Cerebral Cortex Ammonia and Glutamine Metabolism During Liver Insufficiency-Induced Hyperammonemia in the Rat

Hyperammonemia has been suggested to induce enhanced cerebral cortex ammonia uptake, subsequent glutamine synthesis and accumulation, and finally net glutamine release into the blood stream, but this has never been confirmed in liver insufficiency models. Therefore, cerebral cortex ammonia- and glutamine-related metabolism was studied during liver insufficiency-induced hyperammonemia by measuring plasma flow and venous-arterial concentration differences of ammonia and amino acids across the cerebral cortex (enabling estimation of net metabolite exchange), 1 day after portacaval shunting and 2, 4, and 6 h after hepatic artery ligation (or in controls). The intra-organ effects were investigated by measuring cerebral cortex tissue ammonia and amino acids 6 h after liver ischemia induction or in controls. Arterial ammonia and glutamine increased in portacaval-shunted rats versus controls, and further increased during liver ischemia. Cerebral cortex net ammonia uptake, observed in portacaval-shunted rats, increased progressively during liver ischemia, but net glutamine release was only observed after 6 h of liver ischemia. Cerebral cortex tissue glutamine, gamma-aminobutyric acid, most other amino acids, and ammonia levels were increased during liver ischemia. Glutamate was equally decreased in portacaval-shunted and liver-ischemia rats. The observed net cerebral cortex ammonia uptake, cerebral cortex tissue ammonia and glutamine accumulation, and finally glutamine release into the blood suggest that the rat cerebral cortex initially contributes to net ammonia removal from the blood during liver insufficiency-induced hyperammonemia by augmenting tissue glutamine and ammonia pools, and later by net glutamine release into the blood. The changes in cerebral cortex glutamate and gamma-aminobutyric acid could be related to altered ammonia metabolism.     

The Binding Properties of the Particulate and Solubilized Sulfonylurea Receptor from Cerebral Cortex Are Modulated by the Mg2+ Complex of ATP

Glibenclamide closes an ATP-sensitive K+ channel (K-ATP channel) by interaction with the sulfonylurea receptor in the plasma membrane of pancreatic B cells and thereby initiates insulin release. Previous studies demonstrated that the Mg2+ complex of ATP decreases glibenclamide binding to the sulfonylurea receptor from pancreatic islets. The aim of the present study was to examine the effect of adenine and guanine nucleotides on binding of sulfonyl-ureas to the cerebral sulfonylurea receptor. For this purpose, binding properties of the particulate and solubilized site from rat or pig cerebral cortex were analyzed. Maximum recovery of receptors in detergent extracts amounted to 40-50%. Specific binding of [3H]glibenclamide to the solubilized receptors corresponded well to specific binding to microsomes. In microsomes and detergent extracts, the Mg2+ complexes of ATP, ADP, GTP, and GDP inhibited binding of [3H]glibenclamide. These effects were not observed in the absence of Mg2+. In detergent extracts, Mg-ATP (300 microM) reduced the number of high-affinity sites for [3H]-glibenclamide by 52% and increased the dissociation constant for [3H]glibenclamide by eightfold; Mg-ATP was half-maximally effective at 41 microM. Alkaline phosphatase accelerated the reversal of Mg-ATP-induced inhibition of [3H]glibenclamide binding. The data suggest similar control of the sulfonylurea receptor from brain and pancreatic islets by protein phosphorylation.     

The Effect of Cyclohexyladenosine on the Penischemic Increases of Hippocampal Glutamate and Glycine in the Rabbit

We investigated the ability of N6-cyclohexyladenosine (CHA), a potent and selective agonist of the adenosine A1 receptor, to attenuate elevations of levels of extracellular hippocampal glutamate and glycine that result from episodes of transient global cerebral ischemia (TGCI). A total of 30 New Zealand white rabbits were randomly assigned to receive 0 (n = 5), 0.1 (n = 8), 1.0 (n = 6), 10 (n = 6), or 100 (n = 5) microM CHA. The drug was dissolved in artificial CSF (vehicle) and administered via a microdialysis probe placed stereotactically into the dorsal hippocampus. A second microdialysis probe placed into the contralateral hippocampus of each animal was perfused with vehicle alone. Ten minutes of TGCI was induced by neck tourniquet inflation and deliberate hypotension from 0 to 10 min. Microdialysis samples were collected as follows: every 20 min preischemia (at -80, -60, -40, -20, and 0 min); every 5 min during ischemia and in the immediate reperfusion period (at 5, 10, 15, and 20 min); and every 20 min for the remainder of the reperfusion period (at 40, 60, and 80 min). Samples were then analyzed for their concentration of glutamate and glycine by HPLC. Following 10 min of ischemia, glutamate levels increased to a peak of 3.28 +/- 0.55 times baseline and returned to preischemic levels by 40 min, i.e., during reperfusion. Glycine concentrations increased to 5.41 +/- 0.91 times over baseline and remained elevated for the duration of the study.(ABSTRACT TRUNCATED AT 250 WORDS)     

Amino Acid Neurotransmitter Receptor Changes in Cerebral Cortex in Alcoholism: Effect of Cirrhosis of the Liver

Gamma-aminobutyric acidA/benzodiazepine receptor binding sites and the N-methyl-D-aspartate subclass of glutamate receptor sites were assessed in synaptic plasma membrane homogenates of cerebral cortex tissue obtained at autopsy from cirrhotic and noncirrhotic alcoholic patients and matched control subjects. The alcoholic patients consumed an average of greater than 80 g of ethanol/day, the control subjects less than 20 g/day. Postmortem delays up to approximately 100 h caused no significant loss of any of the binding sites; the patient and subject groups were closely matched for age. The affinities (KD) of the receptor sites did not differ between the patient and subject groups, nor between cortical regions. Using three different radioligands ([3H]muscimol, [3H]flunitrazepam, and [3H]diazepam), the gamma-aminobutyric acidA/benzodiazepine receptor complex was found to have greater density (Bmax) in superior frontal gyrus in alcoholic patients (which selectively shows morphological change in alcoholic patients), but was unchanged in motor cortex. Alcoholic patients with cirrhosis had much less pronounced changes. The density of the N-methyl-D-aspartate subclass of glutamate receptors, assessed with [3H]MK-801, did not vary across patient and subject groups.     

Comparison of Rates of Local Cerebral Glucose Utilization Determined with Deoxy[l-14C]glucose and Deoxy[6-14C]glucose

The activity of the pentose phosphate shunt pathway in brain is thought to be linked to neurotransmitter metabolism, glutathione reduction, and synthetic pathways requiring NADPH. There is currently no method available to assess flux of glucose through the pentose phosphate pathway in localized regions of the brain of conscious animals in vivo. Because metabolites of deoxy[1-14C]glucose are lost from brain when the experimental period of the deoxy[14C]glucose method exceeds 45 min, the possibility was considered that the loss reflected activity of this shunt pathway and that this hexose might be used to assay regional pentose phosphate shunt pathway activity in brain. Decarboxylation of deoxy[1-14C]glucose by brain extracts was detected in vitro, and small quantities of 14C were recovered in the 6-phosphodeoxygluconate fraction when deoxy[14C]glucose metabolites were isolated from freeze-blown brains and separated by HPLC. Local rates of glucose utilization determined with deoxy[1-14C]glucose and deoxy[6-14C]glucose were, however, similar in 20 brain structures at 45, 60, 90, and 120 min after the pulse, indicating that the rate of loss of 14CO2 from deoxy[1-14C]glucose-6-phosphate in normal adult rat brain is too low to permit assay pentose phosphate shunt activity in vivo. Further metabolism of deoxy[1-14]glucose-6-phosphate via this pathway does not interfere during routine use of the deoxyglucose method or explain the progressive decrease in calculated metabolic rate when the experimental period exceeds 45 min.     

Regional and Temporal Alterations in Ca2+/Calmodulin-Dependent Protein Kinase II and Calcineurin in the Hippocampus of Rat Brain After Transient Forebrain Ischemia

We have investigated regional and temporal alterations in Ca2+/calmodulin-dependent protein kinase II (CaM kinase II) and calcineurin (Ca2+/calmodulin-dependent protein phosphatase) after transient forebrain ischemia. Immunoreactivity and enzyme activity of CaM kinase II decreased in regions CA1 and CA3, and in the dentate gyrus, of the hippocampus early (6-12 h) after ischemia, but the decrease in immunoreactivity gradually recovered over time, except in the CA1 region. Furthermore, the increase in Ca2+/calmodulin-independent activity was detected up to 3 days after ischemia in all regions tested, suggesting that the concentration of intracellular Ca2+ increased. In contrast to CaM kinase II, as immunohistochemistry and regional immunoblot analysis revealed, calcineurin was preserved in the CA1 region until 1.5 days and then lost with the increase in morphological degeneration of neurons. Immunoblot analysis confirmed the findings of the immunohistochemistry. These results suggest that there is a difference between CaM kinase II and calcineurin in regional and temporal loss after ischemia and that imbalance of Ca2+/calmodulin-dependent protein phosphorylation-dephosphorylation may occur.     

An analog of ACTH/MSH (4–9), ORG-2766, reduces cerebral uptake of morphine

The uptake of morphine was significantly reduced in most regions of the brains of conscious, unrestrained rats within 10 minutes after treatment with an analog of ACTH/MSH (4–9), ORG-2766. The effect was most obvious in regions with significant densities of enkephalin receptors, namely basal ganglia, hippocampus and cortex. The results explain, in part, how some fragments and analogs of ACTH/MSH may antagonize behavioral actions of morphine, even though some of these peptides lack significant opiate receptor binding properties. We believe that this effect of ORG-2766 is related to an action on the permeability characteristics of the brain microvasculature. The underlying mechanism is unknown.     

Gastro-intestinal and neurohormonal peptides in the alimentary tract and cerebral complex ofCiona intestinalis (Ascidiaceae)

Summary Polypeptide-hormone producing cells were localized in the alimentary tract and cerebral ganglion ofCiona intestinalis using cytochemical, immunocytochemical and electron-microscopical methods.Antisera to the following peptides of vertebrate type were employed: bombesin, human prolactin (hPRL), bovine pancreatic polypeptide (PP), porcine secretin, motilin, vasoactive intestinal polypeptide (VIP),-endorphin, leu-enkephalin, met-enkephalin, neurotensin, 5-hydroxytryptamin (5-HT), cholecystokinin (CCK), human growth hormone (GH), ACTH, corticotropin-like intermediate lobe peptide (CLIP) and gastric inhibitory peptide (GIP).Immunoreactive cells were found both in the alimentary tract epithelium and in the cerebral ganglion for bombesin, PP, substance P, somatostatin, secretin and neurotensin. Additionally, in the cerebral ganglion only, there were cells immunoreactive for-endorphin, VIP, motilin and human prolactin. 5-HT positive cells, however, were restricted to the alimentary tract.No immunoreactivity was obtained either in the cerebral ganglion or in the alimentary tract with antibodies to leu-enkephalin, met-enkephalin, CCK, growth hormone, ACTH, CLIP and GIP. Prolactin-immunoreactive and pancreatic polypeptide-immunoreactive cells were argyrophilic with the Grimelius' stain and were found in neighbouring positions in the cerebral ganglion.At the ultrastructural level five differently granulated cell types were distinguished in the cerebral ganglion. Granules were present in the perikarya as well as in axons. The possible functions of the peptides as neurohormones, neuroregulators and neuromodulators are discussed.     

Alterations of Phospholipid Metabolism in Rat Cerebral Cortex Mince Induced by Cationic Amphiphilic Drugs

Abstract Cationic amphiphilic drugs (CADs) of varied clinical use were screened to determine their capacity to alter the pattern of labeling with ³²Pj of cerebral cortex mince phospholipids. The altered phospholipid labeling patterns were qualitatively similar, the prominent features being reduced incorporation into phosphatidylcholine and increased incorporation into phosphatidic acid. Relative potencies were: (±)-propranolol > chlorpromazine = 4,4'-bis(diethylaminoethoxy) α,β -diethyldiphenylethane > desipramine > di-bucaine > pimozide > oxymetazoline = fenfluramine = haloperidol = chloroquine > amphetamine = no drug added. Propranolol was used to study the action of CADs further. Its effect was time- and dose-dependent, but in contrast with pineal gland, no label appeared in phosphatidyl-CMP (CDP-diacylglycerol), nor did dialysis of the mince to reduce diffusible substrates or exogenous addition of substrates cause appearance of liponucleotide. Thus lack of diffusible precursors is not responsible for CAD effects in vitro. Pulse-chase experiments with ³²P₁ and [2-³H]glycerol suggested that inhibition of phosphatidate phosphohydrolase may be partly responsible for the observed alterations in phospholipid labeling in the presence of CADs.     

Uptake of 36Cl and 22Na by the Brain-Cerebrospinal Fluid System: Comparison of the Permeability of the Blood-Brain and Blood-Cerebrospinal Fluid Barriers

Abstract: Transport and permeability properties of the blood-brain and blood-CSF barriers were determined by kinetic analysis of radioisotope uptake from the plasma into the CNS of the adult rat. Cerebral cortex and cerebellum uptake curves for ³⁶Cl and ²²Na were resolved into two components. The fast component (t_½ 0.02–0.05 h, fractional volume 0.04–0.08) is comprised of the vascular compartment and a small perivascular space whereas the slow component (t_½ 1.06–1.69 h, fractional volume 0.92–0.96) represents isotope movement across the blood-brain barrier into the brain extracellular and cellular compartments. Uptake curves of both ³⁶Cl and ²²Na into the CSF were also resolved into two components, a fast component (t_½ 0.18 h, fractional volume 0.24) and a slow component (t_½ 1.2 h, fractional volume 0.76). Evidence suggests that the fast component represents isotope movement across the blood-CSF barrier, i.e., the choroid plexuses, whereas the CSF slow component probably reflects isotope penetration primarily from the brain extracellular fluid into the CSF. The extracellular fluid volume of the cerebral cortex and cerebellum was estimated as ?13% from the initial slope of the curve of brain space versus CSF space curve for both ³⁶Cl and ²²Na. Like the choroid plexuses, the glial cell compartment of the brain appears to accumulate Cl from 2 to 6 times that predicted for passive distribution. The relative permeability of the blood-CSF and blood-brain barriers to ³⁶Cl, ²²Na, and [³H]mannitol was determined by calculating permeability surface-area products (PA). Analysis of the PA values for all three isotopes indicates that the effective permeability of the choroidal epithelium (blood/CSF barrier) is significantly greater than that of the capillary endothelium in the cerebral cortex and cerebellum (blood-brain barrier).