ACID-LABILE PHOSPHATE CONTENT OF INCUBATED BRAIN SLICES

1959 ◽  
Vol 37 (1) ◽  
pp. 999-1010 ◽  
Author(s):  
Hanna M. Pappius ◽  
Dorothy McLean Johnson ◽  
K. A. C. Elliott

During aerobic incubation of brain cortex slices in glucose-containing medium at 38° the pyrophosphate content of the tissue increases rapidly for 15 minutes and then more slowly. The creatine phosphate content increases rapidly during the first 5 minutes and then falls. Anaerobically, the pyrophosphate falls during the first 30 minutes; the creatine phosphate tends to rise initially and then falls to a lower level than that found aerobically. In the absence of glucose and oxygen the levels of both substances fall rapidly to low levels but can be raised again by aerobiosis in the presence of glucose. In the absence of glucose and oxygen in ice-cold medium, or at 38° in the absence of any medium, the levels of pyrophosphate and creatine phosphate are preserved. More inorganic phosphate is produced when slices are incubated than can be accounted for by breakdown of acid-labile phosphate compounds.It is shown that the initial rate of anaerobic glycolysis by slices which have been subjected to various pretreatments is strongly correlated with the pyrophosphate content of the tissue.The failure of slices to concentrate potassium when immersed in sodium-free media is not due to failure to maintain pyrophosphate levels.

1959 ◽  
Vol 37 (8) ◽  
pp. 999-1010 ◽  
Author(s):  
Hanna M. Pappius ◽  
Dorothy McLean Johnson ◽  
K. A. C. Elliott

During aerobic incubation of brain cortex slices in glucose-containing medium at 38° the pyrophosphate content of the tissue increases rapidly for 15 minutes and then more slowly. The creatine phosphate content increases rapidly during the first 5 minutes and then falls. Anaerobically, the pyrophosphate falls during the first 30 minutes; the creatine phosphate tends to rise initially and then falls to a lower level than that found aerobically. In the absence of glucose and oxygen the levels of both substances fall rapidly to low levels but can be raised again by aerobiosis in the presence of glucose. In the absence of glucose and oxygen in ice-cold medium, or at 38° in the absence of any medium, the levels of pyrophosphate and creatine phosphate are preserved. More inorganic phosphate is produced when slices are incubated than can be accounted for by breakdown of acid-labile phosphate compounds.It is shown that the initial rate of anaerobic glycolysis by slices which have been subjected to various pretreatments is strongly correlated with the pyrophosphate content of the tissue.The failure of slices to concentrate potassium when immersed in sodium-free media is not due to failure to maintain pyrophosphate levels.


1977 ◽  
Vol 55 (3) ◽  
pp. 356-362 ◽  
Author(s):  
A. M. Benjamin ◽  
J. H. Quastel

Acetylcholine, in presence of eserine, has little or no effect on the potassium-ion-suppressed concentrative uptakes of GABA and taurine by rat brain cortex slices in contrast with its effect on those of L-glutamate, L-aspartate, and glycine. Potassium ions at a concentration of 30 μequiv./ml in the incubation medium has a marked suppressive effect on the uptakes of GABA and taurine when there is no apparent change in the sodium ion content of the brain tissue. It is concluded that some factor, besides the change in sodium gradient, operates in the mechanism of potassium suppression of GABA and taurine uptakes. Acetylcholine diminishes the potassium-evoked release of endogenous GABA and taurine from brain slices. Its action is Ca2+ dependent and is diminished by atropine. Acetylcholine does not affect the potassium-accelerated release of GABA from brain slices previously loaded with this amino acid. The differences in uptake and release phenomena exhibited by GABA and taurine from those of L-glutamate and L-aspartate may be due to differences between the mechanisms, as well as the sites, of cerebral uptake and release of these two groups of amino acids.


1972 ◽  
Vol 126 (4) ◽  
pp. 851-867 ◽  
Author(s):  
R. Shankar ◽  
J. H. Quastel

1. Tetrodotoxin, at concentrations at which it abolishes generation of action potentials in the nervous system, enhances by about 300% the rate of anaerobic glycolysis of brain-cortex slices from adult rats, or from adult and infant guinea pigs. This occurs to a greater extent in Ca2+-deficient incubation media than in Ca2+-rich media. Tetrodotoxin has no accelerative effect on cerebral aerobic glycolysis. 2. Tetrodotoxin does not affect the rate of anaerobic glycolysis of 2-day-old rat brain-cortex slices, nor that of adult rat kidney medulla, nor that of an extract of an acetone-dried powder of brain. 3. Tetrodotoxin does not affect the rate of penetration of glucose into brain slices. 4. Its effect is not apparent if it is added 10min or later after the onset of anoxia. 5. Its effect diminishes as the concentration of K+ in the incubation medium is increased while that of Na+ is decreased. 6. Its salient effect, at the onset of anoxia, is to diminish influx of Na+ into, and efflux of K+ from, the brain slices. 7. Substances that promote cerebral influx of Na+, e.g. protoveratrine, sodium l-glutamate, diminish the accelerative action of tetrodotoxin. 8. It is concluded that tetrodotoxin exerts its effect on anaerobic glycolysis by suppressing, at the onset of anoxia, the generation of action potentials and thereby the accompanying influx of Na+ and efflux of K+. It is suggested that glycolytic stimulation occurs because a rate-limiting step, e.g. operation of pyruvate kinase, is stimulated by K+ and depressed by Na+. 9. Local anaesthetics behave in a manner similar to that of tetrodotoxin in enhancing cerebral anaerobic glycolysis. 10. Sodium Amytal has a marked effect at relatively high concentration. 11. Tetrodotoxin diminishes efflux of amino acids, particularly glutamate and aspartate, at the onset of anoxia.


1962 ◽  
Vol 40 (1) ◽  
pp. 1439-1448
Author(s):  
J. P. von Wartburg

Rat brain cortex slices were incubated with 5-hydroxytryptophan-3-C14. A method for determination of 5-hydroxytryptamine-C14 and 5-hydroxyindolacetic acid-C14 formed in brain slices is described. Effects of inhibitors of 5-hydroxytryptophan decarboxylase and monoamine oxidase on the metabolic pathway of 5-hydroxytryptophan-3-C14 were measured. α-Methyl dopa (0.33 mM) decreased the level of 5-hydroxyindolacetic acid to a greater amount than that of 5-hydroxytryptamine. Iproniazid (3.3 mM) resulted in an accumulation of 5-hydroxytryptamine and a decrease of 5-hydroxyindolacetic acid formation of 65%. Pheniprazine (0.1 mM) exerted an inhibitory effect on both 5-hydroxytryptophan decarboxylase and monoamine oxidase. Chlorpromazine (0.5 mM) decreased the level of 5-hydroxytryptamine 60% and had a synergistic effect with the inhibition on respiration of brain slices and 5-hydroxytryptophan transport exerted by 0.2 M n-propanol.


1972 ◽  
Vol 50 (6) ◽  
pp. 672-683 ◽  
Author(s):  
N. B. Glick ◽  
J. H. Quastel

Comparisons have been made of the rates of incorporation of [8-14C]adenine into RNA and of the specific activities of labelled ATP, in rat brain cortex slices incubated under conditions affecting nucleotide and RNA synthesis.The presence of 50 mM KCl in the incubation medium causes a considerable reduction of the rate of [8-14C]-adenine incorporation into RNA, with no diminution of the specific activity of the cerebral ATP or of the rate of nucleotide formation from adenine. It is inferred that cerebral RNA synthesis is suppressed by 50 mM KCl in the incubation medium.When K+ is omitted from the incubation medium or in the presence of 198 mM NaCl or 5 mM sodium L-glutamate, both the rate of [8-14C]adenine incorporation into RNA and the specific activity of cerebral ATP are diminished to approximately the same extent. This suggests that the process of RNA synthesis in the brain tissue is but little affected either by the increased ceil concentration of Na+ or by the diminished ATP concentration that obtain under these incubation conditions. The process, however, of [8-14C]adenine incorporation into cell nucleotides is markedly suppressed.The presence of protoveratrine (10 μM) causes at least 40% reduction in the rates of [8-14C]adenine incorporation into both RNA and nucleotides with little reduction in the specific activity of the cerebral ATP. The effects of protoveratrine are abolished by tetrodotoxin, indicating that the effects of protoveratrine are confined to the neurons.It is concluded that reductions of the specific activity of cerebral ATP derived from labelled adenine are due to the diminished rates of nucleotide formation from adenine that occur under specific incubation conditions. Such reductions may give rise to the observed diminutions in the rates of incorporation of labelled adenine into RNA. The relatively small fail in the specific activity of isolated ATP after incubation of brain tissue in the presence of protoveratrine is attributed to the localization of the effects of this drug to the neurons, in which the content and specific activity of ATP are suppressed, while those in the glia are undiminished.


1962 ◽  
Vol 40 (10) ◽  
pp. 1439-1448 ◽  
Author(s):  
J. P. von Wartburg

Rat brain cortex slices were incubated with 5-hydroxytryptophan-3-C14. A method for determination of 5-hydroxytryptamine-C14 and 5-hydroxyindolacetic acid-C14 formed in brain slices is described. Effects of inhibitors of 5-hydroxytryptophan decarboxylase and monoamine oxidase on the metabolic pathway of 5-hydroxytryptophan-3-C14 were measured. α-Methyl dopa (0.33 mM) decreased the level of 5-hydroxyindolacetic acid to a greater amount than that of 5-hydroxytryptamine. Iproniazid (3.3 mM) resulted in an accumulation of 5-hydroxytryptamine and a decrease of 5-hydroxyindolacetic acid formation of 65%. Pheniprazine (0.1 mM) exerted an inhibitory effect on both 5-hydroxytryptophan decarboxylase and monoamine oxidase. Chlorpromazine (0.5 mM) decreased the level of 5-hydroxytryptamine 60% and had a synergistic effect with the inhibition on respiration of brain slices and 5-hydroxytryptophan transport exerted by 0.2 M n-propanol.


1965 ◽  
Vol 43 (7) ◽  
pp. 1041-1051 ◽  
Author(s):  
Edward Majchrowicz

Aliphatic alcohols and the corresponding aldehydes inhibit the oxidation of glucose-U-C14to C14O2, total respiratory carbon dioxide formation, and oxygen consumption by potassium-stimulated rat brain cortex slices. The inhibitory effects of alcohols increase with the increase of the length of carbon chain, which is similar to the inhibitory effects of alcohols on the metabolism of liver slices. Forty millimolar pentanol and ethanol inhibit C14O2formation by 92% and 17% respectively. However, aliphatic alcohols at a fraction of the concentrations used with brain slices severely suppress C14O2formation, total CO2formation, and incorporation of acetate-1-C14and glucose-U-C14into hepatic lipids and proteins.At low concentrations aldehyde inhibition increases rapidly with the concentration, which is in direct contrast to ethanol or propanol whose inhibitory effects change slightly. Three millimolar propionaldehyde, butyraldehyde, and valeraldehyde are approximately 6 times more inhibitory to C14O2formation than the corresponding alcohols at 20 mM; acetaldehyde (3 mM), on the other hand, is approximately 24 times more inhibitory than 20 mM ethanol. These observations show that aldehydes affect the metabolism of brain slices in a different manner than the corresponding alcohols, which is consistent with the conclusion that there is no enzyme system present in the brain cortex slices responsible for the oxidation of alcohols to aldehydes. In contrast to aliphatic alcohols, the inhibitory effects of aldehydes do not increase with the length of aliphatic carbon chain. Of all alcohols and aldehydes tested, the inhibitions caused by acetaldehyde and valeraldehyde are most severe and approximately equal at equivalent concentrations. Three millimolar acetaldehyde and valeraldehyde suppress C14O2formation by 58% and 53% respectively. The effects of 3 mM propionaldehyde and butyraldehyde (29% and 26% respectively) are also approximately equal but smaller than those of either acetaldehyde or valeraldehyde.The observed inhibitory effects of alcohols on the metabolism of rat brain cortex slices support the suggestion that the site of ethanol inhibition is partly associated with that component of the oxidative system which is dependent on normal functioning of the active transport of sodium across the nerve cell membrane and partly due to acetaldehyde which is conveyed via the blood stream from liver to the brain. Similar deductions may apply to other aliphatic alcohols. The inhibitory effects of aldehydes are consistent with the conclusion that the inhibition depends on the properties of the aldehyde group rather than on the length of carbon chain, although their effects on ion transport across the nerve cell membrane have yet to be reported.


1961 ◽  
Vol 39 (12) ◽  
pp. 1811-1827 ◽  
Author(s):  
A. Vardanis ◽  
J. H. Quastel

The effects of tetraethyl lead, tetraethyl tin, triethyl lead, and triethyl tin on the metabolism of rat brain cortex slices have been studied. Tetraethyl lead and tetraethyl tin inhibit the active transport of amino acids into rat brain cortex slices at concentrations and under conditions that show no effect on the glucose metabolism of the slices. Tetraethyl lead and tetraethyl tin inhibit the oxidation of L-glutamate by rat brain slices. This effect can be accounted for on the basis of the inhibitory action of these two substances on the transport of the amino acid into the brain tissue.Tetraethyl lead and tetraethyl tin abolish, at low concentrations, potassium-stimulated brain slice respiration in presence of glucose, having little or no effect on unstimulated brain slice respiration. However, the respiration of rat brain cortex slices previously treated with phospholipase A is highly sensitive to tetraethyl lead.The inhibitory effects of the two tetraethyl compounds show differences from those of their triethyl derivatives indicating that the effects of the former substances are not due to admixture with, or conversion to, the latter substances.The brain slices of rats poisoned with either tetraethyl lead or tetraethyl tin are unable to effect the active transport of amino acids. The appearance of this biochemical abnormality coincides with the manifestation of neuropathological symptoms.The mode of action of tetraethyl lead and of tetraethyl tin on brain metabolism in vitro is discussed. It is suggested that they may act on phospholipid groups concerned with amino acid and cation transport at the cell membrane.


1970 ◽  
Vol 116 (4) ◽  
pp. 641-655 ◽  
Author(s):  
T. Itoh ◽  
J. H. Quastel

1. Acetoacetate or dl-β-hydroxybutyrate increases the rate of oxygen consumption to a smaller extent than that brought about by glucose or pyruvate in adult rat brain-cortex slices but to the same extent as that in infant rat brain-cortex slices. 2. The rate of 14CO2 evolution from [1-14C]glucose considerably exceeds that from [6-14C]glucose in respiring infant rat brain-cortex slices, in contrast with adult brain-cortex slices, suggesting that the hexose monophosphate shunt operates at a greater rate in the infant rat brain than in the adult rat brain. 3. The rate of 14CO2 evolution from [3-14C]acetoacetate or dl-β-hydroxy[3-14C]butyrate, in the absence of glucose, is the same in infant rat brain slices as in adult rat brain slices. It exceeds that from [2-14C]glucose in infant rat brain but is less than that from [2-14C]glucose in adult rat brain. 4. Acetoacetate is oxidized in the brain through the operation of the citric acid cycle, as shown by the accelerating effect of glucose on acetoacetate oxidation in adult brain slices, by the inhibitory effects of malonate in both infant and adult brain slices and by its conversion into glutamate and related amino acids in both tissues. 5. Acetoacetate does not affect glucose utilization in adult or infant brain slices. It inhibits the rate of 14CO2 formation from [2-14C]glucose or [U-14C]-glucose the effect not being wholly due to isotopic dilution. 6. Acetoacetate inhibits non-competitively the oxidation of [1-14C]pyruvate, the effect being attributed to competition between acetyl-CoA and CoA for the pyruvate-oxidation system. 7. Acetoacetate increases the rate of aerobic formation of lactate from glucose with both adult and infant rat brain slices. 8. The presence of 0.1mm-2,4-dinitrophenol diminishes but does not abolish the rate of 14CO2 formation from [3-14C]acetoacetate in rat brain slices. This points to the participation of ATP in the process of oxidation of acetoacetate in infant or adult rat brain. 9. The presence of 5mm-d-glutamate inhibits the rate of 14CO2 formation from [3-14C]acetoacetate, in the presence or absence of glucose. 10. Labelled amino acids are formed from [3-14C]acetoacetate in both adult and infant rat brain-cortex slices, but the amounts are smaller than those found with [2-14C]glucose in adult rat brain and greater than those found with [2-14C]glucose in infant rat brain. 11. Acetoacetate is not as effective as glucose as a precursor of acetylcholine in adult rat brain but is as effective as glucose in infant rat brain slices. 12. Acetoacetate or β-hydroxybutyrate is a more potent source of acetyl-CoA than is glucose in infant rat brain slices but is less so in adult rat brain slices.


1961 ◽  
Vol 39 (3) ◽  
pp. 551-558 ◽  
Author(s):  
P. N. Abadom ◽  
K. Ahmed ◽  
P. G. Scholefield

Tofranil inhibits the respiratory activity of rat brain cortex slices incubated in a glucose-containing medium. It also inhibits the uptake and incorporation of glycine-1-C14at concentrations which have only a slight inhibitory effect on the respiration of slices. Tofranil also inhibits oxidative phosphorylation in both rat liver and rat brain mitochondria but at higher concentrations respiration is greatly affected. Tofranil differs quantitatively from chlorpromazine in its greater inhibitory effect on the ATP–Pi32exchange reaction and its lesser effect on the cytochrome c oxidase activity of rat liver mitochondria.


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