Cerebral metabolism in newborn dogs during reversible asphyxia

Acute, systemic asphyxia was induced in paralyzed, lightly anesthetized 1- to 2-day-old dogs by respiratory arrest. Measurements of arterial blood pressure, acid-base balance, and concentrations of pyruvate, lactate, and glucose in blood were correlated with electroencephalographic activity and with concentrations of high-energy phosphates and glycolytic substrates in the cerebral cortex. Most animals tolerated 15 minutes, but not 20 minutes, of asphyxia with apparently normal behavioral recovery. Asphyxia was always accompanied by bradycardia, systemic hypotension, and a progressive decline in arterial pH; the concentration of blood lactate and the lactate/pyruvate ratio rose. Blood glucose levels were unaffected, at least during the first 10 minutes. Concentrations of glucose and phosphocreatine in cerebral cortex declined rapidly during asphyxia to low levels, but levels of adenosine triphosphate remained within normal limits for up to 5 minutes despite the fact that electrical activity in brain ceased within 2 minutes. The intravenous injection of carbon black into animals asphyxiated for 2^1/2, 5, 10, or 15 minutes revealed substantial reductions in blood flow to brain during asphyxia; however, relative to the cerebral cortex, brainstem structures received a preferential blood supply.     

Cerebral carbohydrate metabolism during severe ischemia in fetal sheep

The effect of cephalic hypotension on brain metabolism was studied in 10 unanesthetized, normoxic (PaO2 greater than 17 mm Hg), late-gestation fetal lambs. Perfusion pressure (cephalic arterial minus sagittal venous pressure) was 40 +/- 1 mm Hg (SEM) during control and was reduced to 10 +/- 1 by occlusion of the Grachio-cephalic artery. Cerebral blood flow was measured with microspheres, and arterial and sagittal vein blood samples were analyzed for oxygen content, glucose, and lactate. During the occlusion, oxygen consumption decreased from 125 +/- 8 to 95 +/- 4 (p less than 0.05) (all values mumol 100 g-1 min-1), and glucose uptake increased from 20 +/- 3 to 25 +/- 1 (p less than 0.05). During the control period, there was no net lactate flux; during the occlusion, lactate excretion was 5.7 +/- 1.4 (p less than 0.005). The control glucose and oxygen uptakes demonstrated a normal 6:1 molar ratio; however, during the occlusion, 9.4 mumol 100 g-1 glucose min-1 were taken up in excess of expected aerobic glucose metabolism. If all of this glucose were anaerobically metabolized to lactate, three times the measured efflux would be produced. The transport properties of the fetal blood-brain barrier may be important factors in perinatal brain injury.     

Cerebral carbohydrate metabolism during acute carbon monoxide intoxication.

The cerebral metabolic effects of 2.5, 5, 7.5, 10, 20, 30 and 60 min exposure to 1% CO were studied in lightly anesthetized rats by measurement of cerebral cortical contents of selected glycolytic and citric acid cylce intermediates, as well as tissue energy phosphates. The initial change in the glycolytic sequence occurred at 2.5 min with decreases in tissue glucose and glucose-6-phosphate and increases in fructose-1-6-diphosphate which indicated an activation of phosphofructokinase and hexokinase. The "crossover" pattern between glucose-6-phosphate and fructose-1,6-diphosphate was present at 5, 7.5 and 10 min, but not at 20, 30 and 60 min and thus confirmed previous observations that detection of phosphofructokinase activation in acute unifactorial cerebral hypoxia requires tissue study during the early phases of the experimental exposure. The initial activation of phosphofructokinase occurred in the absence of detectable changes in the tissue content of ATP, ADP, AMP or phosphocreatine and therefore suggested that an imbalance of tissue energy homeostasis is not a prerequisite for the activation of glycolysis in CO intoxication. One percent CO resulted in an increasing malate/oxaloacetate ratio at 5 min, followed by a decrease in alpha-ketoglutarate and aspartate at 7.5 min which suggested a shift in the aspartate aminotransferase reaction towards the replenishment of oxaloacetate removed via the malate dehydrogenase reaction. Subsequent increases in alpha-ketoglutarate at 10, 20, 30 and 60 min were associated with increases in alanine, indicating a contributing role for a secondary shift of the alanine aminotransferase reaction in the replenishment of alpha-ketoglutarate. A comparison of the CO induced changes in the glycolytic and citric acid cycle pathways with those seen in acute hypoxemia indicates no basic qualitative differences in the metabolic responses of brain tissue to the two conditions.     

Cerebral transport and metabolism of 1-11C-D-glucose during stepped hypoglycemia

Attempts to measure blood-to-brain glucose transport and cerebral glucose metabolism with ¹¹C-glucose have been hampered by methods that require jugular venous sampling or do not adequately account for the efflux of labeled metabolites from the brain. We performed eight positron emission tomography studies with 1-¹¹C-d-glucose in macaques at arterial plasma glucose concentrations of 8.43 to 1.51 μmol ml^?1 (152–27 mg dl^?1) using a model that includes a fourth rate constant to account for regional egrees of all ¹¹C-metabolites. Values for blood-to-brain glucose influx, cerebral glucose metabolism, and brain free glucose concentration agreed closely with values obtained in mammals by other investigators. Values for net extraction fraction corresponded closely to simultaneously measured arteriovenous values. We demonstrated that utilization of a model that includes a fourth rate constant to account for regional egress of all ¹¹C-metabolites with positron emission tomography and 1-¹¹C-d-glucose provides accurate measurements of blood-to-brain glucose transport and cerebral glucose metabolism in vivo without need for jugular venous sampling, even under conditions of severe hypoglycemia.     

二氢石蒜碱对大鼠海马脑片缺氧缺糖损伤的影响

目的探讨二氢石蒜碱(DL)对大鼠海马脑片缺糖缺氧损伤的影响。方法在大鼠离体海马脑片上,应用细胞外记录技术,记录CAl1区群峰电位,以群峰电位(PS)持续时间以及恢复程度作为判断药效标准。结果对照组及二氢石蒜碱组PS持续时间分别为335.13±74.73s和761.50±268.30s,显示DL组缺糖缺氧后PS持续时间明显延长(与对照组比较P＜0.05),而哌唑嗪在1×10-6mol·L-1时无影响,持续缺糖缺氧7min30s后,对照组PS幅度为4.93％±7.78％,而DL(1×10-6mol·L-1)组仍保持48.22％±27.08％;复糖复氧10minDL(1×10-6mol·L-1及1×10-5mol·L-1)组PS恢复较对照组好(P＜0.05),对照组、DL(1×10-6mol·L-1)组、DL(1×10-5mol·L-1)组及酚妥拉明(5×10-5mol·L-1)组PS分别恢复到缺糖缺氧前14.17％±34.70％、94.30％±49.10％、71.00％±41.40％、71.40％±46.02％。结论DL对海马脑片缺糖缺氧损伤有明显的保护作用。其机理可能是为DL阻滞肾上腺素能受体及改善能量代谢,从而提高了海马脑片抗缺氧缺糖损伤的能力。     

Cerebral blood flow during chronic hypoglycemia in the rat

Regional cerebral blood flow (rCBF) was measured in normoglycemic and chronically hypoglycemic rats. Chronic hypoglycemia was produced by continuously infusing insulin for 6-7 days. During chronic hypoglycemia (plasma glucose = 1.97 mumol/ml), rCBF increased in all regions except the cerebellum and hypothalamus. Blood flow increases present during chronic hypoglycemia were not as great as those previously measured during acute hypoglycemia. Therefore, adjustments in the regulation of rCBF occurred during chronic hypoglycemia compared to acute hypoglycemia.     

Modification of glutamate binding sites in newborn brain during hypoglycemia

We have shown that acute insulin-induced hypoglycemia leads to specific changes in the cerebral NMDA receptor-associated ion channel in the newborn piglet. The present study tests the hypothesis that exposure to acute hypoglycemia in the newborn will alter the glutamate binding site of both NMDA and kainate receptors. Studies were performed in 3-6 days-old piglets randomized to control (n=6) or hypoglycemic (n=6) groups. Hypoglycemia was maintained for 120 min using insulin infusion. Saturation binding assays were performed in cerebral cell membranes using (3)H-glutamate or (3)H-kainate to determine the characteristics of the glutamate binding sites of the NMDA and kainate receptors, respectively. The concentration of glucose in cerebral cortex was 10-fold less in hypoglycemic piglets than in controls (P<0.05). Brain ATP was not significantly decreased during hypoglycemia, but phosphocreatine decreased from control of 6.6 +/- 1.3 micromoles/g brain to 3.2 +/- 1.9 micromoles/g brain in hypoglycemic piglets. The B(max) for NMDA-displaceable (3)H-glutamate binding was 992 +/- 64 fmol/mg protein in hypoglycemic animals, significantly higher than the control value of 746 +/- 42 fmol/mg protein. However, the dissociation constant for glutamate was unchanged during hypoglycemia. The (3)H-kainate binding studies demonstrated no change in B(max) of high-affinity kainate receptors during hypoglycemia. In contrast, the affinity of the kainate receptor glutamate binding site significantly increased compared to control. Thus, acute hypoglycemia in the newborn piglet had specific effects on the glutamate binding sites of the NMDA and kainate receptors that could be due to alterations in cell membrane lipids or modification of receptor proteins.     

Effects of fasting and insulin-induced hypoglycemia on brain cell membrane function and energy metabolism during hypoxia-ischemia in newborn piglets.

This study was done to determine the effects of 12 h fasting-induced mild hypoglycemia (blood glucose 60 mg/dl) and insulin-induced moderate hypoglycemia (blood glucose 35 mg/dl) on brain cell membrane function and energy metabolism during hypoxia-ischemia in newborn piglets. Sixty-three ventilated piglets were divided into six groups; normoglycemic control (NC, n=8), fasting-induced mildly hypoglycemic control (FC, n=10), insulin-induced moderately hypoglycemic control (IC, n=10), normoglycemic/hypoxic-ischemic (NH, n=11), fasting-induced mildly hypoglycemic/hypoxic-ischemic (FH, n=12) and insulin-induced moderately hypoglycemic/hypoxic-ischemic (IH, n=12) group. Cerebral hypoxia-ischemia was induced by occlusion of bilateral common carotid arteries and simultaneous breathing with 8% oxygen for 30 min. The brain lactate level was elevated in NH group and this change was attenuated in FH and IH groups. The extent of cerebral lactic acidosis during hypoxic-ischemic insult showed significant positive correlation with blood glucose level (r=0.55, p<0.001). Cerebral Na+, K+-ATPase activity and concentrations of high-energy phosphate compounds were reduced in NH group and these changes were not ameliorated in FH or IH group. Cortical levels of conjugated dienes, measured as an index of lipid peroxidation of brain cell membrane, were significantly elevated in NH, FH and IH groups compared with NC, FC and IC groups and these increases were more profound in FH and IH with respect to NH. Blood glucose concentration showed significant inverse correlation with levels of conjugated dienes (r=-0.35, p<0.05). These findings suggest that, unlike in adults, mild or moderate hypoglycemia, regardless of methods of induction such as fasting or insulin-induced, during cerebral hypoxia-ischemia is not beneficial and may even be harmful in neonates.     

Blood flow and metabolism in heterotopic cerebellar grafts during hypoglycemia

Summary Hypoglycemia-induced disturbances of brain metabolism and neuronal injury exhibit a distinct predilection for forebrain structures, in particular the caudate-putamen, hippocampus and cerebral cortex, whereas the cerebellum is remarkably resistant. In an attempt to assess the biological basis of this differential regional vulnerability, we have used a neural transplantation technique to compare hemodynamic and metabolic changes in cerebellum during servere hypoglycemia with those in heterotopic cerebellar grafts. To this end, the cerebellar anlage of fetal rat brain (day 15 of gestation) was stereotactically transplanted into the vulnerable caudate-putamen. Following a differentiation period of 8 weeks the grafts had developed into an organotypic population of mature cells with laminar histoarchitecture. Host animals were then subjected to insulin-induced hypoglycemia. After 15 min of isoelectric EEG, blood flow was increased throughout the brain but residual glucose consumption was significantly higher in cerebellum (0.29 mol/g per min) and cerebellar grafts (0.22 mol/g per min) as a result of increased glucose extraction. Hypoglycemia caused a depletion of ATP in all brain structures except cerebellum where normal levels were maintained. Correlation of local ATP content and glucose utilization revealed a threshold-like decline of ATP at a glucose utilization rate of 0.27 mol/g per min. ATP, in consequence, was normal in cerebellum but partially depleted in cerebellar grafts. It is concluded that the resistance of cerebellum to hypoglycemia is due to its capacity for higher glucose extraction at low blood glucose levels, and that this unique intrinsic property is preserved after heterotopic transplantation.Supported in part by the Deutsche Forschungsgemeinschaft (SFB 70)     

Cerebral ammonia metabolism in normal and hyperammonemic rats

Brain ammonia is generated from many enzymatic reactions, including glutaminase, glutamate dehydrogenase, and the purine nucleotide cycle. In contrast, the brain possesses only one major enzyme for the removal of exogenous ammonia, i.e., glutamine synthetase. Thus, following administration of [¹³N]ammonia to rats [via either the carotid artery or cerebrospinal fluid (csf)], most metabolized label was in glutamine (amide) and little was in glutamate (plus aspartate). Since blood-and csf-borne ammonia are converted to glutamine largely, if not entirely, in the astrocytes, it is not possible from these types of experiments to predict with certainty the metabolic fate of the bulk of endogenously produced ammonia. By comparing the specific activity ofl-[¹³N]glutamate to that ofl-[amine-¹³N]glutamine following intracarotid [¹³N]ammonia administration it was concluded that metabolic compartmentation is no longer intact in the brains of rats treated with the glutamine synthetase inhibitorl-methionine-SR-sulfoximine (MSO) and that blood and brain ammonia pools mix in such animals. In MSO-treated animals, recovery of label in brain was low (～20% of controls), and of the label remaining, a prominent portion was in glutamine (amide) (despite an 87% decrease in brain glutamine synthetase activity). These data are consistent with the hypothesis that glutamine synthetase is the major enzyme for metabolism of endogenously—as well as exogenously—produced ammonia. The rate of turnover of blood-derived ammonia to glutamine in normal rat brain is extremely rapid (t _1/2≤3 s), but is slowed in the brains of chronically (12–14-wk portacaval-shunted) or acutely (urease-treated) hyperammonemic rats (t _1/2≤10 s). The slowed turnover rate may be caused by an increased astrocytic ammonia, decreased glutamine synthetase activity, or both. In the hyperammonemic rat brain, glutamine synthetase is still the only important enzyme for the removal of blood-borne ammonia. Hyperammonemia causes an increase in brain lactate/pyruvate ratios and decreases in brain glutamate and brainstem ATP, consistent with an interference with the malate-aspartate shuttle. In vitro, pathological levels of ammonia also inhibit brain α-ketoglutarate dehydrogenase complex and, less strongly, pyruvate dehydrogenase complex. The rat brain does not adapt to prolonged hyperammonemia by increasing its glutamine synthetase activity. Therefore, since the brain only has a limited capacity to buffer against excess ammonia, it is important that diseases in which hyper-ammonemia is a prominent feature be treated to reduce the associated hyperammonemia as much as possible.     

Cerebral blood flow and tissue metabolism in experimental cerebral ischemia of spontaneously hypertensive rats with hyper-, normo-, and hypoglycemia

The present study was designed to clarify the effect of blood glucose level on cerebral blood flow and metabolism during and after acute cerebral ischemia induced by bilateral carotid ligation (BCL) in spontaneously hypertensive rats (SHR). Blood glucose levels were varied by intraperitoneal infusion of 50% of glucose (hyperglycemia), insulin with hypertonic saline (hypoglycemia) or hypertonic saline (normoglycemia). Cerebral blood flow (CBF) in the parietal cortex and thalamus was measured by hydrogen clearance technique, and the supratentorial metabolites of the brain frozen in situ were determined by the enzymatic method. In non-ischemic animals, blood glucose levels had no influence on the supratentorial lactate, pyruvate or adenosine triphosphate (ATP) concentrations. In ischemic animals, however, cortical CBF was reduced to less than 1% of the resting value at 3 hours after BCL. However, there were no substantial differences of CBF during and after ischemia among 3 glycemic groups. Cerebral lactate in the ischemic brain greatly increased in hyperglycemia (34.97 +/- 1.29 mmol/kg), moderately in normoglycemia (23.43 +/- 3.13 mmol/kg) and less in hypoglycemia (7.20 +/- 1.54 mmol/kg). In contrast, cerebral ATP decreased in hyperglycemia (0.93 +/- 0.19 mmol/kg) as much as it did in normoglycemia (1.04 +/- 0.25 mmol/kg), while ATP reduction was much greater in hypoglycemia (0.45 +/- 0.05 mmol/kg). At 1-hour recirculation after 3-hour ischemia, ATP tended to increase in all groups of animals, indicating the recovery of energy metabolism. Such metabolic recovery after recirculation was good in hypo- and normoglycemia, and was also evident in hyperglycemia. Our results suggest that hyperglycemia is not necessarily an unfavorable condition in acute incomplete cerebral ischemia.     

Cerebral protein synthesis during long-term recovery from severe hypoglycemia

Regional protein synthesis was investigated in the rat brain during long-term recovery from insulin-induced hypoglycemia with 30 min of cerebral electrical silence. At various time intervals up to 14 days after glucose replenishment, animals received a single dose of L-[3,5-3H]tyrosine and were killed 30 min later. Brains were processed for autoradiography using the stripping film technique. Although hypoglycemia sufficiently severe to cause cessation of EEG activity leads to almost complete inhibition of amino acid incorporation in all "vulnerable" forebrain structures (cerebral cortex, hippocampus, caudoputamen), autoradiographs revealed a very specialized sequence with differential posthypoglycemic restoration of biosynthetic activity in certain neuronal cell types. Three major subpopulations could be distinguished: Neurons that fully regained their protein synthetic capacity within 6 h following hypoglycemia (cortical neurons of layer III-VI, large neurons in the caudoputamen, CA3 and CA4 pyramidal neurons, the majority of granule cells of the dentate gyrus) seemed to escape neuronal necrosis. Prolonged impairment of protein synthesis with only partial restoration during the early posthypoglycemic recovery period (CA1 neurons, most small- to medium-sized neurons of the caudoputamen) carried an increased risk of permanent cell damage. The large majority of these neurons, however, showed full recovery of protein synthesis as late as 7 days after hypoglycemia. Neurons with complete lack of amino acid incorporation after 6 h of recovery (granule cells at the crest of the dentate gyrus, small neurons of the dorsolateral caudoputamen) never resumed protein synthesis, regressed, and died. These studies in conjunction with morphological analysis indicate that the sequential recovery of protein synthesis reflects the extent to which neuronal populations are at risk during severe hypoglycemia.     

Relationship between ion gradients and neurotransmitter release in the newborn rat striatum during anoxia

It has been well documented that mammalian newborns are more resistant to hypoxia than adults. The mechanisms for this tolerance has attracted considerable attention due to its clinical implications. Recently, there has been interest in comparing the mechanisms involved in such tolerance with those of turtle brain, which has shown a remarkable tolerance to anoxia. In the latter, much attention has been paid to the role of neurotransmitters in regulating brain metabolic rate. In order to investigate this phenomenon in the mammalian neonate the pattern of neurotransmitter release with respect to pre- and postdepolarization stages was determined. Microdialysis was used to ascertain levels of neurotransmitters in the striatum of 5-day-old rats. Ion homeostasis was determined with a potassium-selective microelectrode. We report here that during anoxia at the predepolarization stage purines (inosine, hypoxanthine, xanthine and adenosine) were significantly released. However, amino acids (glutamate, gamma-amino butyric acid (GABA), aspartate and taurine) remained low during the first 30 min, but were released during anoxic depolarization. It was concluded that mammalian neonate brain differs from that of the turtle in hypoxic adaptation, which may be consequence of its comparatively undifferentiated state.     

Effects of fasting and insulin-induced hypoglycemia on brain cell membrane function and energy metabolism during hypoxia–ischemia in newborn piglets

This study was done to determine the effects of 12 h fasting-induced mild hypoglycemia (blood glucose 60 mg/dl) and insulin-induced moderate hypoglycemia (blood glucose 35 mg/dl) on brain cell membrane function and energy metabolism during hypoxia–ischemia in newborn piglets. Sixty-three ventilated piglets were divided into six groups; normoglycemic control (NC, n=8), fasting-induced mildly hypoglycemic control (FC, n=10), insulin-induced moderately hypoglycemic control (IC, n=10), normoglycemic/hypoxic–ischemic (NH, n=11), fasting-induced mildly hypoglycemic/hypoxic–ischemic (FH, n=12) and insulin-induced moderately hypoglycemic/hypoxic–ischemic (IH, n=12) group. Cerebral hypoxia–ischemia was induced by occlusion of bilateral common carotid arteries and simultaneous breathing with 8% oxygen for 30 min. The brain lactate level was elevated in NH group and this change was attenuated in FH and IH groups. The extent of cerebral lactic acidosis during hypoxic–ischemic insult showed significant positive correlation with blood glucose level (r=0.55, p<0.001). Cerebral Na⁺, K⁺-ATPase activity and concentrations of high-energy phosphate compounds were reduced in NH group and these changes were not ameliorated in FH or IH group. Cortical levels of conjugated dienes, measured as an index of lipid peroxidation of brain cell membrane, were significantly elevated in NH, FH and IH groups compared with NC, FC and IC groups and these increases were more profound in FH and IH with respect to NH. Blood glucose concentration showed significant inverse correlation with levels of conjugated dienes (r=−0.35, p<0.05). These findings suggest that, unlike in adults, mild or moderate hypoglycemia, regardless of methods of induction such as fasting or insulin-induced, during cerebral hypoxia–ischemia is not beneficial and may even be harmful in neonates.     

Protein synthesis in rat brain in hypoxia, anoxia and hypoglycemia

The effect of cerebral hypoxia on protein synthesis was investigated by exposing rats to 5% O2, and examining polypeptide synthesis and size distribution profiles of ribosomes. The findings were compared with the results from cerebral anoxia (decapitation) and hypoglycemia. In cerebral hypoxia there was suppression of polypeptide synthesis, though to a lesser extent than in cerebral anoxia, while no effect was detected in hypoglycemia. Among 4 different ribosomal fractions used for polypeptide synthesis, the microsome was the most sensitive for hypoxia and anoxia, and the polyribosome after short centrifugation was the least sensitive. The size distribution profiles of 3 different ribosomes revealed an increase in the size of the monomere-dimer complex and a decrease of the polysome peak both in cerebral hypoxia and anoxia. Comparison of the energy state and the extent of lactic acidosis in cerebral hypoxia, anoxia and hypoglycemia available in the literature and the functional and structural state of polyribosomes in the present investigation suggests that intracellular acidosis may be the main cause of the suppression of polypeptide synthesis and disaggregation of polyribosomes in hypoxia, and the depletion of energy reserve may be the main cause in anoxia-ischemia.     

Cerebral ketone metabolism during development and injury

Cerebral metabolism of ketones is a normal part of the process of brain development. While the mature brain relies on glucose as a primary fuel source, metabolism of ketone bodies remains an alternative energy source under conditions of starvation. The neuroprotective properties of brain ketone metabolism make this alternative substrate a viable therapeutic option for various pathologies. Since the ability to revert to utilizing ketones as an alternative substrate is greatest in the younger post-weaned brain, this particular therapeutic approach remains an untapped resource particularly for pediatric pathological conditions.