Cerebral Blood Flow and Metabolism During Hypoglycemia in Newborn Dogs

: Cerebral blood flow (CBF) and cerebral metabolic rates (CMR) were studied in newborn dogs during insulin-induced hypoglycemia. Pups were anesthetized, paralyzed, and artificially ventilated with a mixture of 70% nitrous oxide and 30% oxygen to maintain normoxia and normocarbia. Experimental animals were given regular insulin (0.3 units/gm IV); controls received normal saline. CBF was determined using a modification of the Kety-Schmidt technique employing ¹³³Xe as indicator. Arteriovenous differences for oxygen, glucose, lactate, and β-hydroxybutyrate (β-OHB) were also measured, and CMRo₂ and CMR_substrates calculated. Two groups of hypoglycemic dogs were identified; those in which blood glucose levels were greater than 0.5 mm (group 1), and those in which they were less than 0.5 mm (group 2). CBF did not change significantly from control values of 23 ± 10 ml/min/100 g (mean ±s.d. ) at both levels of hypoglycemia. Similarly, hypoglycemia did not alter CMRo₂, significantly from its initial level of 1.05 ± 0.37 ml O₂/min/100 g. Glucose consumption in brain during normoglycemia accounted for 95% of cerebral energy supply with minimal contributions from lactate (4%) and β-OHB (0.5%). During hypoglycemia, CMR_glucose. declined by 29 and 52% in groups 1 and 2, respectively, while CMR,_lactate increased to the extent that this metabolite became the dominant fuel for oxidative metabolism in brain. The cerebral utilization of β-OHB was unaltered by hypoglycemia. The findings indicate that insulin-induced hypoglycemia in the newborn dog is associated with an increase in cerebral lactate utilization, supplementing glucose as the primary energy fuel and thereby preserving a normal CMRo₂. These metabolic responses may contribute to the tolerance of the immature nervous system to the known deleterious effects of hypoglycemia.     

Endogenous Substrates Utilized by Rat Brain in Severe Insulin-Induced Hypoglycemia

Abstract: Several previous studies have demonstrated that severe hypoglycemia is accompanied by consumption of endogenous brain substrates (glycolytic and citric acid cycle metabolites and free amino acids) and some have shown a loss of structural components as well, notably phospholipids. In the present study, on paralysed and artificially ventilated rats, we measured cerebral oxygen and glucose consumption during 30 min of hypoglycemic coma (defined as hypoglycemia of sufficient severity to cause cessation of spontaneous EEG activity) and calculated the non-glucose oxygen consumption. In an attempt to estimate the missing substrate we measured tissue concentrations of phospholipids and RNA. After 5 min of hypoglycemic coma, tissue phospholipid content decreased by about 8% with no further change during the subsequent 55 min. A similar reduction remained after 90 min of recovery, induced by glucose administration following 30 min of coma. Since no preferential loss of polyenoic fatty acids or of ethanolamine phosphoglycerides occurred, it is concluded that loss of phospholipids was due to phospholipase activity rather than to peroxidative degradation. The free fatty acid concentration increased sixfold after 5 min of coma and remained elevated during the course of hypoglycemia. A 9% reduction in tissue RNA content was observed after 30 min of hypoglycemia. Calculations indicated that available endogenous carbohydrate and amino acid substrates were essentially consumed after 5 min of coma, and that other non-glucose substrates must have accounted for approximately 50μmol·g^?1 of oxygen (8.3 μmol·g^?1 in terms of glucose equivalents) within the 5–30 min period. The 10% reduction in phospholipid-bound fatty acids was more than sufficient (in four- to fivefold excess) to account for this oxygen consumption. However, since no further degradation occurred in the 5–30 min period, there is no simple, direct, quantitative relationship between oxygen consumption and cortical fatty acid oxidation during this interval. The possibility thus remains that unmeasured exogenous or endogenous substrates were utilized.     

Determination of Glucose Utilization Rates in Cultured Astrocytes and Neurons with [<Superscript>14</Superscript>C]deoxyglucose: Progress,Pitfalls, and Discovery of Intracellular Glucose Compartmentation

2-Deoxy-d-[¹⁴C]glucose ([¹⁴C]DG) is commonly used to determine local glucose utilization rates (CMR_glc) in living brain and to estimate CMR_glc in cultured brain cells as rates of [¹⁴C]DG phosphorylation. Phosphorylation rates of [¹⁴C]DG and its metabolizable fluorescent analog, 2-(N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino)-2-deoxyglucose (2-NBDG), however, do not take into account differences in the kinetics of transport and metabolism of [¹⁴C]DG or 2-NBDG and glucose in neuronal and astrocytic cells in cultures or in single cells in brain tissue, and conclusions drawn from these data may, therefore, not be correct. As a first step toward the goal of quantitative determination of CMR_glc in astrocytes and neurons in cultures, the steady-state intracellular-to-extracellular concentration ratios (distribution spaces) for glucose and [¹⁴C]DG were determined in cultured striatal neurons and astrocytes as functions of extracellular glucose concentration. Unexpectedly, the glucose distribution spaces rose during extreme hypoglycemia, exceeding 1.0 in astrocytes, whereas the [¹⁴C]DG distribution space fell at the lowest glucose levels. Calculated CMR_glc was greatly overestimated in hypoglycemic and normoglycemic cells because the intracellular glucose concentrations were too high. Determination of the distribution space for [¹⁴C]glucose revealed compartmentation of intracellular glucose in astrocytes, and probably, also in neurons. A smaller metabolic pool is readily accessible to hexokinase and communicates with extracellular glucose, whereas the larger pool is sequestered from hexokinase activity. A new experimental approach using double-labeled assays with DG and glucose is suggested to avoid the limitations imposed by glucose compartmentation on metabolic assays.     

CEREBRAL METABOLIC AND VASCULAR EFFECTS OF BARBITURATE THERAPY FOLLOWING COMPLETE GLOBAL ISCHEMIA

Complete global cerebral ischemia was induced in dogs by temporary ligation of the ascending aorta for 10min. Prior to the ischemic period, half of the animals were given pentobarbital 30-38 mg/kg, a maneuver previously reported to prevent or attenuate cerebral damage in this same model. Cerebral blood flow (CBF) and cerebral metabolic rate (CMR_O2) were followed from prior to the ischemic period to 6 h post-ischemia. At varying time intervals following ischemia, brain biopsies were obtained and analyzed for cerebral metabolites to determine the cerebral energy state. Only a few differences were observed between pentobarbital-treated and untreated animals. Post-ischemic CMR_O2, stabilized at a significantly lower level in treated than in untreated animals. However, CBF was proportionately lower and thus O₂ delivery relative to O₂ needs in the two groups was comparable. Also in both groups, the CBF and CMR_O2 stabilized at levels significantly below pre-ischemia controls. Cerebral energy stores in both groups were depleted after 10min of ischemia but were restored to near normal within 4min post-ischemia. Total restoration of the adenine nucleotide pool and ATP were delayed as was the return of brain lactate to normal. A 10min period of post-ischemic hyperemia was observed in all animals and in the initial 4min post-ischemia CMR_O2 was also increased. The latter is probably accounted for by the O₂ needs for restoration of cerebral energy and O₂ stores. We conclude that cerebral protection as provided by barbiturates following complete global ischemia cannot be accounted for by any measurable effect on CBF, CMR_O2, or the cerebral energy stores during the initial 6 h post-ischemia.     

Decreased vascular endothelial growth factor response to acute hypoglycemia in type 2 diabetic patients with hypoglycemic coma

IntroductionPlasma vascular endothelial growth factor (VEGF) was shown to increase during acute hypoglycemia and could mediate rapid adaptation of the brain. In this study we examined the neuroendocrine response in patients with type 2 diabetes mellitus (T2DM) in hypoglycemic coma or with acute neuroglycopenic symptoms.MethodsWe prospectively studied 135 consecutive T2DM patients admitted for severe hypoglycemia during a 2-year period. We collected clinical variables and measured plasma concentrations of VEGF, epinephrine, norepinephrine, cortisol and growth hormone at admission and 30 min afterwards.ResultsThirty two patients developed hypoglycemic coma and 103 did not lose consciousness. Median plasma VEGF level of coma patients was 3.1-fold lower at baseline than that of non-coma patients, and even 5.3-fold lower 30 min afterwards. Plasma epinephrine concentration was significantly lower just at baseline in coma patients. On the contrary, there were no differences in concentrations of the other hormones. Multivariate logistic regression analysis showed that VEGF concentration (OR 0.68; CI 0.51–0.95) was a protective factor against the development of coma.ConclusionsVEGF and epinephrine responses to acute hypoglycemia are reduced in T2DM patients who develop hypoglycemic coma. An increased plasma VEGF concentration appeared to be a protective factor against the development of hypoglycemic coma.     

Regional Acetylcholine Metabolism in Brain During Acute Hypoglycemia and Recovery

Insulin-induced hypoglycemia in normothermic rats caused progressive neurological depression and differentially altered regional cerebral acetylcholine metabolism. Reductions of plasma glucose from 7.7 mM (control) to 2.5-1.7 mM (moderate hypoglycemia associated with decreased motor activity) or 1.5 mM (severe hypoglycemia with lethargy progressing to stupor) decreased glucose concentrations in the cerebral cortex, striatum, and hippocampus to less than 10% of control. Moderate hypoglycemia diminished acetylcholine concentrations in cortex and striatum (21% and 45%, respectively) and reduced [1-2H2, 2-2H2]choline incorporation into acetylcholine (62% and 41%, respectively). Severe hypoglycemia did not reduce the acetylcholine concentration or synthesis in cortex and striatum further. The concentrations of choline rose in the cortex (+53%) and striatum (+130%) of animals that became stuporous but a similar rise in [1-2H2, 2-2H2]choline left the specific activities of choline in these structures unchanged. Even severe hypoglycemia did not alter the hippocampal cholinergic system. In rats that developed hypoglycemic stupor and were then treated with glucose, the animals recovered apparently normal behavior, and the concentrations of acetylcholine and the incorporation of [1-2H2, 2-2H2]-choline into acetylcholine returned to control values in the striatum but not in the cerebral cortex. Thus, impaired acetylcholine metabolism in selected regions of the brain may contribute to the early symptoms of neurological dysfunction in hypoglycemia.     

Brain oxygen utilization is unchanged by hypoglycemia in normal humans: lactate, alanine, and leucine uptake are not sufficient to offset energy deficit

During hypoglycemia, substrates other than glucose have been suggested to serve as alternate neural fuels. We evaluated brain uptake of endogenously produced lactate, alanine, and leucine at euglycemia and during insulin-induced hypoglycemia in 17 normal subjects. Cross-brain arteriovenous differences for plasma glucose, lactate, alanine, leucine, and oxygen content were quantitated. Cerebral blood flow (CBF) was measured by Fick methodology using N(2)O as the dilution indicator gas. Substrate uptake was measured as the product of CBF and the arteriovenous concentration difference. As arterial glucose concentration fell, cerebral oxygen utilization and CBF remained unchanged. Brain glucose uptake (BGU) decreased from 36.3+/-2.6 to 26.6+/-2.1 micromol.100 g of brain(-1).min(-1) (P<0.001), equivalent to a drop in ATP of 291 micromol.100 g(-1).min(-1). Arterial lactate rose (P<0.001), whereas arterial alanine and leucine fell (P<0.009 and P<0.001, respectively). Brain lactate uptake (BLU) increased from a net release of -1.8+/- 0.6 to a net uptake of 2.5+/-1.2 micromol.100 g(-1).min(-1) (P<0.001), equivalent to an increase in ATP of 74 micromol.100 g(-1).min(-1). Brain leucine uptake decreased from 7.1+/-1.2 to 2.5 +/- 0.5 micromol.100 g(-1).min(-1) (P<0.001), and brain alanine uptake trended downward (P<0.08). We conclude that the ATP generated from the physiological increase in BLU during hypoglycemia accounts for no more than 25% of the brain glucose energy deficit.     

Fructose 2, 6-Bisphosphate in Hypoglycemic Rat Brain

Abstract: Fructose 2,6-bisphosphate has been studied during hypoglycemia induced by insulin administration (40 IU/kg). No changes in content of cerebral fructose 2,6-bisphosphate were found in mild hypoglycemia, but the level of this compound was markedly decreased in hypoglycemic coma and recovered after 30 min of glucose administration. To correlate a possible modification of the concentration of the metabolite with selective regional damage occurring during hypoglycemic coma, we have analyzed four cerebral areas (cortex, striatum, cerebellum, and hippocampus). Fructose 2,6-bisphosphate concentrations were similar in the four areas analyzed; severe hypoglycemia decreased levels of the metabolite to the same extent in all the brain areas studied. The decrease in content of fructose 2,6-bisphosphate was not always accompanied by a parallel decrease in ATP levels, a result suggesting that the low levels of the bisphosphorylated metabolite during hypoglycemic coma could be due to the decreased 6-phosphofructo-2-kinase activity, mainly as a consequence of the fall in concentration of its substrate (fructose 6-phosphate). These results suggest that fructose 2,6-bisphosphate could play a permissive role in cerebral tissue, maintaining activation of 6-phosphofructo-l-kinase and glycolysis.     

LEVELS OF CEREBRAL CORTICAL GLYCOLYTIC AND CITRIC ACID CYCLE METABOLITES DURING HYPOGLYCEMIC STUPOR AND ITS REVERSAL

Abstract— Blood glucose, cerebral cortical glucose, and eight metabolites of the glycolytic pathway and citric acid cycle were measured during insulin hypoglycemic stupor and during the first 100s after glucose administration. In hypoglycemic mice that had lost righting ability, blood and brain glucose were decreased 89% and 96% respectively, but glucose-6-phosphate fell only 23%. Other glycolytic and citric acid cycle intermediates were decreased 31–77%. Fructose bisphosphate, 3-phosphoglycerate and phosphopyruvate fell more than glucose-6-phosphate, but less than pyruvate and lactate. Citrate fell less than a-ketoglutarate and malate. These results suggest that in severe hypoglycemia there is a decrease in brain glucose utilization, mediated by phosphofructokinase, but probably caused by decreased neuronal activity. An intravenous injection of glucose restored brain glucose to 75% of normal within 10s and caused return of righting ability within 60s. Glucose-6-phosphate, fructose bisphosphate, 3-phosphoglycerate, and phosphopyruvate rose to normal or near normal levels within 60s, whereas pyruvate, lactate, citrate, ã-ketoglutarate, and malate changed little in this period. This suggests that although glucose given to hypoglycemic animals rapidly enters the glycolytic pathway in brain (and behavior is almost normal), total neuronal activity, and hence overall glucose metabolism, remains subnormal for several minutes.     

Effects of Hypoglycemia on Rat Brain Polyribosome Sedimentation Pattern

Insulin-induced hypoglycemia provokes polyribosome disaggregation and accumulation of monomeric ribosomes in the brain of rats with hypoglycemic paresis and coma. The extent of brain polyribosome disaggregation depends on the decrease of blood glucose concentration, and in comatose animals on the duration of hypoglycemia. Cycloheximide prevents the disaggregation of brain polyribosomes induced by hypoglycemia, indicating that hypoglycemia affects brain protein synthesis, decreasing the rate of initiation relative to the rate of elongation of polypeptide chain synthesis.     

Regional levels of glucose,amino acids,high energy phosphates,and cyclic nucleotides in the central nervous system during hypoglycemic stupor and behavioral recovery

The effects of insulin-induced hypoglycemic stupor and subsequent treatment with glucose on mouse cerebral cortical, cerebellar and brain stem levels of glucose, glycogen, ATP, phosphocreatine, glutamate, aspartate and GABA and on cerebral cortical and cerebellar levels of cyclic AMP and cyclic GMP have been measured. Hypoglycemia decreased glucose, glycogen and glutamate levels and had no effect on ATP levels in all three regions of brain. GABA levels were decreased only in cerebellum. Aspartate levels rose in cerebral cortex and brain stem, and creatine phosphate increased in cerebral cortex and cerebellum. In the hypoglycemic stuporous animals, cyclic GMP levels were elevated in cerebral cortex and depressed in cerebellum whereas cyclic AMP levels were unchanged from control values. Intravenous administration of 2.5-3.5 mmol/kg of glucose to the hypoglycemic stuporous animals produced recovery of near normal neurological function within 45 s. Only brain glucose and aspartate levels returned to normal prior to behavioral recovery. These results suggest that of the several substances examined in this study, only glucose and perhaps aspartate have important roles in the biochemical mechanisms producing neurological abnormalities in hypoglycemic animals.     

Catecholamines in the rat brain during hypoglycemic convulsions and coma

The purpose of the study was to investigate the relation between the catecholamines: noradrenaline and dopamine in the rat brain on one hand and hypoglycemic convulsions and coma on the other. Concentrations of noradrenaline in the hypothalamus, brain stem and cerebral cortex were decreased during hypoglycemic convulsions and were lower during coma than those during convulsions. Dopamine concentration in the striatum was decreased during convulsions and coma. It was shown that the decrease in concentration of catecholamines was a result of hypoglycemia but not of insulin action itself. Clonidine- alpha 2 agonist accelerated occurrence and prolonged duration of hypoglycemic convulsions. Haloperidol-dopamine receptor blocker had no effect on the time of occurrence or duration of convulsions and coma. The results indicate that noradrenaline may exert an inhibitory influence on hypoglycemic convulsions. No evidence has been provided to support involvement of dopamine in the control of hypoglycemic convulsions and coma.     

CEREBRAL BLOOD FLOW AND CEREBRAL METABOLIC RATES FOR OXYGEN, GLUCOSE, AND KETONE BODIES IN NEWBORN DOGS

Cerebral blood flow (CBF) and the cerebral metabolic rates for oxygen, glucose, acetoacetate, β-hydroxybutyrate and lactate were measured in 1- to 5-day old Beagle dogs under nitrous oxide anesthesia. CBF was determined by ¹³³Xe washout with mechanically integrated blood samples withdrawn simultaneously from a femoral artery and from the posterior one-third of the superior sagittal sinus. CBF and CMRO₂ in normocapnia (PaCO₂ 40 × 1 mm Hg) were 48 × 5 ml/100 g/min and 2.15 ml/100 g/min, respectively. There was a positive, linear relationship between CBF and PaCO₂, calculated for PaCO₂ values ranging from 26 to 70 mm Hg. Induced hypocapnia (PaCO₂ 31 × 1 mm Hg) or hypercapnia (PaCO₂ 58 × 2 mm Hg) did not alter the CMRO₂. Glucose and acetoacetate were taken up by the brain at all PaCO₂ levels examined; however, the cerebral uptake of glucose always exceeded the combined uptake of ketone bodies by more than a factor of ten. The cerebral metabolic rate for glucose (94.6 × 3.6 μmol/100 g/min) more than accounted for overall cerebral oxygen consumption, and yielded an oxygen:glucose ratio (mol:mol) of 5.1. Thus, as in adult animals, PaCO₂ is an important regulator of cerebral blood flow in puppies, and glucose is the major substrate for oxidative energy production in the immature brain. The oxidation of ketone bodies by the newborn dog brain accounts for not more than 6% of the in vivo cerebral oxygen consumption.     

Regional CNS Levels of Acetylcholine and Choline During Hypoglycemic Stupor and Recovery

During insulin stupor in mice, acetylcholine levels in cerebral cortex, cerebellum. brainstem, striatum, and hippocampus were unchanged from control values despite brain glucose concentrations 3-10% of normal, whereas choline levels rose 2.4-3.6-fold in all five CNS regions. Brain acetylcholine and choline levels did not change during recovery following glucose injection. The data suggest that. in hypoglycemic stupor, (1) overall rates of acetylcholine synthesis and degradation remain balanced within each of the CNS regions studied: (2) the biochemical mechanism that elevates brain choline levels is unlikely to be related only to cholinergic synaptic processes: and (3) brain choline levels need not rise for stupor to occur.     

Cerebral Polyamine Metabolism in Reversible Hypoglycemia of Rat: Relationship to Energy Metabolites and Calcium

Thirty minutes of insulin-induced reversible hypoglycemic coma (defined in terms of cessation of EEG activity) was produced in anesthetized rats. At the end of the hypoglycemic coma or after recovery for 3, 24, or 72 h induced by glucose infusion, the animals were reanesthetized and their brains frozen in situ. Two control groups were used: untreated controls without prior manipulations, and insulin controls, which received injections of insulin followed by glucose infusion to maintain blood glucose within the physiological range. The brains of these latter animals were frozen 3, 24, or 72 h after glucose infusion. Tissue samples from the cortex, striatum, hippocampus, and thalamus were taken to measure ornithine decarboxylase (ODC) activity, and putrescine and spermidine levels, as well as phosphocreatine (PCr), ATP, glucose, and lactate content. In addition, 20-microns thick coronal sections taken from the striatum and dorsal hippocampus were used for histological evaluation of cell damage and also stained for calcium. Insulin in the absence of hypoglycemia produced a significant increase in ODC activity and putrescine level but had no effect on the profiles of energy metabolites or spermidine. During hypoglycemic coma, brain PCr, ATP, glucose, and lactate levels were sharply reduced, as expected. Energy metabolites normalized after 3 h of recovery. In the striatum, significant secondary decreases in PCr and ATP contents and rises in glucose and lactate levels were observed after 24 h of recovery. ODC activity, and putrescine and spermidine levels were unchanged during hypoglycemic coma. After 3 h of recovery, ODC activity increased markedly throughout the brain, except in the striatum. After 24 h of recovery, ODC activity decreased and approached control values 2 days later. Putrescine levels increased significantly throughout the brain after reversible hypoglycemic coma, the highest values observed after 24 h of recovery (p less than or equal to 0.001, compared with controls). After 72 h of recovery, putrescine levels decreased, but still significantly exceeded control values. Reversible hypoglycemic coma did not produce significant changes in regional spermidine levels except in the striatum, where an approximately 30% increase was observed after 3 and 72 h of recovery (p less than or equal to 0.01 and p less than or equal to 0.05, respectively). Twenty-four hours after hypoglycemic coma, intense calcium staining was apparent in layer III of the cerebral cortex, the lateral striatum, and the crest of the dentate gyrus. After 72 h of recovery, the intense calcium staining included also cortical layer II, the septal nuclei, the subiculum, and the hippocampal CA1-subfield.(ABSTRACT TRUNCATED AT 400 WORDS)     

Effects of Alda-1, an Aldehyde Dehydrogenase-2 Agonist,on Hypoglycemic Neuronal Death

Hypoglycemic encephalopathy (HE) is caused by a lack of glucose availability to neuronal cells, and no neuroprotective drugs have been developed as yet. Studies on the pathogenesis of HE and the development of new neuroprotective drugs have been conducted using animal models such as the hypoglycemic coma model and non-coma hypoglycemia model. However, both models have inherent problems, and establishment of animal models that mimic clinical situations is desirable. In this study, we first developed a short-term hypoglycemic coma model in which rats could be maintained in an isoelectric electroencephalogram (EEG) state for 2 min and subsequent hyperglycemia without requiring anti-seizure drugs and an artificial ventilation. This condition caused the production of 4-hydroxy-2-nonenal (4-HNE), a cytotoxic aldehyde, in neurons of the hippocampus and cerebral cortex, and a marked increase in neuronal death as evaluated by Fluoro-Jade B (FJB) staining. We also investigated whether N-(1,3-benzodioxole-5-ylmethyl)-2,6-dichlorobenzamide (Alda-1), a small-molecule agonist of aldehyde dehydrogenase-2, could attenuate 4-HNE levels and reduce hypoglycemic neuronal death. After confirming that EEG recordings remained isoelectric for 2 min, Alda-1 (8.5 mg/kg) or vehicle (dimethyl sulfoxide; DMSO) was administered intravenously with glucose to maintain a blood glucose level of 250 to 270 mg/dL. Fewer 4-HNE and FJB-positive cells were observed in the cerebral cortex of Alda-1-treated rats than in DMSO-treated rats 24 h after glucose administration (P = 0.002 and P = 0.020). Thus, activation of the ALDH2 pathway could be a molecular target for HE treatment, and Alda-1 is a potentially neuroprotective agent that exerts a beneficial effect on neurons when intravenously administered simultaneously with glucose.     

CEREBRAL ENERGY STATE IN INSULIN-INDUCED HYPOGLYCEMIA, RELATED TO BLOOD GLUCOSE AND TO EEG

—Concentrations of phosphocreatine, creatine, ATP, ADP and AMP were measured in the cerebral cortex of rats during insulin-induced hypoglycemia. Blood glucose concentrations were related to clinical symptoms in unanaesthetized animals and to the EEG pattern in paralysed and lightly anaesthetized animals. There was an excellent correlation between blood glucose concentration and EEG pattern. In animals showing a pronounced slowing of the EEG or convulsive polyspike activity for up to 20 min, there were no changes in any of the phosphates. However, after prolonged convulsive activity some animals showed clear signs of energy failure, and in all animals with an isoelectric EEG there was a major derangement of the energy state. Since the majority of those animals did not show signs of cerebral hypoxia or ischemia it is concluded that hypoglycemic coma is accompanied by substrate deficiency of a degree sufficient to induce energy depletion of brain tissue.     

Expression of Aquaporin 4 and Breakdown of the Blood-Brain Barrier after Hypoglycemia-Induced Brain Edema in Rats

Background

Hypoglycemia-induced brain edema is a severe clinical event that often results in death. The mechanisms by which hypoglycemia induces brain edema are unclear.

Methods

In a hypoglycemic injury model established in adult rats, brain edema was verified by measuring brain water content and visualizing water accumulation using hematoxylin and eosin staining. Temporal expression of aquaporin 4 (AQP4) and the integrity of the blood-brain barrier (BBB) were evaluated. We assessed the distribution and expression of AQP4 following glucose deprivation in astrocyte cultures.

Results

Brain edema was induced immediately after severe hypoglycemia but continued to progress even after recovery from hypoglycemia. Upregulation of AQP4 expression and moderate breakdown of the BBB were observed 24 h after recovery. In vitro, significant redistribution of AQP4 to the plasma membrane was induced following 6 h glucose deprivation.

Conclusion

Hypoglycemia-induced brain edema is caused by cytotoxic and vasogenic factors. Changes in AQP4 location and expression may play a protective role in edema resolution.     

Methylene Blue as a Cerebral Metabolic and Hemodynamic Enhancer

By restoring mitochondrial function, methylene blue (MB) is an effective neuroprotectant in many neurological disorders (e.g., Parkinson’s and Alzheimer’s diseases). MB has also been proposed as a brain metabolic enhancer because of its action on mitochondrial cytochrome c oxidase. We used in vitro and in vivo approaches to determine how MB affects brain metabolism and hemodynamics. For in vitro, we evaluated the effect of MB on brain mitochondrial function, oxygen consumption, and glucose uptake. For in vivo, we applied neuroimaging and intravenous measurements to determine MB’s effect on glucose uptake, cerebral blood flow (CBF), and cerebral metabolic rate of oxygen (CMRO₂) under normoxic and hypoxic conditions in rats. MB significantly increases mitochondrial complex I–III activity in isolated mitochondria and enhances oxygen consumption and glucose uptake in HT-22 cells. Using positron emission tomography and magnetic resonance imaging (MRI), we observed significant increases in brain glucose uptake, CBF, and CMRO₂ under both normoxic and hypoxic conditions. Further, MRI revealed that MB dramatically increased CBF in the hippocampus and in the cingulate, motor, and frontoparietal cortices, areas of the brain affected by Alzheimer’s and Parkinson’s diseases. Our results suggest that MB can enhance brain metabolism and hemodynamics, and multimetric neuroimaging systems offer a noninvasive, nondestructive way to evaluate treatment efficacy.     

Magnesium sulfate attenuates increased blood-brain barrier permeability during insulin-induced hypoglycemia in rats

Magnesium probably protects brain tissue against the effects of cerebral ischemia, brain injury and stroke through its actions as a calcium antagonist and inhibitor of excitatory amino acids. The effects of magnesium sulfate on cerebrovascular permeability to a dye, Evans blue, were studied during insulin-induced hypoglycemia with hypothermia in rats. Hypoglycemia was induced by an intramuscular injection of insulin. After giving insulin, each animal received MgSO4 (270 mg/kg) ip, followed by a 27 mg/kg dose every 20 min for 2.5 h. Plasma glucose and Mg2+ levels of animals were measured. Magnesium concentrations increased in the serum following MgSO4 administration (6.05+/-0.57 vs. 2.58+/-0.14 mg/dL in the Mg2+ group, and 7.14+/-0.42 vs. 2.78+/-0.06 mg/dL in the insulin + Mg2+ group, P < 0.01). Plasma glucose levels decreased following hypoglycemia (4+/-0.66 vs. 118+/-2.23 mg/dL in the insulin group, and 7+/-1.59 vs. 118+/-4.84 mg/dL in the insulin + Mg2+ group, P < 0.01). Blood-brain barrier permeability to Evans blue considerably increased in hypoglycemic rats (P < 0.01). In contrast, blood-brain barrier permeability to Evans blue was significantly reduced in treatment of hypoglycemic rats with MgSO4 (P < 0.01). These results indicate that Mg2+ greatly reduced the passage of exogenous vascular tracer bound to albumin into the brain during hypoglycemia with hypothermia. Mg2+ could have protective effects on blood-brain barrier permeability against insulin-induced hypoglycemia.