Effects of glucose load on brain extracellular lactate concentration in conscious rats using a microdialysis technique

Changes in the brain lactate concentration in cerebral extracellular fluid (ECF) during intravenous infusion of glucose and local administration of glucose were investigated in adult, conscious, unrestrained rats, with a microdialysis probe in the posterior hippocampus. The rats were infused intravenously with either 25% sucrose solution or 25% glucose solution at a rate of 16.6 microliters.min-1.100 g-1 for three hours. The blood glucose concentration reached 17.0 +/- 2.6 mM at the end of the glucose infusion, and brain ECF glucose showed a parallel change with the blood glucose concentration and increased to 2.37 +/- 0.30 mM. However, blood and brain ECF glucose concentrations did not change in animals infused with the sucrose solution. On the other hand, the blood lactate concentration in the glucose-infused group also increased from 0.93 +/- 0.18 mM to 2.85 +/- 0.39 mM at the end of the glucose infusion, which was significantly higher than that measured in the sucrose-infused group. The blood lactate level in the glucose-infused group returned to the basal level by the end of the experiment. Brain ECF lactate concentrations increased from 1.21 +/- 0.06 mM to 1.69 +/- 0.11 mM in glucose-infused animals, but did not change in the sucrose-infused animals. The brain ECF lactate concentration showed a positive correlation with the brain ECF glucose concentration in glucose-infused animals. Another group of rats was administered glucose locally for 90 min after substitution of artificial cerebrospinal fluid.(ABSTRACT TRUNCATED AT 250 WORDS)     

Interaction between Ca2+, K+, carbamazepine and zonisamide on hippocampal extracellular glutamate monitored with a microdialysis electrode

1. Multiple components of hippocampal glutamate release were examined by study of Ca2+- and K+-evoked hippocampal extracellular glutamate release using an in vivo microdialysis glutamate biosensor in urethane-anaesthetized rats. In addition, the effects of the antiepileptic drugs, carbamazepine (CBZ) and zonisamide (ZNS) perfused through the probe on glutamate release were assessed. 2. Basal glutamate levels were below detection limits (approximately 0.1 microM). An increase in extracellular KCl (from 2.7 to 50 and 100 mM) increased extracellular hippocampal glutamate levels to 9.2+/-1.4 and 20.0+/-2.6 microM, respectively, calculated from the area under curve (AUC) for 60 min. 3. This KCl-evoked glutamate release consisted of three components: an initial transient rise, a late gentle rise, and late multiple phasic transient rises. 4. An increase in or removal of extracellular CaCl2 levels respectively enhanced and reduced the 50 mM KCl-evoked hippocampal glutamate release (AUC for 60 min) from 9.2+/-1.4 to 12.4+/-2.1 and 5.8+/-0.9 microM. 5. Perfusion with 100 microM CBZ or 1 mM ZNS inhibited both the 50 mM KCl-evoked hippocampal glutamate release (AUC for 60 min) from 9.2+/-1.4 to 5.5+/-1.1 and to 5.8+/-1.3 microM, respectively, as well as the stimulatory effects of Ca2+ on KCl-evoked hippocampal glutamate release. 6. These results suggest that both CBZ and ZNS may reduce epileptiform events by inhibiting excitatory glutamatergic transmission.     

Post-traumatic brain hypothermia reduces histopathological damage following concussive brain injury in the rat

The purposes of this study were (1) to document the histopathological consequences of moderate traumatic brain injury (TBI) in anesthetized Sprague-Dawley rats, and (2) to determine whether post-traumatic brain hypothermia (30 degrees C) would protect histopathologically. Twenty-four hours prior to TBI, the fluid percussion interface was positioned over the right cerebral cortex. On the 2nd day, fasted rats were anesthetized with 70% nitrous oxide, 1% halothane, and 30% oxygen. Under controlled physiological conditions and normothermic brain temperature (37.5 degrees C), rats were injured with a fluid percussion pulse ranging from 1.7 to 2.2 atmospheres. In one group, brain temperature was maintained at normothermic levels for 3 h after injury. In a second group, brain temperature was reduced to 30 degrees C at 5 min post-trauma and maintained for 3 h. Three days after TBI, brains were perfusion-fixed for routine histopathological analysis. In the normothermic group, damage at the site of impact was seen in only one of nine rats. In contrast, all normothermic animals displayed necrotic neurons within ipsilateral cortical regions lateral and remote from the impact site. Intracerebral hemorrhagic contusions were present in all rats at the gray-white interface underlying the injured cortical areas. Selective neuronal necrosis was also present within the CA3 and CA4 hippocampal subsectors and thalamus. Post-traumatic brain hypothermia significantly reduced the overall sum of necrotic cortical neurons (519 +/- 122 vs 952 +/- 130, mean +/- SE, P = 0.03, Kruskal-Wallis test) as well as contusion volume (0.50 +/- 0.14 vs 2.14 +/- 0.71 mm3, P = 0.004).(ABSTRACT TRUNCATED AT 250 WORDS)     

In vivo voltammetric detection of rat brain lactate with carbon fiber microelectrodes coated with lactate oxidase

To allow rat brain lactate measurement in vivo, a specific sensor based on a carbon fiber (phi = 30 microns) microelectrode coated with lactate oxidase was prepared. Combined with the differential normal pulse voltammetry measurement method, such a sensor, with a sensitivity of 9.15 +/- 0.91 mA.M-1.cm-2, provided a lactate linear response in concentrations ranging from 0.1 to 2.0 mM. The measurements performed appeared to be essentially insensitive to usual interference caused by the electroactive compounds present in the brain (ascorbic acid and peptides). In vivo detection performed in the cortex of the anesthetized rat led to the determination of a lactate concentration of 0.41 +/- 0.02 mM. Moreover, to validate the results obtained in vivo, an ex vivo determination of the lactate level was also performed in samples of brain tissue, plasma, and cerebrospinal fluid, using both voltammetry and a clinical analyzer with colorimetric-based detection. A good correlation was observed between the sets of data established by both methods.     

Induction of HLA class II in HTLV-I infected neuronal cell lines

Prenatal methylazoxymethanol acetate (MAMac) injection disrupts cell migration in developing rats. We investigated the electrophysiological characteristics of hippocampal CA1 pyramidal neurons from young MAMac-treated animals (postnatal days 25-35). In vitro intracellular recordings from CA1 cells in MAMac-treated tissue revealed resting membrane potential (mean, -61.5 +/- 1.5 mV), action potential amplitude (mean, 69 +/- 3.1 mV), action potential duration (mean, 2.1 +/- 0.2 ms), input resistance (mean, 51.5 +/- 3.6 M omega) and time constant (mean, 33.2 +/- 1.2 ms) similar to those of CA1 cells from control tissue. However, MAMac-treated tissue could be distinguished as having a higher percentage of cells (62% vs. 10%) which fire a burst of action potentials in response to suprathreshold current injection. The synaptic responses of CA1 cells in MAMac-treated and control tissue were comparable. The CA1 field response to stimulation was also comparable at all stimulus intensities tested (50-1500 microA). Elevation of extracellular potassium concentration ([K+]o) from 3 mM to 6 mM resulted in epileptiform discharge activity in response to stratum radiatum stimulation in all MAMac-treated slices (10/10) but in only one-third of controls (3/9). Spontaneous epileptiform discharges were also observed in the majority (8/13) of MAMac-treated slices bathed in 6 mM KCl but in no controls. These data suggest that MAMac treatment during fetal development not only disrupts normal anatomical organization but also leads to alterations in electrophysiological features of the hippocampal CA1 pyramidal cell region. As such, the MAMac model may provide insights into early onset seizure syndromes associated with developmental abnormalities.     

Steady-state cerebral glucose concentrations and transport in the human brain

Understanding the mechanism of brain glucose transport across the blood-brain barrier is of importance to understanding brain energy metabolism. The specific kinetics of glucose transport have been generally described using standard Michaelis-Menten kinetics. These models predict that the steady-state glucose concentration approaches an upper limit in the human brain when the plasma glucose level is well above the Michaelis-Menten constant for half-maximal transport, Kt. In experiments where steady-state plasma glucose content was varied from 4 to 30 mM, the brain glucose level was a linear function of plasma glucose concentration. At plasma concentrations nearing 30 mM, the brain glucose level approached 9 mM, which was significantly higher than predicted from the previously reported Kt of approximately 4 mM (p < 0.05). The high brain glucose concentration measured in the human brain suggests that ablumenal brain glucose may compete with lumenal glucose for transport. We developed a model based on a reversible Michaelis-Menten kinetic formulation of unidirectional transport rates. Fitting this model to brain glucose level as a function of plasma glucose level gave a substantially lower Kt of 0.6 +/- 2.0 mM, which was consistent with the previously reported millimolar Km of GLUT-1 in erythrocyte model systems. Previously reported and reanalyzed quantification provided consistent kinetic parameters. We conclude that cerebral glucose transport is most consistently described when using reversible Michaelis-Menten kinetics.     

Two distinct subgroups of cholecystokinin-immunoreactive cortical interneurons

Sustained high levels of corticosterone (CORT), one of the major stress-induced hormones in the rat, were suggested as generating 'accelerated brain aging' and were shown to induce both specific brain changes in the hippocampus and learning impairments in young and middle-aged Fischer-344 rats. Evidence that altered calcium (Ca) homeostasis may play a major role in brain aging has accumulated over the last decade. Recently, new data established a connection between glucocorticoids and voltage-activated Ca influx in aged hippocampal neurons. In the present study, an attempt was made to block the CORT-induced 'accelerated aging' by the simultaneous administration of the L-type Ca channel blocker nimodipine. CORT or placebo sustained-release (SR) pellets were implanted subcutaneously in 3 months old Fischer male rats. Each group was further sub-divided between nimodipine and placebo SR treatments. Characteristic CORT-induced morphological changes were observed in pyramidal hippocampal cells, such as at the CA1 and CA4 sub-regions (22.2% +/- 7.7 and 28.6% +/- 8.4 of pyknotic cells without clear nuclei, respectively). Concomitant treatment with nimodipine conferred full protection against CORT-induced morphological changes (e.g. 3.2% +/- 0.8 and 2.1% +/- 1.9 of pyknotic cells in CA1 and CA4, n = 7 rats in each group; P < 0.04). The neuroprotective efficacy of nimodipine supports the theory of Ca involvement in CORT related 'accelerated brain aging'.     

Analysis of ceftriaxone and ceftazidime distribution in cerebrospinal fluid of and cerebral extracellular space in awake rats by in vivo microdialysis

In vivo microdialysis was used to estimate the extracellular concentrations of ceftazidime and ceftriaxone, two expanded-spectrum cephalosporins commonly used in the treatment of bacterial meningitis, in two brain regions (the right corpus striatum and the left lateral ventricle_ of awake, freely moving rats. Antibiotics were administered by constant intravenous infusion at 18 mg/h until steady-state levels were reached. Ceftriaxone levels measured at the steady state in the extracellular space of the corpus striatum (0.80 +/- 0.17 micrograms/ml) were statistically equivalent to those obtained in the cerebrospinal fluid of the lateral ventricle (0.71 +/- 0.15 micrograms/ml). The ratios of these levels in the brain to the steady-state levels in plasma were 0.5 +/- 0.1% for both regions. The postinfusion concentrations of ceftriaxone in the brain declined monoexponentially, with an elimination half-life similar to that obtained in plasma. However, the mean antibiotic concentration of ceftazidime in the striatum (2.2 +/- 0.4 micrograms/ml) was lower (P < 0.001) than that in the lateral ventricle (3.8 +/- 0.5% and 4.0 +/- 1.8%, respectively) were higher than those obtained with ceftriaxone. Moreover, the half-life of ceftazidime elimination from plasma was lower than that obtained in the two brain regions. It was concluded that the in vivo microdialysis technique yields useful data on antibiotic distribution in the extracellular space of the brain, that the distribution may not be homogeneous, and that the decay of postinfusion concentrations in the brain may be different from the decay of postinfusion concentrations in plasma.     

A temporary local energy pool coupled to neuronal activity: fluctuations of extracellular lactate levels in rat brain monitored with rapid-response enzyme-based sensor

A successfully developed enzyme-based lactate microsensor with rapid response time allows the direct and continuous in vivo measurement of lactic acid concentration with high temporal resolution in brain extracellular fluid. The fluctuations coupled to neuronal activity in extracellular lactate concentration were explored in the dentate gyrus of the hippocampus of the rat brain after electrical stimulation of the perforant pathway. Extracellular glucose and oxygen levels were also detected simultaneously by coimplantation of a fast-response glucose sensor and an oxygen electrode, to provide novel information of trafficking of energy substances in real time related to local neuronal activity. The results first give a comprehensive picture of complementary energy supply and use of lactate and glucose in the intact brain tissue. In response to acute neuronal activation, the brain tissue shifts immediately to significant energy supply by lactate. A local temporary fuel "reservoir" is established behind the blood-brain barrier, evidenced by increased extracellular lactate concentration. The pool can be depleted rapidly, up to 28% in 10-12 s, by massive, acute neuronal use after stimulation and can be replenished in approximately 20 s. Glutamate-stimulated astrocytic glycolysis and the increase of regional blood flow may regulate the lactate concentration of the pool in different time scales to maintain local energy homeostasis.     

Glucose-induced intracellular ion changes in sugar-sensitive hypothalamic neurons

In the lateral hypothalamic area (LHA) of rat brain, approximately 30% of cells showed sensitivity to small changes in local concentrations of glucose. These "glucose-sensitive" neurons demonstrated four types of behavior, three of which probably represent segments of a continuous spectrum of recruitment in response to ever more severe changes in blood sugar. Type I cells showed maximum activity 相似文献   

New insights into the compartmentation of glutamate and glutamine in cultured rat brain astrocytes

Studies from several groups have provided evidence that glutamate and glutamine are metabolized in different compartments in astrocytes. In the present study we measured the rates of 14CO2 production from U-[14C]glutamate and U-[14C]glutamine, and utilized both substrate competition experiments and the transaminase inhibitor aminooxyacetic acid (AOAA) to obtain more information about the compartmentation of these substrates in cultured rat brain astrocytes. The rates of oxidation of 1 mM glutamine and glutamate were 26.4 +/- 1.4 and 63.0 +/- 7.4 nmol/h/mg protein, respectively. The addition of 1 mM glutamate decreased the rate of oxidation of glutamine to 26.3% of the control rate, demonstrating that glutamate can effectively compete with the oxidation of glutamine by astrocytes. In contrast, the addition of 1 mM glutamine had little or no effect on the rate of oxidation of glutamate by astrocytes, demonstrating that the glutamate produced intracellularly from exogenous glutamine does not dilute the glutamate taken up from the media. The addition of 5 mM AOAA decreased the rate of 14CO2 production from glutamine to 29.2% of the control rate, consistent with earlier studies by our group. The addition of 5 mM AOAA decreased the rate of oxidation of concentrations of glutamate < or = 0.1 mM by approximately 50%, but decreased the oxidation of 0.5-1 mM glutamate by only approximately 20%, demonstrating that a substantial portion of glutamate enters the tricarboxylic acid (TCA) cycle via glutamate dehydrogenase (GDH) rather than transamination, and that as the concentration of glutamate increases the relative proportion entering the TCA cycle via GDH also increases. To determine if the presence of an amino group acceptor (i.e. a ketoacid) would increase the rate of metabolism of glutamate, pyruvate was added in some experiments. Addition of 1 mM pyruvate increased the rate of oxidation of glutamate, and the increase was inhibited by AOAA, consistent with enhanced entry of glutamate into the TCA cycle via transamination in the presence of pyruvate. Enzymatic studies showed that pyruvate increased the activity of mitochondrial aspartate aminotransferase (AAT). Overall, the data demonstrate that glutamate formed intracellularly from glutamine enters the TCA cycle primarily via transamination, but does not enter the same TCA cycle compartment as glutamate taken up from the extracellular milieu. In contrast, extracellular glutamate enters the TCA cycle in astrocytes via both transamination and GDH, and can compete with, or dilute, the oxidation of glutamate produced intracellularly from glutamine.     

Monosodium glutamate (MSG)-obese rats develop glucose intolerance and insulin resistance to peripheral glucose uptake

Different levels of insulin sensitivity have been described in several animal models of obesity as well as in humans. Monosodium glutamate (MSG)-obese mice were considered not to be insulin resistant from data obtained in oral glucose tolerance tests. To reevaluate insulin resistance by the intravenous glucose tolerance test (IVGTT) and by the clamp technique, newborn male Wistar rats (N = 20) were injected 5 times, every other day, with 4 g/kg MSG (N = 10) or saline (control; N = 10) during the first 10 days of age. At 3 months, the IVGTT was performed by injecting glucose (0.75 g/kg) through the jugular vein into freely moving rats. During euglycemic clamping plasma insulin levels were increased by infusing 3 mU.kg-1.min-1 of regular insulin until a steady-state plateau was achieved. The basal blood glucose concentration did not differ between the two experimental groups. After the glucose load, increased values of glycemia (P < 0.001) in MSG-obese rats occurred at minute 4 and from minute 16 to minute 32. These results indicate impaired glucose tolerance. Basal plasma insulin levels were 39.9 +/- 4 microU/ml in control and 66.4 +/- 5.3 microU/ml in MSG-obese rats. The mean post-glucose area increase of insulin was 111% higher in MSG-obese than in control rats. When insulinemia was clamped at 102 or 133 microU/ml in control and MSG rats, respectively, the corresponding glucose infusion rate necessary to maintain euglycemia was 17.3 +/- 0.8 mg.kg-1.min-1 for control rats while 2.1 +/- 0.3 mg.kg-1.min-1 was sufficient for MSG-obese rats. The 2-h integrated area for total glucose metabolized, in mg.min.dl-1, was 13.7 +/- 2.3 vs 3.3 +/- 0.5 for control and MSG rats, respectively. These data demonstrate that MSG-obese rats develop insulin resistance to peripheral glucose uptake.     

Pharmacokinetics and distribution over the blood-brain barrier of zalcitabine (2',3'-dideoxycytidine) and BEA005 (2', 3'-dideoxy-3'-hydroxymethylcytidine) in rats, studied by microdialysis

Microdialysis was applied to sample the unbound drug concentration in the extracellular fluid in brain and muscle of rats given zalcitabine (2',3'-dideoxycytidine; n = 4) or BEA005 (2', 3'-dideoxy-3'-hydroxymethylcytidine; n = 4) (50 mg/kg of body weight given subcutaneously). Zalcitabine and BEA005 were analyzed by high-pressure liquid chromatography with UV detection. The maximum concentration of zalcitabine in the dialysate (Cmax) was 31.4 +/- 5. 1 microM (mean +/- standard error of the mean) for the brain and 238. 3 +/- 48.1 microM for muscle. The time to Cmax was found to be from 30 to 45 min for the brain and from 15 to 30 min for muscle. Zalcitabine was eliminated from the brain and muscle with half-lives 1.28 +/- 0.64 and 0.85 +/- 0.13 h, respectively. The ratio of the area under the concentration-time curve (AUC) (from 0 to 180 min) for the brain and the AUC for muscle (AUC ratio) was 0.191 +/- 0.037. The concentrations of BEA005 attained in the brain and muscle were lower than those of zalcitabine, with Cmaxs of 5.7 +/- 1.4 microM in the brain and 61.3 +/- 12.0 microM in the muscle. The peak concentration in the brain was attained 50 to 70 min after injection, and that in muscle was achieved 30 to 50 min after injection. The half-lives of BEA005 in the brain and muscle were 5.51 +/- 1.45 and 0.64 +/- 0.06 h, respectively. The AUC ratio (from 0 to 180 min) between brain and muscle was 0.162 +/- 0.026. The log octanol/water partition coefficients were found to be -1.19 +/- 0.04 and -1.47 +/- 0.01 for zalcitabine and BEA005, respectively. The degrees of plasma protein binding of zalcitabine (11% +/- 4%) and BEA005 (18% +/- 2%) were measured by microdialysis in vitro. The differences between zalcitabine and BEA005 with respect to the AUC ratio (P = 0.481), half-life in muscle (P = 0.279), and level of protein binding (P = 0.174) were not statistically significant. The differences were statistically significant in the case of the half-life in the brain (P = 0.032), clearance (P = 0.046), volume of distribution (P = 0.027) in muscle, and octanol/water partition coefficient (P = 0.019).     

Locally infused taurine, GABA and homotaurine alter differently the striatal extracellular concentrations of dopamine and its metabolites in rats

We studied in vivo the effects of locally infused taurine (50, 150, and 450 mM) on the striatal dopamine and its metabolites in comparison with those of GABA and homotaurine, a GABAA receptor agonist, in freely moving rats. The extracellular dopamine concentration was elevated maximally 2.5-, 2- and 4-fold by taurine, GABA and homotaurine, respectively. At 150 mM concentration, at which the maximum effects occurred, homotaurine increased the extracellular dopamine more than taurine or GABA. When taurine and GABA were infused simultaneously with tetrodotoxin the output of dopamine did not differ from that in the presence of tetrodotoxin alone. In comparison, tetrodotoxin did not inhibit the increase in extracellular dopamine caused by homotaurine. Furthermore, omission of calcium from the perfusion fluid inhibited the increase of extracellular dopamine caused by GABA. However, it did not block the increase of dopamine caused by taurine or homotaurine. The present study suggests that the effects of intrastriatal taurine, GABA and homotaurine on the striatal extracellular dopamine differ. Thus, these amino acids seem to affect the striatal dopaminergic neurons via more than one mechanism.     

The influence of the milieu on the rate of neutralisation of herpes simplex virus 1

Minimizing secondary injury after severe traumatic brain injury (TBI) is the primary goal of cerebral resuscitation. For more than two decades, hyperventilation has been one of the most often used strategies in the management of TBI. Laboratory and clinical studies, however, have verified a post-TBI state of reduced cerebral perfusion that may increase the brain's vulnerability to secondary injury. In addition, it has been suggested in a clinical study that hyperventilation may worsen outcome after TBI. OBJECT: Using the controlled cortical impact model in rats, the authors tested the hypothesis that aggressive hyperventilation applied immediately after TBI would worsen functional outcome, expand the contusion, and promote neuronal death in selectively vulnerable hippocampal neurons. METHODS: Twenty-six intubated, mechanically ventilated, isoflurane-anesthetized male Sprague-Dawley rats were subjected to controlled cortical impact (4 m/second, 2.5-mm depth of deformation) and randomized after 10 minutes to either hyperventilation (PaCO2 = 20.3 +/- 0.7 mm Hg) or normal ventilation groups (PaCO2 = 34.9 +/- 0.3 mm Hg) containing 13 rats apiece and were treated for 5 hours. Beam balance and Morris water maze (MWM) performance latencies were measured in eight rats from each group on Days 1 to 5 and 7 to 11, respectively, after controlled cortical impact. The rats were killed at 14 days postinjury, and serial coronal sections of their brains were studied for contusion volume and hippocampal neuron counting (CA1, CA3) by an observer who was blinded to their treatment group. Mortality rates were similar in both groups (two of 13 in the normal ventilation compared with three of 13 in the hyperventilation group, not significant [NS]). There were no differences between the groups in mean arterial blood pressure, brain temperature, and serum glucose concentration. There were no differences between groups in performance latencies for both beam balance and MWM or contusion volume (27.8 +/- 5.1 mm3 compared with 27.8 +/- 3.3 mm3, NS) in the normal ventilation compared with the hyperventilation groups, respectively. In brain sections cut from the center of the contusion, hippocampal neuronal survival in the CA1 region was similar in both groups; however, hyperventilation reduced the number of surviving hippocampal CA3 neurons (29.7 cells/hpf, range 24.2-31.7 in the normal ventilation group compared with 19.9 cells/hpf, range 17-23.7 in the hyperventilation group [25th-75th percentiles]; *p < 0.05, Mann-Whitney rank-sum test). CONCLUSIONS: Aggressive hyperventilation early after TBI augments CA3 hippocampal neuronal death; however, it did not impair functional outcome or expand the contusion. These data indicate that CA3 hippocampal neurons are selectively vulnerable to the effects of hyperventilation after TBI. Further studies delineating the mechanisms underlying these effects are needed, because the injudicious application of hyperventilation early after TBI may contribute to secondary neuronal injury.     

0.45% saline and 5% dextrose in water, but not 0.9% saline or 5% dextrose in 0.9% saline, worsen brain edema two hours after closed head trauma in rats

In this study, we examined the effect of four i.v. fluids (250 mL/kg) on blood glucose and osmolality and brain tissue specific gravity after closed head trauma (CHT) in rats. CHT was delivered at Time 0; blood was sampled at 60 min; fluid infusion began at 75 min and ended at 105 min. Blood was again sampled at 105 and 120 min, and brain tissue specific gravity was determined at 120 min. Five groups (one control and four fluid-treated groups) received CHT, and five other groups (one control and four fluid-treated) did not (n = 9 in each group). 0.45% saline (1/2 NS) and 5% dextrose in water (D5W) accentuated the decrease of brain tissue specific gravity (1.0366 +/- 0.0025 and 1.0368 +/- 0.0028, respectively; mean +/- SD) caused by CHT (1.0395 +/- 0.0036), but 5% dextrose in 0.9% saline (D5NS) and 0.9% saline (NS) did not (1.0431 +/- 0.0042 and 1.0389 +/- 0.0049, respectively). In addition, 1/2 NS decreased blood osmolality (248 +/- 6 mOsm/L), D5W increased blood glucose (1095 +/- 173 mg/dL), D5NS increased blood osmolality (350 +/- 5 mOsm/L) and glucose (1695 +/- 76 mg/dL), and NS caused no significant change. We conclude that administering hypoosmolar i.v. fluids after CHT causes a significant worsening of cerebral edema 2 h after CHT. Implications: We previously reported worse neurological outcome and/or mortality after closed head trauma in rats when 5% dextrose in water or 0.45% saline was given i.v. compared with 0.9% saline or 5% dextrose in 0.9% saline. The present results and our previous findings indicate that worsening of outcome after closed head trauma in rats may be caused more by edema formation than by hyperglycemia.     

NMDA receptor-mediated pilocarpine-induced seizures: characterization in freely moving rats by microdialysis

1. Pilocarpine administration has been used as an animal model for temporal lobe epilepsy since it produces several morphological and synaptic features in common with human complex partial seizures. Little is known about changes in extracellular neurotransmitter concentrations during the seizures provoked by pilocarpine, a non-selective muscarinic agonist. 2. Focally evoked pilocarpine-induced seizures in freely moving rats were provoked by intrahippocampal pilocarpine (10 mM for 40 min at a flow rate of 2 microl min(-1)) administration via a microdialysis probe. Concomitant changes in extracellular hippocampal glutamate, gamma-aminobutyric acid (GABA) and dopamine levels were monitored and simultaneous electrocorticography was performed. The animal model was characterized by intrahippocampal perfusion with the muscarinic receptor antagonist atropine (20 mM), the sodium channel blocker tetrodotoxin (1 microM) and the N-methyl-D-aspartate (NMDA) receptor antagonist MK-801 (dizocilpine maleate, 100 microM). The effectiveness of locally (600 microM) or systemically (10 mg kg(-1) day(-1)) applied lamotrigine against the pilocarpine-induced convulsions was evaluated. 3. Pilocarpine initially decreased extracellular hippocampal glutamate and GABA levels. During the subsequent pilocarpine-induced limbic convulsions extracellular glutamate, GABA and dopamine concentrations in hippocampus were significantly increased. Atropine blocked all changes in extracellular transmitter levels during and after co-administration of pilocarpine. All pilocarpine-induced increases were completely prevented by simultaneous tetrodotoxin perfusion. Intrahippocampal administration of MK-801 and lamotrigine resulted in an elevation of hippocampal dopamine levels and protected the rats from the pilocarpine-induced seizures. Pilocarpine-induced convulsions developed in the rats which received lamotrigine perorally. 4. Pilocarpine-induced seizures are initiated via muscarinic receptors and further mediated via NMDA receptors. Sustained increases in extracellular glutamate levels after pilocarpine perfusion are related to the limbic seizures. These are arguments in favour of earlier described NMDA receptor-mediated excitotoxicity. Hippocampal dopamine release may be functionally important in epileptogenesis and may participate in the anticonvulsant effects of MK-801 and lamotrigine. The pilocarpine-stimulated hippocampal GABA, glutamate and dopamine levels reflect neuronal vesicular release.     

A role for astrocytes in glucose delivery to neurons?

The present paper examines the possible role of astrocytes in the delivery of glycogen-derived glucose for neuronal metabolism. Such a process would require astrocytic expression of glucose-6-phosphatase. The degree and significance of brain expression of glucose-6-phosphatase (EC 3.1.3.9) has been a subject of controversy. Published immunohistochemical data are consistent with expression of glucose-6-phosphatase by astrocytes, both in vivo and in vitro. In this paper additional confirmation of the expression of glucose-6-phosphatase mRNA in rat brain is presented. Although cultured astrocytes demonstrate glucose-6-phosphatase activity in vitro under assay conditions, there is very limited in vitro evidence that this activity confers a glucose-export capacity on astrocytes. Under most conditions in vitro, lactate export predominates, however this may relate to aspects of the in vitro phenotype. Data relating to astrocytic glucose and lactate export are considered in the context of hypotheses of trafficking by astrocytes of substrates for neuronal metabolism, hypotheses that imply and require compartmentation of these substances, in contrast with current formulations of glucose transport into and within brain that imply no glucose compartmentation. Microdialysis studies of the properties of the brain extracellular fluid (ECF) glucose pool in the freely moving rat were performed seeking evidence of glucose compartmentation. Results of these studies do imply compartmentalisation of brain glucose, and are consistent with a model envisaging the majority of glucose reaching the neuron via the astrocytic intracellular space and the ECF. In addition, such studies provide evidence that rises in ECF glucose concentration are not the direct result of local recruitment of cerebral blood flow, but suggest the influence of intermediate, astrocyte-based mechanisms. Astrocytic glucose-6-phosphatase may permit astrocytes to modulate the trans-astrocytic flux of glucose to adjacent neurons in response to signals reflecting increased neuronal demand.     

Accuracy of the one-point in vivo calibration of "wired" glucose oxidase electrodes implanted in jugular veins of rats in periods of rapid rise and decline of the glucose concentration

The hypothesis of the feasibility of one-point in vivo calibration of intravenously implanted glucose sensors during periods of rapid rise and decline of venous blood glucose concentration was tested. Miniature (5 x 10(-4) cm2 mass transporting area) glucose electrodes with 10-90% response times < 2 min, that did not consume oxygen, were implanted in jugular veins of systemically heparinized rats and used in 4-h experiments, during which the blood glucose concentration was amperometrically monitored. The glucose electrodes were made by electrically connecting ("wiring") reaction centers of glucose oxidase through an electron-conducting redox hydrogel to gold electrode surfaces. The redox polymer and enzyme constituting the electrode sensing layer were immobilized by cross-linking, and thus the electrodes had no diffusional and readily leached redox mediator. One hour after their implantation, the electrodes accurately tracked the blood glucose concentration when calibrated in vivo by a one-point calibration, when the glucose concentration was steady, when rising rapidly, and when declining steeply. For an assumed 2-min lag time, the sensor readings were well correlated with the true blood glucose concentrations, with linear regression analysis yielding a slope of 0.97 +/- 0.07 and an intercept (bias) of 0.3 +/- 0.3 mM. The correlation coefficient, r2, was 0.949 +/- 0.020, and the percent difference through the 2-22 mM range was 1.9 +/- 1.0%. The results suggest that, in combination with understanding and modeling of transient physiological differences between the subcutaneous and the blood glucose concentrations, it will be possible to calibrate by one-point in vivo calibration subcutaneously implanted sensors, even while the glucose concentration changes rapidly.