Dose dependent reduction of glucose utilization by pentobarbital in rat brain.

Abstract

Downloaded fromPENTOBARBITAL AND GLUCOSE UTILIZATION/Crane et al. 13permits the calculation of the rate of brain regional glucose utilization from individual animals, independent of measurements of blood flow and blood brain barrier transport. The additional measurement of brain pentobarbital levels by gas chromatography (GC) has enabled us to report the correlation between brain level of pentobarbital and the rate of glucose utilization. Methods Injection of Barbiturate and Radiolabeled 2-DOGWistar rats (200-250 g), of either sex, were injected intraperitoneally with 0.5 ml of Ringer's solution containing variable doses of sodium pentobarbital. After 10 min, the rats were placed in a restraining device (Gerling-Moore, Palo Alto) and their tails warmed for 30 sec in warm water. A 23 gauge needle was inserted into the dilated tail vein and ten fid of ( 3 H(G)) 2-DOG (sp. act., 10 Ci/mmole) in 0.25 ml of Ringers solution injected, followed by a 0.1 ml wash of Ringers solution. Two minutes later, 2 j*Ci of 2-(l-14 C)-DOG (sp. act., 54 mCi/mmole) in 0.25 ml of Ringers solution were injected through the same needle, followed by 0.1 ml of Ringers solution. Two minutes after the 14 C injection, the rats were killed by intense microwave irradiation.9 Cardiac blood was collected into a heparinized syringe for liquid scintillation counting. The time course of 2-DOG uptake and phosphorylation was determined by single ( 3 H) 2-DOG injections, followed by microwaving at varying times after injection. The brains were excised and stored at -20°C for subsequent analysis. Biochemical And Radiochemical AnalysisThe brains were thawed and the cerebral cortices were weighed and placed in Ten Broeck homogenizers with a measured amount of distilled water (=1.6 ml water -cortex wet wt X 0.8) and 50 #g of sodium secobarbital in 4 ml of methanol to serve as an internal standard in GC. Using a modified procedure of Bligh and Dyer, 10 the cortices were homogenized, 2 ml of chloroform were added, and the tissues were rehomogenized. The homogenates were centrifuged, the supernates saved, and the pellets rehomogenized in 7 ml of chloroform-methanol-water (1:2:0.8) and centrifuged. The supernates were combined, 5 ml of chloroform and 5 ml of water added and the aqueous and organic phases separated after centrifugation. The interfaces were washed three times with methanol-water (1:1), and the washes combined with the aqueous phases.The aqueous phases were further fractionated into neutral (containing glucose and 2-DOG) and anionic (containing 2-deoxy-D-glucose-6-phosphate (2-DOGP)) fractions by ion exchange chromatography on DEAE Sephadex A25 columns (1 ml bed volume). The neutral fractions were recovered unbound from the column, while the anionic fractions were eluted with 0.5M pyridine acetate, pH 5.0. Aliquots were removed from both fractions for simultaneous 3 H and 14 C liquid scintillation counting. The neutral fractions were dried by rotary evaporation and taken up in a 0.1M Tris-HCl (buffered to pH 7.5) and 0.02% sodium azide solution f...

Stoichiometric coupling of brain glucose metabolism and glutamatergic neuronal activity

“…The linear relationship between Glc oxidation and glutamate-neurotransmitter cycling is in agreement with previous high resolution 2-deoxyglucose-autoradiographic studies of intact neurons that demonstrated that, above a baseline metabolic level, Glc utilization is proportional to stimulation frequency (and by inference neuronal firing and neurotransmitter release) (19,20). The rate of oxidative Glc metabolism of Ϸ0.08 mol͞min͞g in group C is somewhat lower than previously reported cortical Glc utilization rates under high dose pentobarbital anesthesia (0.18-0.28 mol͞min͞g) (21,22); however, the longer duration of anesthetic depression in this study prior to the measurement of metabolic rates probably accounts for this difference. Essentially, these results indicated that under mild anesthesia, and by inference the awake state, a major fraction of cortical energy production supports glutamatergic synaptic transmission.…”

Cerebral oxygen availability and blood flow during middle cerebral artery occlusion: effects of pentobarbital.

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