bluuebunny/crossref_metadata_2025 huggingface dataset:
CHARACTERIZATION OF THE BASIS FOR DIFFERENCES IN SERUM DBH ACTIVITY IN IMMATURE AND ADULT RATS BY USE OF HOMOLOGOUS ANTIBODY
Authors: Grzanna R. , Nelson M. F. , Weinshilboum R. M. , Dunnette J. , Coyle J. T.
Published: October 2006

Abstract– Rat serum dopamine‐β‐hydroxylase (DBH) activity decreased 5‐7‐fold between 15 and 60 days of age. Immunoprecipitation performed with homologous antibody (guinea‐pig anti‐rat adrenal DBH) showed that during this time period the quantity of antibody necessary to precipitate 50% of the enzymatic activity (AD50) decreased 5‐fold from 0.25 to 0.05 μl/ml. The biochemical properties of rat serum DBH at 15 and 60 days of age were compared to test the hypothesis that there might be different biochemical forms of the enzyme in the blood of immature and adult rats. Thermal stability, apparent Km for tyramine, electrophoretic mobility, pH optima and elution profile on gel filtratioh chromatography were all found to be similar for rat serum DBH at both ages. On the basis of homospecific activity and multiple similarities in biochemical characteristics, it appears that differences in serum activity at the two ages reflect differences in the steady‐state levels of enzyme.

To determine the turnover of serum DBH in the two age groups, the recovery of enzyme activity was monitored after acute clearance of the circulating pool of DBH by treatment with the homologous antiserum. Immunotitration of DBH activity in vivo indicated that the total pool of serum enzyme was 4‐fold greater in the mature rat than in 4‐day‐olds. After treatment of adult rats with 2μl of homologous antiserum, serum DBH activity was reduced by 85% with a half‐life of recovery of 3.0 ± 0.6 days; the estimated fractional rate of degradation was 0.23 ± 0.06 day−1 and the rate of entrance was 2.3 ± 0.2 units/ml/day. After treatment of 4‐day‐old rats with 1 μl of homologous antiserum, serum DBH activity was reduced by 95% with a half‐life of recovery of 3.3 ± 0.5 days: the estimated average fractional rate of degradation was 0.22 ± 0.06 day−1 and the average rate of entrance was 10.7 ± 1.6 units/ml/day. Thus, the several‐fold difference in steady‐state levels of serum DBH in rat pups as compared to adult rats appears to be due to greatly increased rates of entrance of the enzyme in the immature rats.
AXONAL TRANSPORT OF PHOSPHOLIPIDS IN RAT VISUAL SYSTEM1
Authors: Toews Arrel D. , Goodrum Jeffry F. , Morell Pierre
Published: October 2006

Abstract— Seventeen day old rats were injected intraocularly with a phospholipid precursor, [32P]phosphate, and a glycoprotein precursor, [3H]fucose. Animals were killed between 1 h and 21 days later, and structures of the visual pathway (retina, optic nerve, optic tract, lateral geniculate body, and superior colliculus) were dissected. Radioactivity in phospholipids ([32P] in solvent‐extracted material) and in glycoproteins ([3H] in solvent‐extracted residue) was determined. Incorporation of [3H]fucose into retinal glycoproteins peaked at 6–8 h. Labelled glycoproteins were present in superior colliculus by 2h after injection, indicating a rapid rate of transport; maximal labelling was at 8–10 h after injection. Incorporation of [32P]phosphate into retinal phospholipids peaked at 1 day after injection. Phospholipids were also rapidly transported since label was present in the superior colliculus by 3 h after injection: however, maximal labelling did not occur until 5–6 days. These results indicate that newly synthesized phospholipids enter a preexisting pool, part of which is later committed to transport at a rapid rate.

Transported phospholipids were catabolized at the nerve endings with a maximum half‐life of several days; there was minimal recycling of precursor label.

Lipids were fractionated by thin‐layer chromatography, and radioactivity in individual phospholipid classes determined. Choline and ethanolamine phosphoglycerides were the major transported phospholipids, together accounting for approx 85% of the total transported lipid radioactivity. At early time points, the ratio of radioactivity in choline phosphoglycerides to that in ethanolamine phosphoglycerides increased in structures progressively removed from the site of synthesis (retina) but by 2 days approached a constant value. In each structure, choline phosphoglyceride‐ethanolamine phosphoglyceride radioactivity ratios decreased with time, rapidly at first, but plateaued by 2 days. These results indicate that choline phosphoglycerides are committed to transport sooner than ethanolamine phosphoglycerides.

Some experiments were also conducted using [2‐3H]glycerol as a phospholipid precursor. Results concerning incorporation of this precursor into individual phospholipid classes and their subsequent axonal transport were comparable to those obtained using [32P]phosphate, with the following exceptions: (a) incorporation of [2‐3H]glycerol into retinal phospholipids was relatively rapid (near‐maximal levels at 1 h after injection) although transport to the superior colliculus showed an extended time course very similar to [32P]‐labelled lipids; (b) [2‐3H]glycerol was somewhat less efficient than [32P]phosphate in labelling lipids committed to transport relative to labelling those which remained in the retina; and (c) [2‐3H]glycerol did not label plasmalogens.
EFFECTS OF X‐IRRADIATION INDUCED LOSS OF CEREBELLAR GRANULE CELLS ON THE SYNAPTOSOMAL LEVELS AND THE HIGH AFFINITY UPTAKE OF AMINO ACIDS1
Authors: Rohde B. H. , Rea M. A. , Simon J. R. , McBride W. J.
Published: October 2006

Abstract— Crude synaptosomal (P2) preparations were obtained from the cerebella of rats in which the granule cell population had been selectively reduced by X‐irradiation treatment and from the cerebella of control animals. In the P2 fraction from control cerebella, the level of glutamate was greater than any other of the 5 amino acids measured and was 2‐fold higher than taurine, which was present at the next highest level. The content of taurine was slightly higher than that found for aspartate and was 3‐fold greater than that observed for GABA. Alanine and glycine were present in the lowest amounts. The levels of glutamate and aspartate were significantly (P < 0.05) lower by 25 and 15%, respectively, in the P2 fraction isolated from the X‐irradiated cerebella in comparison to control values. The content of taurine, GABA, glycine, and alanine were not changed by the X‐irradiation treatment. The uptake of 1.0 μm‐l‐[3H]glutamate and l‐[3H]aspartate was reduced approx 20% by X‐irradiation treatment, whereas the uptake of 1.0 μm‐[3H]GABA and [3H]taurine was unchanged. A more detailed kinetic analysis of l‐[3H]glutamate uptake revealed there was a 20% decrease in the Vmax value with X‐irradiation treatment and no change in the apparent Km value.

In a second study, the uptake of l‐[3H]glutamate, l‐[3H]aspartate and [3H]GABA was measured using P2 fractions obtained from the cerebella of rats in which the population of granule, stellate and basket cells had been reduced by X‐irradiation treatment. The uptake of 1.0μm‐l‐[3H]glutamate, l‐[3H]aspartate and [3H]GABA was significantly (P < 0.05) reduced to 57, 68, and 59%, respectively, of control values. A more detailed kinetic analysis of [3H]GABA uptake revealed no significant change in the apparent Km and a 35% decrease in the Vmax value.

The data are discussed in terms of glutamate being the excitatory neurotransmitter released from granule cells and GABA being the inhibitory neurotransmitter released from basket cells.
STEREOSPECIFIC EFFECTS OF (‐)SULPIRIDE ON BRAIN DOPAMINE METABOLISM AND PROLACTIN RELEASE
Authors: Hofmann M. , Jommi G. C. , Montefusco O. , Tonon G. C. , Spano P. F. , Trabucchi M.
Published: October 2006

Abstract— Sulpiride, an unusual neuroleptic, was resolved into its two enantiomers by a newly developed procedure. (‐)Sulpiride is several times more active than (+)sulpiride in increasing rat prolactin secretion and DOPAC concentrations in rat strialum and limbic areas. These results are in agreement with clinical data showing that (‐)sulpiride is the active form of the drug.
IN VITRO SYNTHESIS OF THE MYELIN BASIC PROTEINS IN THE DEVELOPING MOUSE BRAIN: PROPERTIES OF A HOMOGENATE SYSTEM
Authors: Carey Gregory D. , Campagnoni Anthony T.
Published: October 2006

—An in vitro system using mouse brain homogenates has been developed to study the synthesis of the myelin basic proteins. Incorporation of [3H]leucine into protein in this system did not require additional energy sources. The system was slightly stimulated by glucose and strongly inhibited by puromycin. Myelin basic proteins were isolated from incubation mixtures by conventional techniques of solvent extraction and column chromatography, and finally separated into the large and small components by polyacrylamide gel electrophoresis in an acetic acid‐urea system. Gels were stained, sliced, dissolved, and counted, and relative rates of incorporation of label into the two basic proteins were determined at several ages. The ratio of radioactivity incorporated into the small (S) and large (L) basic proteins, over a 30 min incubation period, was found to increase from 0.97 at 10 days to 1.59 at 21 days and decline thereafter.

These data generally agree with earlier studies on the in vivo synthesis of the myelin basic proteins in mice. An interesting feature of the time course was that incorporation of [3H]leucine into the purified myelin basic proteins relative to incorporation into total protein in the homogenate increased almost 2‐fold during the course of the 30‐min incubation. This suggested that post‐translational processing of at least one of the two basic proteins was occurring. To examine this possibility further, experiments were conducted in which incorporation was allowed to proceed for 2–5 min, before being inhibited with puromycin; the incubation was then continued for up to 25 min longer. Although total incorporation was inhibited immediately after puromycin addition, label was found to continue to accumulate in the basic proteins to the extent of 30–100% above controls. These data support the notion that the MBPs are synthesized as precursors and then processed to yield authentic myelin basic proteins and that this processing can occur in vitro.
RESOLUTION OF BRAIN TROPONIN COMPLEX
Authors: Mahendran C. , Berl S.
Published: October 2006

Affinity chromatography was used to partially purify the troponin complex from crude regulatory proteins obtained from bovine brain cortex. Three components were obtained from this partially purified troponin complex by treatment with 6 M‐urea and 1 mM‐EGTA followed by chromatography on DEAE‐Sephadex‐A50. The effects of the three components on skeletal muscle actin activated MgATPase activity of muscle myosin (ATP phosphohydrolase, EC 3.6.1.3.) suggested that they were analogous to that of the skeletal muscle troponins I, C, and T. The apparent molecular weights of the brain troponin subunits (I, C, and T) were 18, 700, 14, 000 and 36, 400, respectively. The molecular weights of the former two proteins were less than those reported for the analogous skeletal muscle troponins. Thus, brain actomyosin complex may be regulated in a manner similar to that of striated muscle actomyosin.
EFFECT OF TEMPERATURE AND IONOPHORES ON THE PERMEABILITY OF SYNAPTOSOMES
Authors: CARVALHO C. A. M. , CARVALHO A. P.
Published: October 2006

Synaptosomes swell rapidly in isosmotic solutions of glycerol or urea, but the swelling in solutions of larger non‐electrolytes, such as erythritol, glucose or sucrose is slower. The permeability of synaptosomes to non‐electrolytes is temperature dependent, and the low activation energies for the permeation of urea (13 kcal/mol) and erythritol (9.5 kcal/mol) indicate that the penetration of non‐electrolytes into the synaptosomes does not imply complete dehydration of the molecules.

The relative permeability of synaptosomes to cations, as measured by the rate of swelling in isosmotic solutions of acetate salts is in the order: NH+4 > Na+ > Li+ > K+ > Ca2+. The ionophores, X‐537A and nigericin, or valinomycin + FCCP, which promote exchange of cations for H+, cause swelling of synaptosomes in solutions of potassium salts of acetate or propionate, but not in KCI, whereas H+ release is higher in KCI medium. This suggests that the organic unions cross the membrane after combining with H+ to form the respective weak acids. The relative permeability to anions is in the order: acetate ≃ propionate > Cl− > SO2‐4≃ maleate ≃ succinate.

The energies of activation for the permeability of synaptosomes to potassium acetate in the presence of X‐537A or gramicidin D are 13 kcal/mol and 7.5 kcal/mol, respectively, which reflects different mechanisms of action for the two ionophores in the membranes.
Characterization of Corticosterone‐Induced Protein Synthesis in Hippocampal Slices
Authors: Etgen Anne M. , Martin Michelle , Gilbert Russell , Lynch Gary
Published: October 2006

: Corticosterone significantly increases the incorporation of [3H]leucine into specific cytosol protein(s) isolated from in vitro hippocampal slices prepared from adult male albino rats. The present study showed that in slices coincubated with glucocorticoid plus a protein synthesis inhibitor (1 mm‐cycloheximide), no such enhancement of amino acid incorporation was observed, suggesting that the hormone acts in the hippocampus to increase de novo protein synthesis. Further experiments demonstrated that the steroid‐induced protein synthesis was first detectable (+ 5.7%) following a 30‐min exposure of slices to corticosterone; slices incubated for 1 or 2 h both showed a 12% increase in synthesis of the affected protein(s) when compared with controls. In an attempt to determine whether the glucocorticoid alteration of protein metabolism was receptor‐mediated, hippocampal slices were also incubated with 10 nm‐progesterone, a steroid known to compete for corticosterone binding to its cytosol receptor. Progesterone alone, which does not translocate cytoplasmic receptors to the nucleus, did not alter hippocampal protein metabolism and effectively blocked the induction by corticosterone of the 54K protein(s). These studies provide evidence that in the rat hippocampus corticosterone interacts with high‐affinity steroid receptors to regulate the synthesis of specific protein(s).
A Rapid and Simple Microprocedure for Myelin Isolation
Authors: Keenan Roy W. , Jones Marjorie
Published: October 2006

In order to conduct enzymatic studies on myelin from discrete regions of brain or spinal cord, we required a method capable of yielding purified myelin from tissue samples weighing 50 mg or less. Microprocedures that have been reported such as that of Detering and Wells (1976) or Fagg et al. (1978) were not suitable for rapid and routine myelin preparations. In this communication, the results of experiments in which we were able to isolate myelin from as little as 5 mg of whole brain are reported. Data are presented to show that the myelin isolated by this procedure is comparable to the myelin isolated by the macro‐scale procedure of Norton and Poduslo (1973a).
Catecholamine Concentration in the Developing Rat Brain after γ‐Hydroxybutyric Acid
Authors: Lundborg P. , Hedner T. , Engel J.
Published: October 2006

The effect of the GABA receptor agonist γ‐hydroxybutyric acid (GHBA) on brain catecholamine concentration was investigated in 1 to 28 day old rats. The infant rats were given GHBA in various doses (375–1500 mg/kg) and the effects on whole brain or regional brain concentration of dopamine (DA) and noradrenaline (NA) were measured. Brain DA concentration increased in a dose‐dependent way already from two days of postnatal age. In the regional brain study of the 14‐ and 28‐day‐old animals the increase in DA concentration was found to be almost exclusively located in the striatal region. Generally, no changes in NA concentration were found in the whole brain or various brain regions at any of the ages after GHBA. It is concluded that the inhibitory striatal‐nigral neurons, utilizing GABA as a transmitter, are functionally developed during early postnatal age.
Studies on the Mechanism of Action of Avermectin B1a: Stimulation of Release of γ‐Aminobutyric Acid from Brain Synaptosomes
Authors: Pong Sheng‐Shung , Wang Ching C. , Fritz Lawrence C.
Published: October 2006

Avermectin B1a, a novel macrocyclic lactone antiparasitic agent, causes a marked and sustained increase of γ‐aminobutyric acid release from rat brain synaptosomes. A concentration of 8‐10 μM of avermectin B1a produced the maximal effect (310 ± 30% of the control), while the half‐maximal level was achieved at 2‐3 μM. The drug also stimulated γ‐aminobutyric acid release (251 ± 11% of the basal level) from synaptosomes in calcium‐free medium, which was 28 ± 4% lower than that in the 1.8 mm‐Ca2+ medium. The compound did not, however, affect the synaptosomal release of glutamate. At the lobster neuromuscular junction, avermectin B1a reduced the input resistance of muscle fibers in control Ringer's solution as well as in Ringer's solution in which Co2+ was substituted for Ca2+. This observation is in accord with the Ca2+ independent stimulation of γ‐aminobutyric acid release seen with synaptosomes. A good correlation between antiparasitic activity and γ‐aminobutyric acid‐releasing activity has been found among various derivatives of avermectin B1a, which suggests that the ability of the drug to release this neurotransmitter may be the basis of its antiparasitic action.
The Sequential Alterations of Endoneurial Cholesterol and Fatty Acid in Wallerian Degeneration and Regeneration
Authors: Yao Jeffrey K. , Natarajan V. , Dyck Peter James
Published: October 2006

The endoneurial nonpolar lipids were serially examined throughout Wallerian degeneration and regeneration. Following nerve crush in the rat, the endoneurial content of cholesterol falls and cholesteryl ester content rises dramatically. The maximal alteration of this ratio corresponds reasonably well with events of myelin ovoid dissolution to sudanophilic amorphous lipids. Thereafter, as regenerative events overshadow degenerative events the ratio is slowly restored toward normal. The increased cholesteryl esters are probably synthesized within endoneurium from free fatty acids which become available when myelin is degraded. The endoneurial free fatty acid content presumably represents the net effect of phospholipid degradation, cholesterol esterification, cholesteryl ester hydrolysis, and fascicular entry and exit. Free fatty acids become significantly elevated by 12 days, probably reach a peak between 16 and 60 days, and thereafter return to normal with fiber regeneration. The fatty acid composition of cholesteryl esters from crushed nerves is markedly different from those of normal sciatic nerves. The altered fatty acid composition of cholesteryl esters from myelin suggests that both synthesis and hydrolysis exhibit substrate specificity toward chain length and unsaturation, with oleate being the most favored substrate.
A Radioassay for GM1 Ganglioside Concentration in Cerebrospinal Fluid
Authors: Ginns Edward , French Joseph
Published: October 2006

A radioassay for the rapid determination of GM1, ganglioside concentration in small volumes of CSF from individual patients is described. The assay utilizes the high‐affinity interaction between cholera enterotoxin and GM1 ganglioside. The lower detection limit of GM1 ganglioside by this radioassay under the described incubation conditions is 2.5 ng/ml. The radioassay‐determined lumbar CSF GM1 ganglioside concentrations in a small group of patients with diverse neurologic disorders are presented. The radioassay GM1 ganglioside concentration is in good agreement with the GM1 ganglioside concentration determined, in one patient, by the tlc‐densitometry technique.
Biosynthesis of Peripheral Nervous System Myelin Proteins In Vitro
Authors: Smith Marion Edmonds
Published: October 2006

Myelin prepared from spinal roots and sciatic nerves of young rats (24 days of age) after incubation with [3H] amino acids in glucose‐Krebs Ringer bicarbonate contained a considerable amount of radioactivity incorporated into the proteins. When these were separated on SDS‐polyacrylamide gels, peaks of radioactivity were found to correspond to the P0, 23K, P1, and P2 proteins. Although very little Fast‐Green‐stained protein was visible in the high molecular weight region of the gels (>27,000), much radioactivity was located in this area. Incorporation of [3H] amino acids into the myelin proteins continued up to 3 h and was dependent on glucose, but not on O2‐CO2 replacement of air in the flask, whereas cycloheximide inhibited incorporation by 80%. Much less amino acid was incorporated into myelin protein of old rats. High molecular weight proteins appeared to be more metabolically active than other myelin proteins, as estimated by the percent of total counts incorporated/amount of Fast‐Green staining, and their activity decreased less with age. The 23K and P° proteins incorporated [3H] amino acid in approximate proportion to their con‐tent. P0 was less, and P2 somewhat more metabolically active than their con‐tents would predict. [3H]fucose also was incorporated in vitro into P0 and P1 (probably 19K) proteins as well as into the high molecular weight proteins, while glucosamine served less well as a glycoprotein precursor. This preparation may be useful for examining further parameters of PNS myelin synthesis, assembly, and the effects of neurotoxic agents.
Thiamine and Cholinergic Transmission in the Electric Organ of Torpedo
Authors: Eder Lorenza , Dunant Yves , Loctin Françoise
Published: October 2006

Abstract: The electrogenic tissue of Torpedo was found to phosphorylate in vitro external [14C]thiamine by a saturable process. The rate of this metabolisation was increased when acetate, an efficient precursor of acetylcholine (ACh) synthesis in this tissue, was added to the incubation medium, thus increasing the turnover of ACh. Nerve stimulation did not release significant amounts of the previously accumulated [14C]thiamine. Exogenous thiamine modified the size of the electroplaque potential (e.p.p.). At concentrations higher than 10−3 M the nerve‐electroplaque transmission was depressed after a transient increase of the e.p.p. Such a depression was due to a strong decrease of the ACh release. At concentrations equal to or lower than 10−3 M, thiamine affected transmission in a rather complex fashion. ACh release was decreased or increased depending on concentration, time of application, and mode of stimulation. Oxythiamine, a structural antimetabolite of thiamine, affected the transmission in a very characteristic manner at 10−5 M and higher concentrations. The amplitude of the e.p.p. was increased and, more strikingly, its duration was prolonged. These changes were not due to an inhibition of cholinesterase activity but to an enhancement of the evoked release of ACh either on single‐impulse or repetitive stimulation. Another antimetabolite, pyrithiamine, had no effect on the transmission nor on ACh release. From this and our previous work, it is proposed that thiamine is involved, directly or indirectly, in the process of ACh release. The possible mechanisms of this involvement are discussed.
Photochemical Damage in the Albino Rat Retina: Morphological Changes and Endogenous Amino Acids
Authors: Oraedu A. C. I. , Voaden M. J. , Marshall J.
Published: October 2006

Abstract: Endogenous amino acids were measured in retinas of rats exposed for up to 48 h to fluorescent light. Typical light damage was seen in photo–receptor cells after 30 h exposure to a maximum luminance of 1544 scotopic lux; and, from this time, taurine levels were significantly reduced. In contrast, the concentrations of other amino acids increased. After 18 h exposure to light, GABA, glycine, glutamate, and aspartate levels were raised in the photo‐receptor cells, and GABA, glutamate, and glutamine levels in the inner retina. When ‘exposed’ animals were returned to their normal environment for 72 h, photoreceptor degeneration progressed and taurine concentrations were further reduced: the results suggest that the loss was from damaged photo–receptor cells. At this time the concentrations of the other amino acids measured had, in general, returned to normal
Retrograde Inhibition of Transmitter Release by ATP
Authors: Israël M. , Lesbats B. , Manaranche R. , Meunier F. M. , Frachon P.
Published: October 2006

Abstract: After labelling ACh tissue stores in Torpedo electric organ prisms with radioactive acetate, the release of ACh and ATP triggered by electrical stimulation or KCI depolarization was measured in the same perfusate samples. The luciferin‐luciferase reaction for ATP was first counted, then the radioactive content of the sample determined. Further evidence showing that ATP release resulted from postsynaptic transmitter action was that carbachol could induce the release of ATP. A dose‐response curve was obtained. Curare or α‐bungarotoxin block the release of ATP elicited by carbachol. When triggered by KCI depolarization the increased efflux of ACh and ATP returned to low levels in spite of the maintained depolarization. After two successive KCI depolarizations, it was possible to dissociate the release of both substances. The efflux of ATP was exhausted while ACh release was maintained. If the second KCI depolarization was delayed ATP release recovered, but the release kinetics of ACh and ATP were sustained. The exhaustion of endogenous ATP release or the action of exogenous ATP had little or no effect on the release of ACh triggered by KCI depolarization. On the contrary, the release of ACh induced by electrical stimulation was sensitive to the action of adenine nucleotides, and a quantitative estimation of the inhibition of ACh release by ATP and adenosine could be made. At the onset of stimulation ATP release predominated, being gradually replaced by adenosine, which can be reuptaken. This would terminate the inhibitory action of the nucleotide. Carbachol inhibits evoked ACh release, while the effect of α‐bungarotoxin was to increase spontaneous ACh release. These effects could be respectively mediated by an increased or a reduced release of ATP resulting from the postsynaptic action of ACh agonists or antagonists. However, a direct presynaptic effect of these substances is not excluded. It seems possible that the action of ATP on ACh release can be explained through its inhibition of the depolarization‐evoked Ca2+ entry.
Occurrence and Distribution of 5‐Hydroxytryptophol in the Rat
Authors: Cheifetz Sela , Warsh Jerry J.
Published: October 2006

Abstract: The distribution of the serotonin metabolites 5‐hydroxytryptophol (5‐HTOL) and 5‐hydroxyindoleacetic acid (5‐HIAA) was determined in the rat by a sensitive and specific gas chromatography‐mass spectrometric assay. 5‐HTOL occurred in all tissues assayed, with highest concentrations in small intestine (mean ± S.E.M. = 193 ± 13 mg/g), lung (78.8 ± 13.2 mg/g), and liver (64.1 ± 4.9 mg/g). Brain 5‐HTOL concentrations (9.80 ± 0.36 mg/g) were only 1% of brain 5‐HIAA levels. Conjugated 5‐HTOL accounted for a significant fraction of the total 5‐HTOL concentrations in all tissues and varied from 20% in heart to 70% in kidney. In plasma and urine, 5‐HTOL occurred almost completely in conjugated form. Except for liver, 5‐HIAA concentrations were substantially greater than 5‐HTOL in all tissues, plasma, and urine. Highest 5‐HIAA concentrations occurred in brain (787 ± 28 mg/g), lung (744 ± 52 mg/g), and small intestine (424 ± 35 mg/g). 5‐HTOL concentrations in plasma and urine were about 25% of the respective 5‐HIAA levels. It is concluded that significant biotransformation of serotonin to 5‐HTOL in the rat occurs in the intestine, liver, and lung while in brain formation of 5‐HTOL represents a minor pathway of serotonin metabolism.
The Effects of Electroshock Convulsions on Calcium Transport within Synaptic Terminals
Authors: Delgado‐Escueta A. V. , Victor S. , Davidson D.
Published: October 2006

Abstract: Calcium transport was assessed within synaptic terminals isolated from cerebral cortices of rats which experienced one maximal electroshock (ES) convulsion daily. No significant change in calcium content [(Ca1)] of synaptosomes was present after 2 consecutive days of maximal convulsions. After 4 and 6 days of maximal seizures, (Ca1) rose 20% and 37%, respectively. 15Ca2+ influx within synaptosomes in vitro increased after 6 days of ES convulsions (1.94 ± 0.4 μmol/g protein/min in ES convulsions versus 1.54 ± .03 μmol/g protein/min in controls). The higher rate of 45Ca2+ influx in convulsed animals was accounted for by elevated internal sodium [(Na1)] values. Maximal 45Ca2+ efflux decreased after ES convulsions (0.48 μmol/g protei/min in ES convulsions versus 0.8 μmol/g protei/min in controls). The slower rate of 45Ca2+ efflux after convulsions was also accounted for by elevated (Na1). Our results suggest that (Ca1) increased within synapses after in vivo ES convulsions secondary to a primary ionic event, namely, elevated (Na1).
Intermediate Filaments from Bovine, Rat, and Human CNS: Mapping Analysis of the Major Proteins
Authors: Chiu Fung‐Chow , Korey Barry , Norton William T.
Published: October 2006

Abstract: Intermediate filaments were isolated by an axon‐flotation method from bovine, rat, and human CNS. Gel electrophoresis showed four major proteins, having molecular weights of about 50,000, 70,000, 160,000, and 210,000, to be present in filaments of all three species. Small differences in molecular weights and major differences in relative distribution of the filament proteins were observed among species. In bovine and rat brain the predominant protein was the 50,000 band, but in human brain the 70,000 band was present in greatest amount. Each filament protein of the three species was studied by peptide mapping using limited proteolysis and cyanogen bromide cleavage. Within the same molecular weight group, filament proteins from different species gave similar maps with both techniques. Some degree of heterogeneity was also observed. However, filament proteins of different molecular weights of the same species gave distinctly different maps. These studies rule out the possibility that filament proteins from different molecular weight groups are related to each other by oligomerization; nor is it likely that the lower molecular weight proteins are derived from the subunit of molecular weight 210,000.
Effects of Normo‐ and Hyperbaric Pressure on an Acetylcholine‐Binding Proteoglycolipid from Rat Gastrocnemius Tissue1
Authors: Taylor Richard F. , Robertson Robert F.
Published: October 2006

Abstract: The effects of hyperbaric pressure in helium‐oxygen atmospheres on a proteoglycolipid, isolated from rat gastrocnemius tissue, with binding specificity for cholinergic ligands have been studied at six pressure levels in the ranges of 14.7 to 400 psi (1–27 atm). The binding of acetylcholine, α‐bungarotoxin, tubocurarine, decamethonium, and nicotine to the proteoglycolipid decreased with increasing pressure, the greatest binding losses occurring in the 14.7–100 psi range. The loss of [11C]acetylcholine and 125I‐labeled α‐bungarotoxin binding to the proteoglycolipid with increasing pressure was augmented in the presence of cholinergic antagonists such as nonradioactive α‐bungarotoxin, tubocurarine, and decamethonium. The presence of hexamethonium, a ganglionic cholinergic antagonist, in the reaction mixtures appeared to prevent loss of acetylcholine and aα‐bungarotoxin binding with increasing pressure. Saturation of the proteoglycolipid with acetylcholine at each pressure level indicated that the conformation of the isolated macromolecule may be distorted with increasing pressure, resulting in alterations to specific ligand‐binding sites. The significance of these results is discussed in relation to the effects of pressure on in vivo neural processes.
Extension of the Labeled Precursor Method of Measuring Neurotransmitter Kinetics to Non‐Steady‐State Conditions: Application to Striatal Dopamine Metabolism
Authors: Paden Charles M. , Young Stephen J. , MacGregor Ronald J.
Published: October 2006

Abstract: A solution of the differential equations describing the single open compartment model of neurotransmitter kinetics which does not assume a steady state is presented. In addition, a computer procedure is described for analyzing data obtained following the pulse injection of radiolabeled precursors, which eliminates the use of graphical methods in calculating fractional rate constants of synthesis and catabolism. This procedure also provides statistically valid estimates of the standard errors of the rate constants, allowing for tests of the significance of experimental effects. The ability of this approach to discriminate between changes in synthesis and catabolism under non‐steadystate conditions is demonstrated by analyzing the kinetics of striatal dopamine metabolism following (a) administration of α‐methyl‐ρ‐tyrosine and (b) lesion of the substantia nigra. The utility of this method and precautions in its use are discussed.
Measurement of the Membrane Potential of Isolated Nerve Terminals by the Lipophilic Cation [3H]Triphenylmethylphosphonium Bromide
Authors: Hansson Elisabeth , Jacobson Ingemar , Venema Richard , Sellström Åke
Published: October 2006

Abstract: The lipophilic cation [3H]triphenylrnethylphosphonium bromide ([3H]TPMP+) was investigated as a measure of the membrane potential of synaptosomes. Conditions under which [3H]TPMP+ achieved an equilibrium distribution were tested. The toxicity of TPMP has been studied relative to its inhibitory effects on [3H]y‐aminobutyric acid ([3H]GABA) transport. In some experiments the distribution of 86RbZ+ and [3H]TPMP+ was changed upon incubation in the presence of elevated levels of K+, ouabain, or KCN, or at 0°C in a way that would be expected from the membrane potential. In normal incubation conditions a membrane potential of ∼−60 mv was calculated.
Biosynthesis of the Brain‐specific 14‐3‐2 Protein in a Cell‐free System from Wheat Germ Extract Directed with Poly(A)‐containing RNA from Rat Brain
Authors: Sakimura K. , Yoshida Y. , Nabeshima Y. , Takahashi Y.
Published: October 2006

Abstract: 14‐3‐2 Protein is a neuron‐specific protein with a molecular weight of 46,000. Poly(A)‐containing RNA was prepared from free polysomes of rat whole brains by means of phenol‐chloroform extraction and oligo (dT)‐cellulose chromatography. This RNA directed the synthesis of 14‐3‐2 protein in a cell‐free, protein‐synthesizing system derived from wheat germ. 14‐3‐2 Protein was not detected in the products of endogenous incorporation and the products directed with liver poly(A)‐containing RNA. These results indicate that mRNA for 14‐3‐2 protein contains the poly(A) sequence and resides only in the brain.
Predominance of the Cytoplasmic Form of Brain Hexokinase in Cultured Astrocytes
Authors: Lusk James A. , Manthorpe C. Marston , Kao‐Jen Judy , Wilson John E.
Published: October 2006

Abstract: Astrocytes have been cultured from neonatal rat brain according to the flask culture procedure of Booher and Sensenbrenner. Approximately 80% of the hexokinase (ATP: d‐hexose 6‐phosphotransferase, EC 2.7.1.1) activity is found in the soluble fraction in homogenates of these cells, in contrast to only 20% of the total activity in the soluble fraction of whole brain homogenates. The hexokinase from the cultured astrocytes has been compared with the cytoplasmic and glucose‐6‐P‐solubilized mitochondrial enzymes from whole brain. In kinetic properties and pH‐activity relationships, the glial hexokinase was similar to the cytoplasmic enzyme but different from the mitochondrial enzyme of whole brain. Using immunohistochemical methods for detecting hexokinase localization at the electron microscopic level, most of the cells showed prominent staining of cytoplasmic areas. If the cultured astrocytes are accepted as valid models for astrocytes in situ, these results support the suggestion of Bigl and co‐workers that the predominant form of hexokinase in glial cells is the cytoplasmic enzyme.
High‐Affinity Uptake of Spermine by Slices of Rat Cerebral Cortex
Authors: Harman R. J. , Shaw G. G.
Published: October 2006

Abstract: The accumulation of the polyamine spermine into 0.1‐mm prisms of rat cerebral cortex has been investigated at both 37°C and at 4°C. Kinetic analysis of the temperature‐sensitive portion of uptake indicates two high‐aftinity saturable components together with an unsaturable component at high concentrations. The ‘very high’– affinity saturable system (Km= 3.8 nM) was temperature‐ and sodium‐dependent, and significantly reduced by metabolic inhibitors, findings that are consistent with an active transport system for spermine into brain tissue. The ‘high’– affinity saturable component (Km= 0.44 μM) was sodium‐dependent and inhibited by ouabain, but only partially susceptible to inhibition by 2,4‐dinitrophenol and sodium cyanide. The significance of these results with respect to the function of spermine in the central nervous system is discussed.
Changes in Particulate Neuraminidase Activity During Normal and Staggerer Mutant Mouse Development
Authors: Wille Wolfgang , Trenkner Ekkhart
Published: October 2006

Abstract: The activity of particulate neuraminidase (sialidase, EC 3.2.1.18) in wild‐type mice and the neurological mutant Staggerer was studied during development. Peak activity of this enzyme was observed at postnatal day 3 (P3) in three tissues of normal mice: cerebellum, cerebrum, and liver. In Staggerer, however, neuraminidase peak activity was observed at P27 in the cerebellum, whereas the activity was close to normal in Staggerer cerebrum and liver. Activities of other glycosidases in Staggerer (α‐glucosidase (pH 3.7), α‐ glucosidase (pH 6.0), N‐acetyl‐β‐hexosaminidase, β‐glucosidase, and β‐galactosidase) did not show significant variation compared with wild‐type at P27 in any of the three tissues. This indicates that the late activity peak of particulate neuraminidase activity in the Staggerer cerebellum is neuraminidase‐specific and not due to a general increase of lysosomal enzymes.
Degradation of Arachidonoyl‐Labeled Phosphatidylinositols by Brain Synaptosomes
Authors: Der Orchid M. , Sun Grace Y.
Published: October 2006

Abstract: Incubation of synaptosomes together with 1‐acyl‐2‐[14C]arachi‐donoyl‐sn‐glycerophosphoinositols (GPI) and sodium deoxycholate yielded diacylglycerols and free arachidonic acid. Diacylglycerol formation is attributed to hydrolysis by the diacyl‐GPI‐specific phospholipase C (EC 3.1.4.10), and this reaction requires sodium deoxycholate for optimal activity. The free arachidonic acid formed is attributed to hydrolysis of diacyl‐GPI by phospholipase A (EC 3.1.1.5). Free fatty acid release was observed during incubation, even in the absence of bile salts, but this process was preferentially stimulated by sodium taurocholate. The release of fatty acids was not specific for diacyl‐GPI, as similar release was obtained during incubation with other phosphoglycerides. In the presence of deoxycholate (2 mg/ml), the release of diacylglycerols was maximal at a diacyl‐GPI concentration around 1.0 mM. However, the free fatty acid release was linear with respect to the substrate at least up to 1.4 mM. The rate of diacylglycerol release from diacyl‐GPI was more rapid in the initial 30 min, whereas the free fatty acid release was linear with time up to 2 h. Under this incubation condition, calcium was found to stimulate both types of hydrolytic action, although the concentration needed to achieve this stimulation was rather high. This type of labeled precursor is potentially useful for studies of the different modes of diacyl‐GPI degradation by enzymes in brain subcellular membranes.
Peptidase Regulation of Gonadotropin‐Releasing Hormone Levels During In Vitro Incubations of the Rat Hypothalamus
Authors: Powers C. Andrew , Johnson Donald C.
Published: October 2006

Abstract: The fate of gonadotropin‐releasing hormone (GnRH) was examined by a GnRH radioimmunoassay during in vitro incubations of the rat medial basal hypothalamus (MBH). There was a progressive disappearance of exogenous GnRH during MBH incubations. The GnRH degradation could be explained by the release of peptidases from the MBH into the incubation medium. The cytoplasmic marker lactic dehydrogenase (LDH) was also released into the incubation medium. The peptide antibiotic bacitracin produced a dose‐dependent inhibition of GnRH degradation during MBH incubations; 1 mM‐bacitracin completely inhibited exogenous GnRH degradation during 4‐h incubations. Bacitracin also produced dose‐dependent increases in the recovery of endogenous GnRH released from the MBH under basal conditions or when stimulated with the depolarizing agent veratrine. Veratrine also was found to decrease the GnRH peptidase activity significantly but not the LDH activity during MBH incubations. The present results indicate that peptidase activity can be an important regulator of endogenous GnRH released from the hypothalamus during in vitro incubations.
Reversal of Axonal Transport: Similarity of Proteins Transported in Anterograde and Retrograde Directions
Authors: Bisby M. A.
Published: October 2006

Abstract: Reversal of axonal transport of endogenous labeled protein was studied in intact and injured nerve axons. Nerve crushes were used to collect labeled protein transported in anterograde and retrograde directions in rat sciatic nerve motoneuron axons after administration of L‐[35S]methionine to the vicinity of the cell bodies. The collected proteins were characterized by sodium dodecyl sulfate polyacrylamide gel electrophoresis and subsequent fluorography. In injured nerves, where the nerves were ligated distally at the time of precursor injection, the polypeptide composition of proteins moving in anterograde and retrograde directions, 9–11 h after precursor injection, was identical, indicating that reversal at a ligature is a nonselective process. In intact nerves, protein moving in the anterograde direction 22–24 h after injection was different from that found 9–11 h after injection, and was also different from protein moving in the retrograde direction 22–24 h after injection. However, protein moving in the retrograde direction 22–24 h after injection was similar to protein moving in the anterograde direction 9–11 h after injection. Thus it appears that the same group of proteins originally transported into the axon are later returned toward the cell body. In intact axons, also, reversal was nonselective, except that one major labeled polypeptide was reduced in amount in the protein moving in the retrograde direction.
Specificities of Human and Rat Brain Enzymes of Cholesterol Ester Metabolism Toward Very Long Chain Fatty Acids: Implication for Biochemical Pathogenesis of Adrenoleukodystrophy
Authors: Ogino Tadashi , Suzuki Kunihiko
Published: October 2006

Abstract: Specificities of the cholesterol‐esterifying enzyme and the three cholesterol esterases in rat brain with respect to the chain length of fatty acids were examined. For each of the hydrolases, activities toward cholesteryl lignocerate and cerotate were generally less than 1% of that toward cholesteryl oleate. However, both lignoceric and cerotic acids were esterified at rates approximately 10% of that for oleic acid. In postmortem human control and adrenoleukodystrophy brains, the esterifying activity toward cerotic acid was on the average 25% of that toward oleic acid. The abnormal accumulation of cholesterol esters with very long chain fatty acids observed in adrenoleukodystrophy can therefore occur in the absence of deficient activities of the cholesterol esterases, if the free fatty acid pool of the brain contains an abnormal amount of very long chain fatty acids.
Characterization of Alkylgalactolipids from Calf Brain by High Performance Liquid Chromatography
Authors: Yahara Shoji , Kishimoto Yasuo
Published: October 2006

Minor nonpolar galactolipids were isolated from the total lipids of calf brain stem by column chromatography and were separated by preparative thin‐layer chromatography into four groups. The material recovered from the bottom band of the thin‐layer chromatography consisted of monogalactosyl diglyceride and its 1‐0‐alkyl isomer, alkylgalactolipid, present in a molar ratio of 11 :9. After perbenzoylation. they were separated by preparative thin‐layer chromatography and characterized. The fatty acid compositions of these lipids were similar to each other and to those of the ester‐linked fatty acids of cerebroside esters. The major alkyl group of alkylgalactolipid was palmityl, and the other, minor components were oleyl. myristyl, and stearyl ethers. Perbenzoylated derivatives of these lipids were further separated by reverse‐phase high performance liquid chromatography. The chromatograms from these two lipids were similar; however, most of the peaks were still mixtures of homologs containing different fatty acids or an alkyl group.
β‐ Alanine, a Possible Neurotransmitter in the Visual System?
Authors: Sandberg Mats , Jacobson Ingemar
Published: October 2006

Abstract— The chemically evoked efflux of endogenous amino acids from perfused rabbit superior colliculus (SC) slices was studied. Amino acids in the perfusates were determined fluorimetrically with a precolumn derivatisation method and subsequent separation on an HPLC column. Potassium‐induced depolarisation caused a calcium‐dependent release of β‐alanine, GABA, glutamate, and aspartate. Veratridine‐evoked efflux was essentially similar in selectivity and could be blocked by tetrodotoxin (TTX). The results are indicative of a neurotransmitter role of β‐alanine, GABA, glutamate, and aspartate in the SC of the rabbit.
Characterization of [3H]Clonidine Binding to Two Sites in Calf Brain Membranes
Authors: Braunwalder A. , Stone G. , Lovell R. A.
Published: October 2006

Abstract: Kinetic studies showed that under appropriate conditions, [3H]clonidine binds to two distinct receptor sites in calf cortex membranes. At 23°C, binding was obtained at a low‐affinity site (dissociation constant, KD= 5.4 nm) and a high‐affinity site (KD= 1.1 nm). In contrast, at 0°C, selective binding occurred to the low‐affinity site only. Consequently, at 0°C, it was possible to evaluate the interaction of drugs with the low‐affinity receptor directly. On the other hand, competition with the high‐affinity receptor could be ascertained by generating displacement curves representing the differential between specific binding values obtained at 23 and 0°C. Guanine nucleotides selectively decreased binding to the high‐affinity site without apparent influence on the low‐affinity [3H]clonidine binding. The activities of various pharmacological agents at the low‐ and high‐affinity clonidine receptors are discussed and compared with WB‐4101 binding data.
Immunofluorescence and Histochemical Methods for Neural M1 Pyruvate Kinase Localization
Authors: Gali P. , Hauw J. J. , Boutry J. M. , Hartmann L.
Published: October 2006

Abstract: The distribution of pyruvate kinase (ATP pyruvate phosphotransferase, EC 2.7.1.40) in the nervous system has been studied by both immunofluorescence and a histochemical procedure using nitro blue tetrazolium. The localization in various parts of rat central nervous system in situ, cerebellar and cerebral cortex, was compared to that found in vitro in cultures of cerebellum, spinal ganglia, cerebral astrocytes, and skin fibroblasts. (1) Pyruvate kinase was found predominantly in the cytoplasm of neuronal cell bodies. (2) Large neurons were better visualized than small ones. (3) No glial localization was clearly demonstrated in situ, although this does not rule out the presence of some M1 pyruvate kinase. (4) Regions expected to be rich in nerve terminals, such as the cerebellar glomeruli or the cerebellar molecular layer, showed intense staining even when the cell bodies themselves were negative. This was expected, owing to the previous demonstration of the presence of M1 pyruvate kinase in nerve ending by subcellular fractionation methods. (5) The localization was similar in situ and in tissue culture, except that nerve processes were better seen in the latter and astrocytes were sometimes stained in vitro. (6) Variation in intensity of staining was observed in similar cell types in the same section or in the same culture. This could represent different metabolic or functional or maturational states.