bluuebunny/crossref_metadata_2025 huggingface dataset:
Peripheral Nerve Phospholipid Composition: Development in Normal Nerve and Age‐Dependent Changes in Wallerian Degenerated Nerve
Authors: Hofteig John H. , Vo Phung N. , Yates Allan J. , Leon Karen S.
Published: October 2006

Abstract: The phospholipid composition of normal peripheral nerve as a function of developmental age as well as that of Wallerian‐degenerated nerve as a function of age at nerve transection and duration of Wallerian degeneration have been quantitated in rabbit sciatic nerve. During development, increases in the proportions of ethanolamine plasmalogen, sphingomyelin, and combined phosphatidyl serine plus phosphatidyl inositol and decreases in the proportions of phosphatidyl choline and phosphatidyl ethanolamine correlated well with the concurrent myelin accretion. During Wallerian degeneration, age‐dependent changes in phospholipid composition were observed. The large and statistically significant increase in the proportion of phosphatidyl choline and decrease in the proportion of ethanolamine plasmalogen were manifest promptly in nerves transected at 2 weeks of age but in a delayed manner in nerves transected at 8, 12, and 20 weeks of age. The rate of loss of individual phospholipids was greater in nerves transected at younger ages. The findings from normal developing peripheral nerve may well serve as baseline data for subsequent studies of phospholipid composition in pathological peripheral nerve. The Findings from Wallerian‐degenerated peripheral nerve provide additional evidence for age‐dependent chemical changes occurring in Wallerian‐degenerated peripheral nerve that may be of significance in explaining the superior functional recovery from peripheral nerve injury observed in younger compared with older subjects.
Evidence for Glucocorticoid Target Cells in the Rat Optic Nerve. Physicochemical Characterization of Cytosol Binding Sites
Authors: Meyer Jerrold S. , McEwen Bruce S.
Published: October 2006

Abstract: Cytosolic dexamethasone (DEX) binding sites were studied in the Wallerian‐degenerating rat optic nerve (ON), a tissue that is rich in neuroglial cells but devoid of neuronal perikarya and processes. For comparison, hippocampal (HI) and anterior pituitary (AP) cytosols were studied in parallel. Binding sites in these three tissues were found to be quite similar in almost all respects. The sites have a high affinity for DEX (KD= 2.5–3.5 nM), are present at a high concentration (Bmax= 360–365 fmol/mg cytosol protein), and possess a binding specificity typical of glucocorticoid receptors in other organs. Most experiments supported the assumption of a single DEX‐binding species in each tissue. Saturation analyses consistently yielded linear Scatchard plots over the range of DEX concentrations tested. Density gradient centrifugation in each case revealed a single peak with a sedimentation coefficient of 7–8S at low ionic strength and 4–4.5S in the presence of 0.3 M KCl. Isoelectric focusing similarly localized most of the binding in each cytosol to a single large peak with an isoelectric point of approximately 6.0. Dissociation rate determinations, on the other hand, suggested the possibility of two different binding sites in each tissue. These studies show that glucocorticoid binders present in cells of the ON possess the same characteristics as the cytoplasmic receptors found in HI, AP, and other recognized glucocorticoid target tissues.
Schwann Cell Surface Proteins and Glycoproteins
Authors: Pleasure David , Hardy Mattie , Kreider Barbara , Stern Janet , Doan Hung , Shuman Sandra , Brown Spencer
Published: October 2006

Abstract: To identify surface sialoglycoproteins of rat Schwann cells and to compare molecular weights of these sialoglycoproteins with those present in rat peripheral nervous system myelin, we prepared Schwann cells from sciatic nerves of 1–3‐day‐old rats and cultured them in monolayer. Surface sialoglycoproteins of the cultured cells were tritium‐labeled by the periodateborohydride procedure and compared with sialoglycoproteins of adult rat peripheral nervous system myelin by fluorography following polyacrylamide slab gel electrophoresis in sodium dodecyl sulfate. Three radioactive bands with apparent molecular weights of 114,000–132,000, 105,000–115,000, and 44,000–56,000 were observed in both the Schwann cell and myelin preparations. Bands of similar apparent molecular weights were noted in Schwann cells metabolically radiolabeled with d‐[1,6‐3H]glucosamine. A band co‐migrating with myelin P0 glycoprotein was the most intensely radiolabeled of all peptides in periodate‐B3H4−treated myelin, but was present in only trace amounts in periodate‐B3H4− or d‐[1,6‐3H]glucosamine radiolabeled Schwann cells. Many presumably non‐myelin glycoproteins were identified in the cultured Schwann cells by the periodate‐borohydride procedure and by incubation of the cells with d‐[1,6‐3H]glucosamine. An immunoprecipitation technique was used to detect radiolabeled peptides in a nonionic detergent extract of freshly prepared, surface‐radioiodinated Schwann cells that were bound by a rabbit anti‐Schwann cell serum preabsorbed with rat fibroblasts. Many radioactive peptides were detected in the immunoprecipitate, but the two most intensely radiolabeled had apparent molecular weights of 105,000–115,000 and 95,000–106,000. This study has identified a number of glycoproteins synthesized by cultured rat Schwann cells which resemble in apparent molecular weight the glycoproteins expressed in rat peripheral nervous system myelin and has defined Schwann cell surface proteins recognized by a specific anti‐rat Schwann cell antiserum.
Synaptosomal Calcium Uptake Systems: Prostaglandins Are Probably Not Involved in the Regulation of Calcium Fluxes Into and Within the Nerve Endings
Authors: Denzlinger Claudio , Hertting Georg , Jackisch Rolf
Published: October 2006

Abstract: 45Ca2+ uptake measurements were performed on intact and osmotically lysed synaptosomes from rat brain to study the possible influence of prostaglandins (PGs) on Ca2+ movements into and within the nerve endings. The K+‐induced 45Ca2+ uptake of intact synaptosomes was not influenced by several inhibitors of PG synthesis. 45Ca2+ uptake in lysed synaptosomal preparations was promoted by ATP and seemed to be largely attributable to mitochondria, as it was inhibited by mitochondrial poisons. This Ca2+ uptake was strongly reduced by PG synthesis inhibitors but also by PG precursor fatty acids. Both PG synthesis inhibitors and precursors, according to their relative efficacy in blocking Ca2+ uptake, were able to induce Ca2+ efflux from preloaded intrasynaptosomal organelles. The PGs E2, F2α, D2, and thromboxane B2 were without effect on 45Ca2+ uptake in lysed synaptosomal preparations. On the basis of our results it does not seem likely that PGs influence Ca2+ availability by modulating Ca2+ fluxes into or within the nerve endings. The observed inhibitory effects of PG synthesis inhibitors and precursors on the intrasynaptosomal Ca2+ uptake might be due to unspecific impairment of mitochondrial functions.
αγ‐Enolase in the Rat: Ontogeny and Tissue Distribution
Authors: Jørgensen Ole Steen , Centervall Gyda
Published: October 2006

Abstract: The rat brain enolases are dimers composed of α and γ subunits. At pH 8.6 αγ‐enolase seemed to be stable, and no evidence was found for the possible formation of αγ‐enolase from αα‐enolase and γγ‐enolase in the course of rat brain homogenization. During ontogeny of the rat forebrain, αγ‐enolase was formed before γγ‐enolase. The half‐maximal specific concentrations were reached at postnatal days 14 and 23, respectively. The distribution of αγ‐ and γγ‐enolase in various rat brain areas was also investigated. In all areas both forms were present. In neuroendocrine tissues αγ‐enolase was present at a much higher concentration than γγ‐enolase. The ratio between γγ‐enolase and αγ‐enolase may be indicative of the degree of neuronal maturation, a conclusion further substantiated by the high ratio observed in cerebellum and the low ratio observed in olfactory bulbs, both compared with the ratio in forebrain.
Biochemical and Electrophysiological Demonstrations of the Actions of β‐Bungarotoxin on Synapses in Brain
Authors: Halliwell J. V. , Tse C. K. , Spokes J. W. , Othman I. , Dolly J. O.
Published: October 2006

Abstract: Homogeneous β‐bungarotoxin interacts irreversibly with rat olfactory cortex and produces permanent inhibition of neurotransmission (half‐time of blockade for 230 nM toxin is 25 min). Binding occurs in the absence of divalent cations, but the rate of synaptic blockade is increased by Ca2+, which activates the intrinsic phospholipase A2 activity of the toxin. Other observable actions of the toxin, seen with rat cerebrocortical synaptosomes, are an increase in the release of acetylcholine, glutamate and γ‐aminobutyrate and impairment of transmitter uptake, which are all insensitive to tetrodotoxin. Inactivation of the toxin's phospholipase activity by chemical modification with p‐bromophenacyl bromide diminishes the observed concomitant efflux of the neurotransmitters and lactate dehydrogenase. Collectively, the results support the idea that the toxin binds specifically and irreversibly to component(s) on nerve terminals and this together with the resultant phospholipolysis leads eventually to synaptic blockade. Such a proposal would account for the unique toxicity of the protein relative to phospholipase A2 enzymes.
An Evaluation of γ‐Glutamyl Dipeptide Derivatives as Antagonists of Amino Acid‐Induced Na+ Fluxes in Rat Striatum Slices
Authors: Tal N. , Goldberg O. , Luini A. , Teichberg V. I.
Published: October 2006

Abstract: Various γ‐glutamyl dipeptide derivatives of the l and d configuration were tested as antagonists of excitatory amino acid‐induced [22Na+] efflux from rat striatum slices. The N‐methyl‐d‐aspartate‐ and kainic acid‐induced responses were much more affected by the presence of these peptides than were the l‐glutamate‐, l‐aspartate‐, and quisqualate‐induced responses. The dipeptides produced different patterns of antagonism of the responses to N‐methyl‐d‐aspartate and kainic acid, indicating that these latter excitants interact with distinct receptors.
In Vivo Changes in γ‐Aminobutyric Acid Output from the Cerebral Cortex Induced by Inhibitors of γ‐Aminobutyric Acid Uptake and Metabolism
Authors: Moroni F. , Mulas A. , Moneti G. , Pepeu G.
Published: October 2006

Abstract: The effects of inhibitors of γ‐aminobutyric acid (GABA) metabolism or uptake on GABA output from the cerebral cortex was studied by means of a collecting cup placed on the exposed cortex of rats anaesthetized with urethane. GABA was identified and quantified by a mass‐fragmentographic method. Ethanolamine‐O‐sulphate (10−2M) applied directly on the cerebral cortex caused a long‐lasting twofold increase in GABA output, whereas dl‐2, 4‐diaminobutyric acid (5 × 10−3M) caused a sevenfold increase and β‐alanine was inactive. The results indicate that glial uptake has little effect on GABA inactivation in the cerebral cortex. The inhibition of neuronal uptake seems a more effective tool to increase GABA concentration in the synaptic cleft, and consequently also in GABA output, than the inhibition of GABA metabolism.
Ontogenetic Development of the Striatal [3H]Spiperone Binding: Regulation by Sodium and Guanine Nucleotide in Rats
Authors: Nomura Yasuyuki , Oki Keiko , Segawa Tomio
Published: October 2006

Abstract: Ontogenetic development of specific [3H]spiperone binding to crude synaptic membranes and its regulation by Na+ and GTP was investigated in the rat striatum, (d)‐Butaclamol more effectively inhibited [3H]spiperone binding than (l)‐butaclamol. The ratio of inhibitory activity of (d)‐ and (l)‐butaclamol for [3H]spiperone binding was not different between 1‐, 7‐, and 70‐day‐old animals but eight‐ to ninefold lower at 18 days of gestation than during the postnatal period. A Scatchard plot of specific binding indicated the presence of two types of binding: low‐affinity (KD=1.51 nm) and high‐affinity (KD= 0.09 nM) binding on day 70. Only one component (KD= 0.075 nM) was observed on days 1 and 7 and both types of binding were found on day 15. Bmax gradually increased with age and reached a peak on day 30, followed by a decline on days 70 and 360. Na+, 100 mM, significantly increased specific binding on days 1, 7, 15, and 70. GTP, 50 μM, completely reversed the Na+‐induced decrease in IC50 of apomor‐phine on both days 15 and 70, but not on day 7. It is suggested that receptors could recognize ligand stereospecificity on day 1. The density in dopamine receptors in the striatum reaches a peak on day 30, followed by a decrease on days 70 and 360. In addition, regulation by Na+ and GTP in agonist binding to dopamine receptors seems to become functional between 1 and 2 weeks after birth.
Poly(Adenylate) Polymerase Activity of Rat Brain
Authors: Schumm D. E. , Richter R.
Published: October 2006

Abstract: The poly(adenylate)[poly(A)] polymerase of rat brain, as in rat liver, is located primarily in the nuclear sap when nuclei are prepared under hypertonic conditions. The enzyme can be released from nuclei in two forms. Form I is prepared by gentle incubation of nuclei at 0°C in hypotonic buffer. It has a Mn optimum of 0.6 mM and a pH optimum between 8 and 9. The ATP concentration curve plateaus at 0.2 mM. The optimal poly(A) primer concentration is 600 μg/ml, which is three times higher than that for the enzyme similarly prepared from liver. The time course of the reaction for the form I enzyme is increasing over the first 40 min and becomes nearly linear thereafter. Form I is not stimulated by either calcium or cyclic nucleotides, but is inhibited by polyamines, pyrophosphate, and high concentrations of GTP. Form II enzyme is prepared by homogenization of nuclei in hypotonic buffer. It has the same ATP and poly(A) optima as the form I enzyme but displays linear kinetics over a 60‐min time course. It is slightly stimulated by cGMP and cAMP and strongly inhibited by spermine, sodium pyrophosphate, and high concentrations of GTP.
The Effect of Visual Deprivation on β‐Adrenergic Receptors in the Visual Centres of the Rat Brain
Authors: Schliebs Reinhard , Burgoyne Robert D. , Bigl Volker
Published: October 2006

Abstract: The levels of binding of [3H]dihydroalprenolol to β‐adrenergic receptors in the visual centres and frontal cortex from brains of control, dark‐reared and monocularly deprived rats were compared. Receptor binding is changed in monocularly deprived rats in the lateral geniculate nuclei and superior colliculi of both sides. Scatchard analyses indicated that the changes in the [3H]dihydroalprenolol binding in the lateral geniculate nuclei were due to alterations in both receptor affinity and receptor number. No effect of dark‐rearing could be detected.
Acetylcholinesterase Distribution in Axotomized Frog Motoneurons
Authors: Sinicropi D. V. , Michels K. , Mcllwain D. L.
Published: October 2006

Abstract: The distribution of acetylcholinesterase (AChE; EC 3.1.1.7) activity was examined in the perikarya and proximal axonal stumps of frog motoneurons injured by ventral root transection. Based upon measurements of net AChE accumulation in the proximal stumps of transected ventral roots, and upon orthograde clearances of AChE reported by others, it was determined that an amount of AChE equivalent to at least 0.7–2 times the perikaryal content of this enzyme enters the motor axon each day. A progressive decrease in the rate of AChE accumulation in transected axons during the first 3 days after ventral rhizotomy raised the possibility that excess enzyme might accumulate elsewhere within the axotomized motoneurons. However, AChE accumulation was detected only near the cut ends of the ventral roots and was not appreciably increased within injured motoneuronal cell bodies and proximal dendrites, which were isolated by a new method combining bulk and single‐cell isolation techniques. These data suggest that AChE turnover is altered rapidly in response to axonal injury, thereby avoiding large perikaryal accumulations of this enzyme.
H‐Pro‐[3H]Leu‐Gly‐NH2: Plasma Profile and Brain Uptake Following Subcutaneous Injection in the Rat
Authors: Verhoef J. , Prins A. , Veldhuis H. D. , Witter A.
Published: October 2006

Abstract: Following subcutaneous injection of the tripeptide H‐Pro‐[3H]Leu‐Gly‐NH2 ([3H]PLG) in rats, the profile of intact peptide and its radioactively labeled metabolites was examined both in plasma and in brain tissue. [3H]PLG and metabolites were determined in trichloroacetic acid extracts by reverse‐phase paired‐ion HPLC. Maximal plasma levels of unmetab‐olized PLG were reached 6–8 min after administration, after which they decreased with an elimination half‐life of 20 min. The uptake of [3H]PLG in the brain ranged from 0.0013% to 0.0017% of the administered dose per g tissue at 5–30 min following subcutaneous injection. After comparing these results with our previous findings with intravenous injection of [3H]PLG, it seemed likely that the subcutaneous route of administration might be more effective in eliciting CNS effects of PLG than the intravenous route of administration. The metabolite profiles in plasma and brain point to an initial cleavage of PLG at the NH2‐terminal side and a very rapid degradation of the peptide intermediate H‐Leu‐Gly‐NH2.
Ethylenediamine as a Specific Releasing Agent of γ‐Aminobutyric Acid in Rat Striatal Slices
Authors: Lloyd H. G. E. , Perkins M. N. , Stone T. W.
Published: October 2006

Abstract: Following incubation with [14C]y‐aminobutyric acid (GABA) or [3H]dopamine, slices of rat striatum were superfused with media containing 36 mM K+ or ethylenediamine (EDA), 1 or 5 mM. Both K+ and EDA induced a release of [14C]GABA, the K+‐induced release being largely Ca2+‐dependent, while the EDA‐induced release was not. Whereas K+ also evoked a Ca2+‐dependent release of [3H]dopamine, EDA evoked no release of dopamine. EDA may therefore have potential as a specific GABA releasing agent.
Aminergic Receptors in Drosophila melanogaster: Properties of [3H]Dihydroergocryptine Binding Sites
Authors: Dudai Yadin , Zvi Shoshana
Published: October 2006

Abstract: [3H]Dihydroergocryptine ([3H]DHE) binds to a particulate preparation from Drosophila melanogaster heads at a level of 2.4 ± 0.4 pmol/mg protein, with an apparent dissociation constant of 2.0 ± 0.5 nM. The binding sites are inactivated by heat, pronase treatment, detergents, and sulfhydryl and disulfide reagents. [3H]DHE binding is inhibited by low concentrations of serotonergic and α‐adrenergic ligands. The specificity of the binding sites, as revealed by displacement studies, differs from that of [3H]DHE binding sites in various vertebrate tissues. The [3H]DHE binding sites may correspond to serotonergic receptors, and possibly, to additional classes of receptors for putative neurotransmitters in Drosophila.
Methionine Sulfoximine In Vivo Modifies the tRNALys Pool of Developing Rat Brain
Authors: Sellinger Otto Z.
Published: October 2006

Abstract: tRNA was extracted from brains of 3‐, 8‐, and 18‐day‐old rats that were injected intracerebrally, 45 min before death, with [3H]methyl methionine or [8‐3H]guanosine, and intraperitoneally, 3 h before death, with l‐methionine‐dl‐sulfoximine (MSO), a methylation‐activating convulsant agent. Although there was no effect of age or of MSO on the per gram yield of tRNA, its specific radioactivity (dpm/A260) was highest at 3 days in both the control and the MSO groups. Age‐ and MSO‐related changes in the tRNALys content of the brain tRNA pool were investigated by means of benzoylated DEAE‐ cellulose (BDC) and reverse‐phase chromatography (RPC). BDC chromatography revealed tRNALys species in the brains of the MSO‐treated animals that were absent in control brains. Of particular interest was the finding that differences in RPC‐5 chromatographic mobility between control and MSO‐tRNALys species were abolished by conversion to lysyl‐tRNA, suggesting that the MSO‐elicited change(s) in tRNALLys structure involved the binding site(s) for lysine. Two additional findings were made: (a) lysine acceptance by the [3H]methyl‐labeled tRNALys purified from brains of the MSO‐treated animals was higher than that of controls at 18 days; and (b) omission of the BDC chromatographic step accentuated the differences in mobility on RPC‐5 columns between tRNALys species of control and MSO‐treated brains. Lastly, we found that some tRNALys species present in the MSO‐treated brains contained significantly different proportions of N2‐methyl guanine and 1‐methyl adenine, relative to controls. These MSO‐elicited changes in the methyl base content of tRNALys of immature rat brain are the first evidence of an alteration of brain tRNA structure by a centrally acting excitatory agent.
Estrogen Modulates Neural Control of Muscle Glucose 6‐Phosphate Dehydrogenase
Authors: Schaerf Frederick W. , Patz Thomas , Max Stephen R.
Published: October 2006

Abstract: Exogenously administered estrogens can enhance the rate of increase of glucose 6‐phosphate dehydrogenase activity in rat extensor digitorum longus muscles following denervation. In this communication, we report that the effect of denervation on glucose 6‐phosphate dehydrogenase activity is modified by variations in endogenous estradiol during the estrous cycle.
Measurement of Cerebral Glucose Utilization Using Washout After Carotid Injection in the Rat
Authors: Oldendorf William H. , Pardridge William M. , Braun Leon D. , Crane Paul D.
Published: October 2006

Abstract: The carotid injection technique, used previously to quantitate the kinetics of blood‐brain barrier transport of metabolic substrates, may be modified to analyze the rate of cerebral glucose utilization. A 0.2‐ml solution of [14C]glucose (GF) and [3H]methylglucose (M), an internal reference, is rapidly injected into the carotid artery, followed by microwave fixation of brain at various times up to 4 min after injection. The brain radioactivity is separated into a fraction containing neutral hexoses (GF and M) and a fraction containing metabolites of glucose. The GF/M ratio is related to the rate constant (k3) of brain glucose utilization by the simple, linear equation: In(GF/M) = In(GF°/M°) –k3t, where GF°/M°= the brain uptake index of glucose, relative to methylglucose, at 5‐15 s after injection, and t= the time after carotid injection, e.g., 1–4 min. It is assumed that (a) the rate of influx due to recirculation of label is minimal during the 4‐min circulation period; and (b) the rate constants of glucose efflux (k2) and methylglucose efflux (k2*) are identical. Independent estimates of k2 and k2* showed these parameters to be identical: k2= 0.14 + 0.08 min‐I; k2*= 0.14 ± 0.02 min‐I. A logarithmic plot of GF/M ratios versus time was linear (r = 0.99), and was described by the slope k2= 0.21 ± 0.02 min−1. Assuming glucose is uniformly distributed in brain, then the glycolytic rate = k3× brain glucose = (0.21 min−1) (2.6 μmol g−1) = 0.55 μmol min−1 g−1 for the cortex of the barbiturate‐anesthetized rat. These studies provide the basis for a simple method of measurement of regional brain glycolysis that does not require either the use of correction factors, e.g., the lumped constant, or the use of differentially labeled glucose.
Characterization of Infant Rat Cerebral Cortical Membrane Proteins Phosphorylated In Vivo: Identification of the ACTH‐Sensitive Phosphoprotein B‐50
Authors: Oestreicher A. B. , Zwiers H. , Gispen W. H. , Roberts S.
Published: October 2006

Abstract: This study on the phosphorylation in vivo of membrane proteins in cerebral cortices of infant rats reports the identification of the adrenocorticotropin (ACTH)‐sensitive phosphoprotein B‐50 as one of the substrate proteins that are rapidly phosphorylated in vivo following intracisternal administration of 2 mCi [32P]orthophosphate. Rats were sacrificed 30 min after isotope injection. A fraction enriched in membranes, designated neural membranes (NM), was isolated from the cerebral cortices according to the procedure used for preparation of synaptic plasma membranes (SPM) from adult brain. This NM fraction was characterized by electron microscopy. The proteins of NM were analyzed by sodium dodecyl sulfate‐polyacrylamide gel electrophoresis and autoradiography. Numerous protein bands of NM in infant rat brain were phosphorylated in vivo. Attention was focussed on the 32P‐labeled protein bands in the molecular weight range of 47K‐67K. In this region one phosphoprotein band (MW 48K) was more highly labeled than the other bands. The electrophoretic behavior of three of these labeled bands, designated a, c, and e (MW 48K, 55K, and 62K, respectively) was compared with that of protein bands that were phosphorylated in vitro in cerebral membranes isolated from noninjected infant rats. The effects of ACTH1‐24 and cyclic AMP in the in vitro system were also studied to probe for the presence of specific membrane proteins known to be sensitive to these modulators. On incubation of NM with [γ‐32P]ATP in the presence and absence of ACTH1‐24in vitro, phosphorylation of a 48K protein band was inhibited in a dose‐dependent fashion by the neuropeptide. Two‐dimensional electrophoretic separation of NM proteins labeled in vivo indicated that the 48K band had an isoelectric point of 4.5, identical to that of the ACTH‐sensitive B‐50 protein previously identified. Cyclic AMP stimulated phosphorylation in vitro of two protein bands (MW 55K and 59K) in NM preparations. This result indicates that the in vivo labeled band c may correspond to the cyclic AMP‐sensitive 55K protein, whereas phosphoprotein band e, labeled in vivo, appears to be different from the cyclic AMP‐sensitive 59K protein band. These observations indicate that neural membranes isolated from infant rat cerebral cortices contain a variety of proteins that can be phosphorylated in vivo. Several of these, for example, the 48K protein band, have the properties of synaptic plasma membrane proteins of adult rat brain that have been characterized by their sensitivity to neuromodulators in endogenous phosphorylating systems in vitro.
Lysoplasmalogenase–A Microsomal Enzyme from Rat Brain
Authors: Gunawan Johannes , Debuch Hildegard
Published: October 2006

Abstract: An enzymic activity of rat brain that liberates radioactive free aldehydes from 1‐[1‐14C]alk‐1′‐enyl‐sn‐glycero‐3‐phosphoethanolamine (lysoplasmalogen) is described. It was present mainly in microsomal fractions (crude) of brains of rats of different ages. The highest specific enzyme activity was found in 21‐day‐old animals. The formation of free aldehyde was dependent on the amount of enzyme protein as well as the amount of substrate added, and was linear to the incubation time up to 60 min. The pH optimum was between 7.1 and 7.3. Bivalent cations (Mg2+, Ca2+) and detergents inhibited the reaction. However, the same cell fractions as well as extracts of acetone‐dried powder of brain from young or old rats possessed no enzyme activity for liberating the aldehyde from the acylated substrates: 1‐[1‐14C]alk‐1′‐enyl‐2‐acyl‐sn‐glycero‐3‐phosphoethanolamine (plasmalogen) or plasmalogen of ox corpus callosum.
Serine Transhydroxymethylase Activity in Vertebrate Retina
Authors: Dasgupta P. , Narayanaswami A.
Published: October 2006

Abstract: The presence of serine transhydroxymethylase (STHM; EC 2.1.2.1) in crude homogenates of rat, guinea pig and goat retina has been demonstrated. There was a variation in the levels of retinal STHM activity in the three species. Substantial STHM activity was found in all the three retinal preparations in the absence of pyridoxal‐5‐phosphate in the incubation medium. There was direct correlation (r= 0.995; p < 0.001) between the STHM activity and the glycine content in rat telencephalon, midbrain, medulla‐pons, spinal cord and retina. These findings suggest that STHM is probably an important enzyme for the synthesis of neural glycine in the vertebrate retina.
The Synthesis, Storage, and Release of Propionylcholine by the Electric Organ of Torpedo marmorata
Authors: O'Regan S.
Published: October 2006

Abstract: Little is known about the specificity of the mechanisms involved in the synthesis and release of acetylcholine for the acetyl moiety. To test this, blocks of tissue from the electric organ of Torpedo were incubated with either [1‐14C]acetate or [1‐14C]propionate, and the synthesis, storage, and release of [1‐14C]acetylcholine and [14C]propionylcholine were compared. To obtain equivalent amounts of the two labeled choline esters, a 50‐fold higher concentration of propionate than of acetate was needed. Following subcellular fractionation, similar proportions of [14C]acetylcholine and [14C]propionylcholine were recovered with synaptosomes and with synaptic vesicles. Furthermore, both labeled choline esters were protected to a similar extent from degradation during homogenization of tissue in physiological medium, indicating that the two choline esters were equally well incorporated into synaptic vesicles. Yet depolarization of tissue blocks by 50 mM KCI released much less [14C]propionylcholinc than [14C]acetylcholine. During field stimulation of the tissue blocks, the difference between the releasibility of the two choline esters was less marked, but acetylcholine was still released in preference to propionylcholine. Evidence for specificity of the release mechanism was also obtained when the release of the two choline esters in response to field stimulation was compared in tissue blocks preincubated with both [3H]choline and [14C]propionate.
Metabolism of Deoxyuridine in Rabbit Brain
Authors: Suleiman Suleiman A. , Spector Reynold
Published: October 2006

Abstract: The metabolism of [3H]deoxyuridine by rabbit brain was investigated in vitro and in vivo. In vitro, brain slices from various regions of brain and from all age groups accumulated [3H]deoxyuridine from artificial CSF. Within the slices, a portion of the accumulated [3H]deoxyuridine was metabolized to [3H]deoxyuridine phosphate, with subsequent conversion to [3H]thymidine phosphate, and ultimately [3H]DNA. The percentage of the [3H]deoxyuridine phosphorylated and subsequently converted into [3H]DNA was highest at birth and declined to adult levels in 3‐month‐old rabbits. Thymidine, when added to the incubation medium with the [3H]deoxyuridine, was approximately 10 times as potent as unlabeled deoxyuridine in inhibiting the intracellular phosphorylation and conversion of [3H]deoxyuridine to [3H]thymidine phosphate in brain slices. In vivo, 2.5 h after intraventricular injection of [3H]deoxyuridine, over 90% of the [3H]deoxyuridine was cleared from the central nervous system at all ages. However, in both newborn and 3‐month‐old rabbits, approximately 40 and 12%, respectively, of the 3H remaining in brain was phosphorylated and converted to [3H]thymidine phosphates; and 11 and 4%, respectively, of the 3H remaining in brain was converted to [3H]DNA. These results show that both immature and mature rabbit brain is able to incorporate deoxyuridine into DNA. Thus, all the enzymes involved in this conversion, including thymidylate synthetase (EC 2.1.1.45), are present and active in brain throughout life.
The Effect of Some Precursor Amino Acids and Enzyme Inhibitors on the Mouse Striatal Concentration of Tyramines and Homovanillic Acid
Authors: Juorio A. V. , Boulton A. A.
Published: October 2006

Abstract: The parenteral administration of L‐phenylalanine or p‐tyrosine increases the mouse striatal concentration of p‐tyramine, an effect that is enhanced by monoamine oxidase inhibition and reduced by an L‐aromatic aminoacid decarboxylase inhibitor. Striatal m‐tyramine was increased following administration of L‐phenylalanine or m‐tyrosine and enhanced further by monoamine oxidase inhibition. It was also observed that m‐tyrosine is a better substrate for decarboxylation than p‐tyrosine, and that p‐tyrosine decarboxylation was blocked by NSD 1055, while that of m‐tyrosine was enhanced. The results obtained indicate that both isomers of tyramine are formed in the mouse striatum by hydroxylation of L‐phenylalanine to p‐ or m‐tyrosine followed by decarboxylation by a specific decarboxylase; an alternative pathway could be first the decarboxylation of phenylalanine to β‐phenylethylamine, followed by its hydroxylation to p‐ or m‐tyramine.
Acetylarsenocholine: A Cholinergic Agonist
Authors: Hedlund Britta , Norin Harald , Christakopoulos Alex , Alberts Peteris , Bartfai Tamas
Published: October 2006

Abstract: The pharmacological properties of acetylarsenocholine, an arsenic analogue of acetylcholine, were investigated. Acetylarsenocholine behaved as a cholinergic ligand both in the central and peripheral nervous system. It bound to nicotinic receptors in rat medulla‐pons with a KD of 15 μM and to muscarinic receptors in rat cerebral cortex with a KD of 10 μM. It behaved also as an agonist at presynaptic muscarinic receptors in guinea pig ileum myenteric plexus preparation. Arsenocholine is an alternative substrate for choline acetyltransferase and acetylarsenocholine is an alternative substrate for acetylcholinesterase.
Calcium Requirement for Fast Axonal Transport in Frog Motoneurons
Authors: Larivière L. , Lavoie P.‐A.
Published: October 2006

Abstract: Calcium is required to sustain fast axonal transport in sensory neurons of frog and cat. We studied the Ca2+ dependence of fast axonal transport in the motoneurons of the lower spinal cord from frog. The accumulation of acetylcholinesterase at a crush on the ventral roots was used to follow axonal transport. Two types of experiments were performed: modification of the medium bathing the ventral roots, alone, and modification of the medium bathing the spinal cord and ventral roots. Incubation (17‐18 h) of the ventral roots in Ca2+‐free medium markedly inhibited acetylcholinesterase transport, a finding that demonstrates a Ca2+ requirement for fast axonal transport in motoneurons; when 4 mM MgCl2 was added to the Ca2+‐free medium, transport was also greatly reduced. During incubation of the ventral roots in normal medium supplemented with 0.18 mM CoCl2 transport proceeded normally; but when the Co2+ concentration was raised to 1.8 mM, transport was diminished as drastically as in the Ca2+‐free medium. Incubation of the spinal cord and ventral roots in medium containing 0.18 mM CoCl2 did not reduce the accumulation of acetylcholinesterase at the crush. Similarly, accumulation of acetylcholinesterase at a crush on the dorsal root was not significantly reduced by exposure of the dorsal root ganglion and root to 0.18 mM Co2+. Exposure of sensory cell bodies to 0.18 mM Co2+ thus produces differential effects on transport of acetylcholinesterase and on transport of newly synthesized radiolabeled protein.
Biosynthesis of Somatostatin‐Like Immunoreactivity by Frog Retinas In Vitro
Authors: Yamada Tadataka , Basinger Scott
Published: October 2006

Abstract: Somatostatin‐like immunoreactivity has been localized to a wide variety of central nervous system neurons, including the retina. We utilized the unique advantages the retina provides for in vitro studies of nerves to examine the biosynthesis of somatostatin. Extracts of frog retinas pulse‐labeled with [35S]cysteine for various time periods revealed uptake of radioactivity into material adsorbable by anti‐somatostatin antibody linked to affinity beads. This uptake increased in a curvilinear fashion for 4 h and was inhibited by cycloheximide (0.2 mM) or by boiling the retinas prior to labeling. Pulse‐chase experiments revealed that affinity‐adsorbable radioactivity from retinal extracts decreased with time of incubation in chase medium; 89% of this decrease could be accounted for by increases in the affinity‐adsorbable radioactivity of the chase medium. Chromatography of the retinal extracts on Sephadex G50 (superfine) revealed four elution peaks, whereas only one peak, co‐eluted with somatostatin‐14, could be identified in the medium. Chromatographic elution patterns of affinity‐adsorbable radioactivity from extracts of pulse‐labeled retinas incubated in chase medium for various times showed a gradual shift of radioactivity from the earlier‐eluting peaks to the later ones. These studies indicate that biosynthesis of somatostatin occurs in frog retinas in vitro. The retina may be a useful model for further study of peptidergic neurons.
Axonal Transport of Choline Lipids in Normal and Regenerating Rat Sciatic Nerve
Authors: Gould R. M. , Spivack W. D. , Sinatra R. S. , Lindquist T. D. , Ingoglia N. A.
Published: October 2006

Abstract: Biochemical methods were used to study the time course of transport of choline phospholipids (labeled by the injection of [3H]choline into the ventral horn of the lumbar spinal cord) in rat sciatic nerve. Autoradiographic methods were used to localize the transported lipid within motor axons. Transported phospholipid, primarily phosphatidylcholine, present in the nerve at 6 h, continued to accumulate over the following 12 days. No discrete waves of transported lipid were observed (a small wave of radioactive phospholipid moving at the high rate would have been missed); the amounts of radioactive lipid increased uniformly along the entire sciatic nerve. In light‐microscope autoradiographs, a class of large‐caliber axons, presumably motor axons, retained the labeled lipid. Some lipid, even at 6 h, was seen within the myelin sheaths. Later, the labeling of the myelin relative to axon increased. The continued accumulation of choline phospholipids in the axons probably signifies their prolonged release from cell bodies and their retention in various axonal membranes, including the axolemma. The build‐up of these phospholipids in myelin probably represents their transfer from the axons to the myelin sheaths surrounding them. When nerves are crushed and allowed to regenerate for 6 or 12 days, choline phospholipids transported during these times enter the regenerating nerve. In light and electron microscope autoradiographs, transported lipid was seen to be localized primarily in the regenerating axons. However, grains overlay the adjacent Schwann cell cytoplasm, indicating transported lipids were transferred from the regenerating axons to the associated Schwann cells. In addition, some cells not associated with growing axons were labeled, suggesting that phosphatidylcholine and possibly acetylcholine, carried to the regenerating axons by axonal transport, were actively metabolized in the terminal, with released choline label being used by other cells. These results demonstrate that axonal transport supplies mature and growing axons and their glial cells with choline phospholipids.
Stimulation of Tubulin‐Dependent ATPase Activity in Microtubule Proteins from Porcine Brain by Vinblastin
Authors: Fujii Toshihiro , Kondo Yoshiyuki , Kumasaka Michiyo , Ohki Kosuke
Published: October 2006

Abstract: Vinblastine, a plant alkaloid which inhibits tubulin polymerization, stimulated an ATPase activity in microtubules. When microtubule proteins were separated into microtubule‐associated proteins (MAPs) and tubulin by phosphocellulose column chromatography, vinblastine did not stimulate an ATPase activity recovered in the MAPs fraction unless tubulin was present. Therefore, vinblastine is considered to act through its binding to the tubulin molecule on MAPs ATPase. Divalent cations that activate tubulin‐dependent MAPs ATPase activity were also required for the stimulation by vinblastine. In the presence of Ca2+ and vinblastine the ATPase activity was most active and the extent of stimulation reached about 200% of the original level in the absence of vinblastine. Half‐maximal stimulation was attained when the molar ratio of vinblastine to tubulin was 0.5. The concentration of tubulin for half‐maximal stimulation was increased in the presence of vinblastine, while divalent cation requirements were decreased. Several factors such as KC1 (100 mM), alkaline pH (pH 7.5), and low temperature (10°C) were not responsible for the disappearance of the stimulation. Vincristine stimulated tubulin‐dependent MAPs ATPase activity as vinblastine did, whereas the activity was scarcely affected by colchicine, podophyllotoxin, strychnine, and chlor‐promazine. Actin had no effect on MAPs ATPase activity in the absence and presence of vinblastine when it was used in place of tubulin.
Origin of the Bicarbonate Stimulation of Torpedo Electric Organ Synaptic Vesicle ATPase
Authors: Rothlein Joan E. , Parsons Stanley M.
Published: October 2006

Abstract: The origin of the stimulation by bicarbonate of the ATPase found in cholinergic synaptic vesicles isolated from Torpedo californica electric organ was studied. Given in descending order of potency, the anions formate, malonate, acetate, thiosulfate, cyanate, biphosphite, biselenite, and bisulfite also stimulate to lesser extents. The anion channel blockers 4‐acetamido‐4′‐isothiocyanostilbene‐2,2′‐disulfonic acid (SITS) and 4,4′‐diisothiocyanostil‐bene‐2,2′‐disulfonic acid (DIDS) inactivated the ATPase at micromolar concentrations, but there was no indication that the reagents acted to block an anion channel. The LineweaverBurk plot of the basal ATPase activity was biphasic, indicating high‐ and low–affinity states with Kms of 0.0098 ′ 0.0013 and 1.2 ′ 0.1 mM for Vmaxs of 52 ′ 2 and 82 ′ 9 nmol/min/mg protein, respectively. It became nearly linear in the presence of bicarbonate, indicating conversion of all of the ATPase to one high affinity‐high velocity state with a Km of 0.054 ′ 0.009 mM and a Vmax of 232 ′ 0.051 nmol/min/mg protein. The pH activity profile for basal ATPase was bell‐shaped and optimal between pH 6 and 8. The bicarbonate stimulation was maximal at about pH 7. The ATPase could be solubilized in 1.6% Lubrol WX with full retention of the bicarbonate stimulation. Although ouabain and oligomycin act at high concentrations to inhibit bicarbonate stimulation of the ATPase, there is no contamination by the Na+‐K+ ATPase, mitochondrial ATPase, or other common ATPases. The study suggests that bicarbonate acts to stimulate the ATPase by acting directly on it at an allosteric site on the outside of the synaptic vesicle.
Factors Contributing to the Poor Myelination in the Brain of the Snell Dwarf Mouse
Authors: Noguchi Tetsuya , Sugisaki Tetsuro , Takamatsu Ken , Tsukada Yasuzo
Published: October 2006

Abstract: Conventional histological examination of the pituitary does not distinguish Snell dwarf mutants (dw/dw) from their normal littermates (+/?) in the neonatal stage. However, immunohistochemical examination of pituitaries of litters born to heterozygous Snell parents revealed that in approximately 25% of the glands examined, the number of positive cells was very low in the neonatal stage. We attempted to delineate the events resulting in the poor myelination in the brain of the Snell dwarf mouse, and to devise an immunohistochemical method for identifying the mutant neonate. Differences in the brain weights of the dw/dw and +/? mice first became apparent on the 10th day of age, and from this time on no further increase in the weight of the dwarf mouse brain was recorded. Increase in CNPase activity was found to be suppressed in the cerebrum and brain stem throughout the developmental stage, but not in the other parts of the brain. The yield of isolated myelin decreased by 58% in the mutant mouse, but CNPase activity was equivalent to that of control myelin. Differences in DNA content per cerebrum from the dw/dw and +/? mice first became apparent on the 10th day of age. Henceforth, the dw/dw mice showed no further increase, although the +/? mice continued to increase. [3H]Thymidine incorporation into the DNA fraction in vivo on the 7th day of age, when glial cell proliferation in the cerebrum is most active, was suppressed to about 50% of the control level in all parts of the dwarf brain. These findings indicate that the poor myelination found in the mutant cerebrum is a hypomyelination due to reduced oligodendroglial proliferation caused by lack of circulating growth hormone.
Postnatal Development of the Acetylcholine System in Different Parts of the Rat Cerebellum
Authors: Kása P. , Bánsághy K. , Rakonczay Z. , Gulya K.
Published: October 2006

Abstract: The components of the cholinergic nervous system, i.e., choline acetyltransferase, acetylcholinesterase, sodium‐dependent high‐affinity choline uptake, acetylcholine, and the muscarinic acetylcholine receptors, in the developing archi‐ and paleocerebellum of the rat have been investigated by biochemical methods. A close correlation between the development of the different elements of the system has been demonstrated in the two areas. The cholinergic structure develops first in the archicerebellum, which displays high levels of choline acetyltransferase, acetylcholinesterase, acetylcholine, and sodium‐dependent high‐affinity choline uptake. The paleocerebellum receives a sparser cholinergic innervation during development. The differences in the values for these components in the cerebellum as a whole may reflect the development of cholinergic and noncholinergic neuronal structures. It is concluded that the development of the cholinergic system cannot be analyzed in the cerebellum as a whole; rather specific regions such as the archi‐, paleo‐, or neocerebellum must be examined.
Solid‐Phase Immunoassays for Quantitation of Antibody to Bovine White Matter Proteolipid Apoprotein
Authors: Macklin Wendy B. , Lees Marjorie B.
Published: October 2006

Abstract: Two solid‐phase immunoassays have been developed for quantitation of antibodies to bovine white matter proteolipid apoprotein. Conditions were established for optimal specific antibody binding. Water‐soluble proteolipid apoprotein was bound to microtiter plates and plates were incubated with test serum. Goat anti‐rabbit IgG conjugated with horeseradish peroxidase was used as the second antibody for an enzyme‐linked immunospecific assay and 125I‐labeled protein A for a radioimmunoassay. Both procedures have been used to follow the time course of anti‐proteolipid antibody production in rabbits and to compare different immunization protocols.
Mass Fragmentographic Method for the Determination of Trace Amounts of Putative Amino Acid Neurotransmitters and Related Compounds from Brain Perfusates Collected In Vivo
Authors: Wolfensberger M. , Amsler U.
Published: October 2006

Abstract: A mass fragmentographic method for the determination of trace amounts of amino acid neurotransmitter candidates from brain perfusates is described. The analytical procedure includes the measurements of glycine, β‐alanine, γ‐aminobutyric acid, proline, aspartic acid, and glutamic acid; αalanine, leucine, and sarcosine, undergoing gas chromatographic coelution, are detected simultaneously. Amino acids extracted from dried perfusate residues are converted to the corresponding N‐pentafluoropropionyl hexafluoroisopropyl esters by a single‐step procedure. Gas chromatographic separation of the amino acid derivatives is achieved on a packed glass column filled with trifluoropropylsilicone as stationary phase. The limit of detection for the different derivatives (signal‐to‐noise, 3:1) ranges from 50 femtomol to 1 picomol. Deuterium‐labeled amino acid analogues are used as internal standards for quantitative measurements. The mass spectral characteristics of the derivatives are compared and discussed. The technique has been applied to the assay of amino acids released in vivo within the pigeon optic tectum, demonstrating the capabilities of the present analytical approach.