercise on the peroneus longus and brevis during perturbed gait.

Ankle inversion injuries represent the most common trauma sustained by athletes. Muscle fatigue from activity may contribute to a delay in the response of the ankle proprioceptors and dynamic restraints during unexpected inversion. The purpose of this investigation was to determine changes in peroneal average EMG, peak EMG, and time to peak EMG following a task failure exercise. Thirty-two subjects (age 20+/-1.43 yrs; 21 male, 11 female) with no lower extremity injuries reported for data collection. Data were collected from each subject's dominant leg using surface electromyography (EMG). EMG electrodes were applied over the peroneus longus (PL) and brevis (PB) using a standard protocol Subjects walked at a fixed pace on a 6.1 m runway with one section that could be unexpectedly dropped into 30 degrees of inversion upon foot contact. Trials with perturbed and unperturbed gait were randomized to reduce prediction of the unexpected inversion. Once 3 trials of perturbed gait were recorded, subjects completed an isotonic activity that isolated the peroneals. The task was completed to failure. Immediately following the task failure exercise, subjects walked on the perturbation runway once again until 3 trials of perturbed gait were recorded. Analysis revealed no significant differences with regard to average muscle activity between pre- and post-task failure exercise for the PL (F1,31 = 0.133; p = 0.718) or for the PB (F1,31 = 0.795; p = 0.380). There was also no significant difference in peak muscle activity pre- to post-task failure for the PL (F1,31 = 0.032; p = 0.859) or the PB (F1,31 = 0.156; p = 0.695). Finally, there was no significant difference in time-to-peak muscle activity pre- to post-task failure for the PL (F1,31 = 0.830; p = 0.369) or the PB (F1,31 = 1.037; p = 0.316). We concluded that the task failure exercise did not contribute to changes in peroneal activity during perturbed gait. These results indicate that peroneal fatigue does not play a significant role in the incidence of inversion ankle sprains.

Tissue specific differences in the regulation of the UDP glucuronosyltransferase 2B17 gene promoter.

The human UDP glucuronosyltransferase UGT2B17, glucuronidates androgens and is expressed in the liver and the prostate. Although evidence suggests that variations in UGT2B17 expression between tissues may be a critical determinant of androgen response, the factors that regulate UGT2B17 expression in the liver and prostate are unknown. In this study, we have isolated a 596 bp promoter of the UGT2B17 gene and studied its regulation in the liver cell line, HepG2 and the prostate cell line, LNCaP. The transcription start site of UGT2B17 was mapped and proteins that bound to the proximal promoter were detected by DNase1 footprint analysis. A region (-40 to -52 bp) which resembled a hepatocyte nuclear factor 1 (HNF1) binding site bound proteins in nuclear extracts from HepG2 cells, but did not bind proteins from LNCaP nuclear extracts. In HepG2 cells, HNF1alpha bound to this region and activated the UGT2B17 promoter, as assessed by functional and gel shift assays. HNF1alpha activation of the promoter was prevented by mutation or deletion of the putative HNF1 site. The related transcription factor HNF1beta, which is present in HepG2 cells, did not activate the promoter. The UGT2B17 promoter could also be activated by exogenous HNF1alpha in LNCaP cells. However, because these cells do not contain HNF1alpha, other transcription factors must regulate the UGT2B17 promoter. Cotransfection experiments showed that HNF1beta, elevates promoter activity in LNCaP cells. This activation did not involve the putative HNF1 region (-40 to -52 bp) since mutation of this region did not affect promoter activation by HNF1beta. These results suggest that the UGT2B17 promoter is regulated by different factors in liver-derived HepG2 and prostate-derived LNCaP cells.

Characterization of cortical depolarizations evoked in focal cerebral ischemia.

Cortical tissue surrounding acute ischemic infarcts undergoes repetitive spontaneous depolarizations. It is unknown whether these events are episodes of spreading depression (SD) elicited by the elevated interstitial K+ ([K+]e) in the ischemic core or whether they are evoked by transient decreases of the local blood flow. Electrophysiologically, depolarization caused by SD or by ischemia (ID) can be distinguished by their characteristic patterns of [K+]e rise: During SD, [K+]e rises abruptly, while in ID, this fast rate of increase is preceded by a slow rate lasting minutes. To characterize the depolarizations, we occluded the right middle cerebral artery (MCA) in rats and inserted two K(+)-sensitive microelectrodes into the cortex surrounding the evolving infarct. Repeated increases in [K+]e arose spontaneously following MCA occlusion. [K+]e increased during these transients from a resting level of 3-6 to 60 mM. One-third of these transient increases in [K+]e were biphasic, consisting of a slow initial increase to 10-12 mM, which lasted for minutes, followed by an abrupt increase, a pattern characteristic of ID. The remaining two-thirds exhibited a steep monotonic increase in [K+]e (< 10 s), characteristic of SD. The duration of the transients was a function of the pattern of [K+]e increase: ID-like transients lasted an average 10.7 +/- 5.1 min, whereas the duration of SD-like transients was 5.7 +/- 3.4 min. Both types of K+ transients occurred in an apparently random fashion in individual animals. A K+ transient was never observed solely at one electrode.(ABSTRACT TRUNCATED AT 250 WORDS)

The effect of glutamate receptor blockade on anoxic depolarization and cortical spreading depression.

We examined the effect of blockade of N-methyl-D-aspartate (NMDA) and non-NMDA subtype glutamate receptors on anoxic depolarization (AD) and cortical spreading depression (CSD). [K+]e and the direct current (DC) potential were measured with microelectrodes in the cerebral cortex of barbiturate-anesthetized rats. NMDA blockade was achieved by injection of (+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine maleate [MK-801; 3 and 10 mg/kg] or amino-7-phosphonoheptanoate (APH; 4.5 and 10 mg/kg). Non-NMDA receptor blockade was achieved by injection of 2,3-dihydroxy-6-nitro-7-sulfamoylbenzo(F)quinoxaline (NBQX; 10 and 20 mg/kg). MK-801 and APH blocked CSD, while NBQX did not. In control rats, the latency from circulatory arrest to AD was 2.1 +/- 0.1 min, while the amplitude of the DC shift was 21 +/- 1 mV, and [K+]e increased to 50 +/- 6 mM. All variables remained unchanged in animals treated with MK-801, APH, or NBQX. Finally, MK-801 (14 mg/kg) and NBQX (40 mg/kg) were given in combination to examine the effect of total glutamate receptor blockade on AD. This combination slightly accelerated the onset of AD, probably owing to circulatory failure. In conclusion, AD was unaffected by glutamate receptor blockade. In contrast, NMDA receptors play a crucial role for CSD.

Extracellular pH changes during spreading depression and cerebral ischemia: mechanisms of brain pH regulation.

We have examined the extracellular pH (pHe) during spreading depression and complete cerebral ischemia in rat parietal cortex utilizing double-barrelled H+ liquid ion exchanger microelectrodes. The baseline pHe of the parietal cortex was 7.33 at a mean arterial PCO2 of 38 mm Hg. Following spreading depression and cerebral ischemia, highly reproducible triphasic changes in pHe occurred, which were intimately related to the negative deflection in tissue potential (Ve). The changes in pHe for spreading depression (n = 23) were a small initial acidic shift, beginning before the rapid change in Ve, followed by a rapid transient alkaline shift of 0.16 pH units, the onset of which coincided with the negative deflection in Ve. A prolonged acidic shift of 0.42 pH units then occurred. The maximal decrease in pHe was to 6.97 and the mean duration of the triphasic pHe change was 7.8 min. The lactate concentration in brain cortex increased from baseline 1.2 mM to 7.0 mM (n = 6) during the maximal acidic change in spreading depression. In addition, lactate levels correlated well with resolution of the pHe changes during spreading depression. The triphasic pHe changes following complete cerebral ischemia were an initial acidic shift of 0.43 pH units which developed over 2 min, then an alkaline shift of 0.10 pH units coincident with the negative deflection in Ve, and a final acidic shift of 0.26 pH units. The terminal pHe was 6.75. Superfusion of the cortex with inhibitors of carbonic anhydrase (acetazolamide), Na+/H+ counter transport (amiloride), and Cl-/HCO-3 countertransport (4,4'-diisothiocyanostilbene-2,2'-disulfonic acid) altered the triphasic pHe changes in a similar fashion for both spreading depression and cerebral ischemia, providing insights into the pHe regulatory mechanisms in mammalian brain.

Correlation between N-acetylaspartate levels and histopathologic changes in cortical infarcts of mice after middle cerebral artery occlusion.

The aim of the present study was to evaluate the use of the endogenous neuronal compound N-acetylaspartate (NAA) as a marker of neuronal damage after focal cerebral ischemia in mice. After occlusion of the middle cerebral artery (MCAO) the ischemic cortex was sampled, guided by 2,3,5-triphenyltetrazolium chloride (TTC) staining, and the NAA concentration was measured by high-pressure liquid chromatography (HPLC). Conventional histology and immunohistological methods using antibodies against neuron-specific enolase (NSE), neurofilaments (NF), synaptophysin, glial fibrillary acidic protein (GFAP), and carbodiamide-linked NAA and N-acetylaspartylglutamate (NAAG). The level of NAA rapidly declined to 50% and 20% of control levels in infarcted tissue after 6 hours and 24 hours, respectively. No further decrease was observed during the observation period of 1 week. Within the first 6 hours the number of normal-appearing neurons in the infarcted cortical tissue decreased to 70% of control, of which the majority were eosinophilic. After 24 hours almost no normal-appearing neurons were seen. The number of eosinophilic neurons decreased steadily to virtually zero after 7 days. The number of immunopositive cells in the NSE, NF, and synaptophysin staining within the infarct was progressively reduced, and after 3 to 7 days the immunoreactions were confined to discrete granulomatous structures in the center of the infarct, which otherwise was infested with macrophages. This granulomatous material also stained positive for NAA. The number of cells with positive GFAP immunoreactions progressively increased in the circumference of the infarct. They also showed increased immunoreaction against NAA and NSE. The study shows that the level of NAA 7 days after ischemia does not decline to zero but remains at 10% to 20% of control values. The fact NAA is trapped in cell debris and NAA immunoreactivity is observed in the peri-infarct areas restricts its use as a marker of neuronal density.

Changes in N-acetyl-aspartate content during focal and global brain ischemia of the rat.

N-Acetyl-aspartate (NAA) is almost exclusively localized in neurons in the mature brain and might be used as a neuronal marker. It has been reported that the NAA content in human brain is decreased in neurodegenerative diseases and in stroke. Since the NAA content can be determined by nuclear magnetic resonance techniques, it has potential as a diagnostic and prognostic marker. The objective of this study was to examine the change of NAA content and related substances following cerebral ischemia and compare the results to the damage of the tissue. We used rats to study the changes of NAA, N-acetyl-aspartyl-glutamate (NAAG), glutamate, and aspartate contents over a time course of 24 h in brain regions affected by either permanent middle cerebral artery occlusion (focal ischemia) or decapitation (global ischemia). The decreases of NAA and NAAG contents following global brain ischemia were linear over time but significant only after 4 and 2 h, respectively. After 24 h, the levels of NAA and NAAG were 24 and 44% of control values, respectively. The concentration of glutamate did not change, whereas the aspartate content increased at a rate comparable with the rate of decrease of NAA content. This is consistent with NAA being preferentially degraded by the enzyme amidohydrolase II in global ischemia. In focal ischemia, there was a rapid decline of NAA within the first 8 h of ischemia followed by a slower rate of reduction. The reductions of NAA and NAAG contents in focal ischemia were significant after 4 and 24 h, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Extracellular pH in the rat brain during hypoglycemic coma and recovery.

It has previously been shown that hypoglycemic coma is accompanied by marked energy failure and by loss of cellular ionic homeostasis. The general proposal is that shortage of carbohydrate substrate prevents lactic acid formation and thereby acidosis during hypoglycemic coma. The objective of the present study was to explore whether rapid downhill ion fluxes, known to occur during coma, are accompanied by changes in extra- and/or intracellular pH (pHe and/or pHi), and how these relate to the de- and repolarization of cellular membranes. Cortical pHe was recorded by microelectrodes in insulin-injected rats subjected to 30 min of hypoglycemic coma, with cellular membrane depolarization. Some rats were allowed up to 180 min of recovery after glucose infusion and membrane repolarization. Arterial blood gases and physiological parameters were monitored to maintain normotension, normoxia, normocapnia, and normal plasma pH. Following depolarization during hypoglycemia, a prompt, rapidly reversible alkaline pHe shift of about 0.1 units was observed in 37/43 rats. Immediately thereafter, all rats showed an acid pH shift of about 0.2 units. This shift developed during the first minute, and pHe remained at that level until repolarization was induced. Following repolarization, there was an additional, rapid, further lowering of pHe by about 0.05 units, followed by a more prolonged decrease in pHe that was maximal at 90 min of recovery (delta pHe of approximately -0.4 units). The pHe then slowly normalized but was still decreased (-0.18 pH units) after 180 min when the experiment was terminated. The calculated pHi showed no major alterations during hypoglycemic coma or after membrane repolarization following glucose administration.(ABSTRACT TRUNCATED AT 250 WORDS)

Influence of MK-801 on brain extracellular calcium and potassium activities in severe hypoglycemia.

The purpose of the present study was to examine the effect of blockade of N-methyl-D-aspartate (NMDA) receptors on the depolarization associated with severe hypoglycemia, which is commonly preceded by one or a few transient depolarizations reminiscent of cortical spreading depression (CSD). In the cerebral cortices of rats [K+]e and [Ca2+]e were measured with ion-selective microelectrodes. NMDA blockade was achieved by injection of MK801 in doses that block CSD. In control rats, the latency from the time point when blood glucose reached minimal levels to onset of ionic shifts was 33.2 +/- 3.5 min, and [K+]e rose from 3.2 +/- 0.2 to 55 +/- 5 mM. All variables remained unchanged in rats treated with MK801. In another four rats treated with MK801, [Ca2+]e declined from 1.06 +/- 0.22 to 0.12 +/- 0.02 mM. Plasma glucose measurements indicated that the cortex depolarized at a plasma glucose concentration between 0.7 and 0.8 mM, i.e., within a narrow range, suggesting a threshold phenomenon. In conclusion, activation of NMDA receptors seems of minor importance for hypoglycemic depolarization. The ionic transients that precede the persistent hypoglycemic depolarization are probably mediated by mechanisms distinct from those of electrically induced CSD.

Seizure-induced damage to substantia nigra and globus pallidus is accompanied by pronounced intra- and extracellular acidosis.

Status epilepticus of greater than 30-min duration in rats gives rise to a conspicuous lesion in the substantia nigra pars reticulata (SNPR) and globus pallidus (GP). The objective of the present study was to explore whether the lesion, which encompasses necrosis of both neurons and glial cells, is related to intra- and extracellular acidosis. Using the flurothyl model previously described to produce seizures, we assessed regional pH values with the autoradiographic 5,5-dimethyl[2-14C]oxazolidine-2,4-dione technique. Regional pH values were assessed in animals with continuous seizures for 20 and 60 min, as well as in those allowed to recover for 30 and 120 min after seizure periods of 20 or 60 min. In additional animals, changes in extracellular fluid pH (pHe) were measured with ion-selective microelectrodes, and extracellular fluid (ECF) volume was calculated from the diffusion profile for electrophoretically administered tetramethylammonium. In structures such as the neocortex and the hippocampus, which show intense metabolic activation during seizures, status epilepticus of 20- and 60-min duration was accompanied by a reduction of the "composite" tissue pH (pHt) of 0.2-0.3 unit. Recovery of pHt was observed upon termination of seizures. In SNPR and in GP, the acidosis was marked to excessive after 20 and 60 min of seizures (delta pHt approximately 0.6 after 60 min).(ABSTRACT TRUNCATED AT 250 WORDS)

Cortical spreading depression is associated with arachidonic acid accumulation and preservation of energy charge.

The present study aimed to study the relation between the release of arachidonic acid (AA) and the energy state in cerebral cortices of rats during single episodes of cortical spreading depression (CSD). The changes in concentrations of AA, labile phosphate compounds [ATP, ADP, AMP, and phosphocreatine (PCr)], and glycolytic metabolites (lactate, pyruvate, glucose, and glycogen) were studied during and following the large change of the local direct current (DC) potential. Free AA increased markedly during the DC shift, continued to increase during the subsequent 3 min, and returned to control levels at 4-5 min after CSD. PCr decreased by 38% in the first minutes following the DC shift, while ADP increased by 38%. Both returned to normal within a few minutes. ATP, AMP, and energy charge remained constant throughout the experimental period. Glucose decreased by 47% and glycogen by 34% for a few minutes following CSD, while lactate increased by 105% at 2-3 min and by 77% at 4-5 min after CSD. The metabolites returned to control levels at 10 min after CSD. Considering the constant energy charge at all time points during CSD, it is suggested that the AA rise reflects augmented phospholipase activity due to either increased intracellular [Ca2+] or receptor stimulation or both. The possibility that N-methyl-D-aspartate receptors play a role in the release of AA, and that free AA in turn could be part of the mechanism of CSD, is discussed.

Calcium movements in traumatic brain injury: the role of glutamate receptor-operated ion channels.

Ion-selective microelectrodes were used to study acute effects of N-methyl-D-aspartate (NMDA) and alpha-amino-3-hydroxy- 5-methyl-4-isoxazole (AMPA) receptor blockade on posttraumatic calcium disturbances. An autoradiographic technique with 45 Ca2+ was used to study calcium disturbances at 8, 24, and 72 h. Compression contusion trauma of the cerebral cortex was produced by a 21-g weight dropped from a height of 35 cm onto a piston that compressed the brain 2 mm. Pre- and posttrauma interstitial [Ca2+] ([Ca2+]e) concentrations were measured in the perimeter, i.e., the shear stress zone (SSZ) and in the central region (CR) of the trauma site. For the [Ca2+]e studies the animals were divided into controls and groups pretreated with dizocilipine maleate (MK-801) or with 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo[F]quinoxaline (NBQX). In all groups, [Ca2+]e decreased from pretrauma values of approximately 1 mM to posttraumatic values of 0.1 mM in both the CR and the SSZ. This was followed by a slow restitution toward pretraumatic levels during the 2-h observation period. There was no significant difference in recovery pattern between controls and pretreated animals. Accumulation of 45Ca2+ and serum proteins was seen in the entire SSZ, while neuronal necrosis was confined to a narrow band within the SSZ. The CR was unaffected apart from occasional eosinophilic neurons and showed no accumulation of 45Ca2+. Posttraumatic treatment with MK-801 or NBQX had no obvious effect on neuronal injury in the SSZ. We conclude that (a) acute [Ca2+]e disturbances in compression contusion brain trauma are not affected by blockade of NMDA or AMPA receptors, (b) 45Ca2+ accumulation in the SSZ reflects mainly protein accumulation due to blood-brain barrier breakdown rather than cell death, and (c) acute cellular Ca2+ over-load per se does not seem to be a major determinant of cell death after cerebral trauma in our model.

Regional changes in interstitial K+ and Ca2+ levels following cortical compression contusion trauma in rats.

Brain trauma is associated with acute functional impairment and neuronal injury. At present, it is unclear to what extent disturbances in ion homeostasis are involved in these changes. We used ion-selective microelectrodes to register interstitial potassium ([K+]e) and calcium ([Ca2+]e) concentrations in the brain cortex following cerebral compression contusion in the rat. The trauma was produced by dropping a 21 g weight from a height of 35 cm onto a piston that compressed the cortex 1.5 mm. Ion measurements were made in two different locations of the contused region: in the perimeter, i.e., the shear stress zone (region A), and in the center (region B). The trauma resulted in an immediate increase in [K+]e from a control level of 3 mM to a level > 60 mM in both regions, and a concomitant negative shift in DC potential. In both regions, there was a simultaneous, dramatic decrease in [Ca2+]e from a baseline of 1.1 mM to 0.3-0.1 mM. Interstitial [K+] and the DC potential normalized within 3 min after trauma. In region B, [Ca2+]e recovered to near control levels within 5 min after ictus. In region A, however, recovery of [Ca2+]e was significantly slower, with a return to near baseline values within 50 min after trauma. The prolonged lowering of [Ca2+]e in region A was associated with an inability to propagate cortical spreading depression, suggesting a profound functional disturbance. Histologic evaluation 72 h after trauma revealed that neuronal injury was confined exclusively to region A. The results indicate that compression contusion trauma produces a transient membrane depolarization associated with a pronounced cellular release of K+ and a massive Ca2+ entry into the intracellular compartment. We suggest that the acute functional impairment and the subsequent neuronal injury in region A is caused by the prolonged disturbance of cellular calcium homeostasis mediated by leaky membranes exposed to shear stress.

Brain acidosis in experimental pneumococcal meningitis.

Purulent meningitis is a serious disease that often has a lethal outcome or gives lasting complications due to brain damage. The processes causing brain dysfunction or damage are still not uncovered nor are the reasons for the characteristic increase of CSF lactate, or the decrease of glucose levels and of pH. We studied rabbits with experimentally induced purulent meningitis (Streptococcus pneumoniae). Ten hours after the inoculation into cisterna magna the rabbits developed symptoms of meningitis, with stiffness of the neck, tachypnea, and fever. The CSF level of lactate and the number of leukocytes were significantly increased and the glucose level was decreased. Brain interstitial pH, as measured by ion selective microelectrodes, was significantly decreased from the normal level of 7.4 to 6.9. The levels of energy metabolites in brain cortex, including glucose, were not different between controls and infected animals, and the lactate level was not elevated more than could have been explained by passive diffusion from the CSF. This shows that the brain tissue is not the source of CSF lactate nor the sink for glucose in CSF. The marked acidification of brain interstitial space and CSF demonstrates that purulent meningitis causes a significant disturbance of brain ion homeostasis that could be, at least in part, responsible for the brain dysfunction. We suggest that activated leukocytes consume CSF glucose and produce lactic acid and secrete protons, which causes the CSF and interstitial acidosis.

N-Acetylaspartate distribution in rat brain striatum during acute brain ischemia.

Brain N-acetylaspartate (NAA) can be quantified by in vivo proton magnetic resonance spectroscopy (1H-MRS) and is used in clinical settings as a marker of neuronal density. It is, however, uncertain whether the change in brain NAA content in acute stroke is reliably measured by 1H-MRS and how NAA is distributed within the ischemic area. Rats were exposed to middle cerebral artery occlusion. Preischemic values of [NAA] in striatum were 11 mmol/L by 1H-MRS and 8 mmol/kg by HPLC. The methods showed a comparable reduction during the 8 hours of ischemia. The interstitial level of [NAA] ([NAA]e) was determined by microdialysis using [3H]NAA to assess in vivo recovery. After induction of ischemia, [NAA]e increased linearly from 70 micromol/L to a peak level of 2 mmol/L after 2 to 3 hours before declining to 0.7 mmol/L at 7 hours. For comparison, [NAA]e was measured in striatum during global ischemia, revealing that [NAA]e increased linearly to 4 mmol/L after 3 hours and this level was maintained for the next 4 h. From the change in in vivo recovery of the interstitial space volume marker [14C]mannitol, the relative amount of NAA distributed in the interstitial space was calculated to be 0.2% of the total brain NAA during normal conditions and only 2 to 6% during ischemia. It was concluded that the majority of brain NAA is intracellularly located during ischemia despite large increases of interstitial [NAA]. Thus, MR quantification of NAA during acute ischemia reflects primarily changes in intracellular levels of NAA.