bluuebunny/crossref_metadata_2025 huggingface dataset:
Dopaminergic Mechanisms in the Teleost Retina
Authors: Dowling John E. , Watling Keith J.
Published: October 2006

Abstract: The ability of dopamine, dopamine agonists, other proposed retinal neurotransmitters, depolarizing agents and light to stimulate adenylate cyclase activity in pieces of intact carp retina has been examined. The evidence indicates that a dopamine‐sensitive adenylate cyclase is the only neurotransmitter activated adenylate cyclase in the carp retina. That is, only dopamine, or agents that activate dopamine receptors, appear to stimulate cyclic AMP synthesis in the retina. Depolarizing agents such as K+ or veratridine also increase retinal cyclic AMP levels, but apparently by releasing endogenous stores of dopamine. For example, the increase of retinal cyclic AMP levels induced by 45 mM‐K+ is blocked by 5 mM‐Co2+ or 100 μM‐haloperidol, a dopamine antagonist. Flashing lights slightly increase cyclic AMP levels in the retina, an effect that is likewise abolished by haloperidol.
Highly Sensitive Immunoassays for Three Forms of Rat Brain Enolase
Authors: Kato Kanefusa , Suzuki Fujiko , Umeda Yumiko
Published: October 2006

Abstract: Highly sensitive enzyme irnmunoassay systems for three forms (αα, αγ, and γγ) of rat brain enolase were prepared by use of specific antisera to two distinct subunits (α and γ) of the isozymes and β‐D‐galactosidase from Escherichia coli as label. Less than fmol‐levels of the homologous dimer forms (αα and γγ) could be determined with the corresponding antibody F(ab′)2‐bound solid‐phase and the antibody Fab′‐βD‐galactosidase complex. The hybrid form (αγ) could also be assayed specifically by use of the antibody to one subunit as solid‐phase, and the antibody to another subunit as labelled complex with a minimum detectable sensitivity of 1 fmol.
Electroconvulsive Shock and Reserpine: Effects on β‐Adrenergic Receptors in Rat Brain
Authors: Kellar Kenneth J. , Cascio Caren S. , Bergstrom Debra A. , Butler Judith A. , Iadarola Paul
Published: October 2006

Abstract Electroconvulsive shock (ECS) administered once daily for up to 14 days decreases β‐adrenergic receptor binding in the cortex and hippocampus in a time‐dependent manner. The decrease in binding in the cortex lasts at least 1 week after the last shock. In the striatum, hypothalamus, or cerebellum, 14 days of ECS did not produce significant changes in β‐adrenergic receptor binding. The brain regional pattern of β‐adrenergic receptor changes suggests that repeated ECS affects β‐adrenergic receptors in brain regions that receive a noradrenergic innervation activated by ECS. The effects of ECS on neurotransmitter receptor binding appear to be highly selective. Of five receptors in the cortex and three receptors in the hippocampus measured, only β‐adrenergic receptor binding is decreased. Chronic footshock stress does not alter β‐adrenergic receptor binding sites in the cortex, indicating that the effects of ECS are not due to stress alone. The effects of ECS on reserpine‐induced alterations in β‐adrenergic receptor binding sites were also examined. Ten days of ECS following chronic reserpine injections reverses the increased binding of β‐adrenergic receptors
Characterization of Immunoreactive Thyrotropin Releasing Hormone in Human Fetal Cerebellum
Authors: Parker C. Richard
Published: October 2006

Abstract— High concentrations (411 ± 30 pg/mg protein; mean ± S.E., n= 12) of immunoreactive TRH (TRHi) were detected in extracts of human fetal cerebellum (13‐26 weeks gestation). The TRHi in the cerebellar extracts, when subjected to gel filtration and high performance liquid chromatography (HPLC), co‐migrated with synthetic TRH. In each instance, no TRHi was detected which did not share identical chromatographic mobilities with synthetic TRH. Like synthetic TRH, the TRHi in extracts of fetal cerebellum and hypothalamus was insoluble in diethyl ether and was efficiently degraded when incubated at 37°C with adult rat serum. No significant degradation of TRHi occurred when the incubation was conducted at 0°C or when the extracts were incubated with rat serum that had been preheated at 60°C for 20 min. Neither synthetic TRH nor TRHi in extracts of fetal cerebellum or hypothalamus was degraded when incubated with human umbilical cord serum at 37°C or 0°C. The results of this study are supportive of the view that the TRHi in extracts of human fetal cerebellum is identical to TRH.
Comparison of Proteoglycans Extracted by Saline and Guanidinium Chloride from Cultured Chick Retinas
Authors: Morris John E. , Ting Yuan‐Pin
Published: October 2006

Abstract: To compare the loosely associated sulfated proteoglycans with those tightly bound to membranes, retinas from 14‐day chick embryos were subjected to progressively disruptive techniques. The most easily removed proteoglycans were isolated from the medium in which the tissue was labeled with [35S]sulfate. On the average, 25% of the glycosaminoglycans were in the labeling medium, 39% were in proteoglycans extracted from the tissue in the balanced salt solution, 32% were in a 4 m‐guanidinium chloride (GuCl) fraction, and 4% remained unextracted. These glycosaminoglycans contained, respectively, 28, 28, 40, and 4% of the incorporated [35S]sulfate. On the basis of electrophoretic mobility and TLC of chondroitinase digests, the ratio of 35S in chondroitin sulfate to that in heparan sulfate was 4–7 times higher in the medium and balanced salt extracts than in the GuCl extracts. In both extracts there was more 35S in chondroitin‐6‐sulfate than in chondroitin‐4‐sulfate. Dialysis of the extracts against 0.5 M‐NaCl resulted in the precipitation of about 12% of the glycosaminoglycans in the saline extracts and about 40% in GuCl extract. These subfractions, which were relatively enriched in heparan sulfate, were largely soluble in dithiothreitol in 8 m‐urea (DTT). Similarities between the proteoglycans in the medium and those extracted by balanced salt solutions suggest that the saline‐extracted proteoglycans were for the most part loosely associated with cell surfaces or extracellular matrices, whereas the GuCl‐extracted proteoglycans probably were bound to membranes.
Regional Distribution of Kininase in Rat Brain
Authors: Kariya K. , Kawauchi R. , Okamoto H.
Published: October 2006

Abstract: Kininase activity, which inactivates kinins, was measured in seven regions of the rat brain (i.e., the cerebral cortex, cerebellum, striatum, midbrain, hippocampus, hypothalamus, medulla oblongata), and in the spinal cord with a bioassay method using bradykinin as the substrate. Specific kininase activities in the cerebellum and striatum were higher than those in the other five regions or the spinal cord. Angiotensin‐converting enzyme activity, which was measured fluorometrically using Hip‐His‐Leu as substrate, showed high activity in the striatum and cerebellum. These findings suggest that the presence of high concentrations of peptidases plays a role in the degradation of kinins and/or other peptides in these areas.
Comparative Effects of Ethanol and Malnutrition on the Development of Catecholamine Neurons: A Long‐Lasting Effect in the Hypothalamus
Authors: Detering N. , Collins R. M. , Hawkins R. L. , Ozand P. T. , Karahasan A.
Published: October 2006

Abstract: Exposure to ethanol during only the last week of gestation in the rat resulted in significantly decreased weight and levels of norepinephrine in the hypothalamus of the offspring after 26 weeks on standard diet compared with appropriate controls.
Lipid and Protein Alterations of Spinal Cord and Cord Myelin of Multiple Sclerosis
Authors: Yu Robert K. , Ueno Kunihiro , Glaser Gilbert H. , Tourtellotte Wallace W.
Published: October 2006

Abstract: A comprehensive study was carried out to clarify the chemical compositions of spinal cord, cord myelin, and myelin subfractions of multiple sclerosis (MS). The protein compositions of normal‐appearing cerebral white matter and cerebral plaque and periplaque tissues were also analyzed for comparison. MS whole cord samples were found to contain higher amounts of water compared with normal samples. The total lipid contents were below normal. Among the individual lipids, cholesterol content remained unchanged, whereas cholesteryl esters appeared increased in MS cords. The acidic phospholipid concentrations were found to be lower than normal. Glycolipids, such as cerebrosides GM4, GM1, and GD1b, which are abundant in myelin, were all decreased. However, the concentrations of GM3 and GD3, which are more characteristic of reactive astrocytes, were highly elevated. The total protein content of MS cord samples was decreased, and the decrease was attributable to the loss of myelin proteins as evidenced by the low recovery of myelin. The concentrations of myelin‐specific proteins, such as proteolipid protein and myelin basic protein, were significantly reduced. Other changes in the protein compositions included the accretion of two low molecular weight proteins of approximately 11,000 and 12,000, and the appearance of a periodic acid‐Schiff‐positive protein with the same electrophoretic mobility as the P0 protein. Analysis of the isolated myelin indicated that it had a grossly normal protein composition. However, the two low molecular weight proteins and the P0 protein appeared to be enriched in an upper‐phase cord subtraction. We attribute the appearance of the two low molecular weight proteins to the breakdown of proteolipid protein and/or myelin basic protein as a result of demyelination, and the appearance of P0 to the involvement of PNS myelin. The latter finding provides the first biochemical evidence that in MS cord, remyelination can be achieved in part by invading Schwann cells and/or by the small number of Schwann cells that may be present in the cord.
Higher GABA Concentrations in Fallopian Tube Than in Brain of the Rat
Authors: Erdö S. L. , Rosdy B. , Szporny L.
Published: October 2006

Abstract: The GABA content was determined simultaneously in two peripheral organs, i.e., ovary and Fallopian tube. Moreover, the effects of inhibitors of glutamate decarboxylase or γ‐aminobutyrate transaminase (GABA‐T) on the GABA concentrations of the two organs were examined, to point out similarities and differences between central and peripheral pathways of GABA biosynthesis and degradation. In ovary, GABA concentration was found to be about 30% of that in total brain tissue. Furthermore, isoniazid and thiosemicarbazide caused significant reduction of GABA levels in peripheral organs. In contrast to the CNS, aminooxyacetic acid failed to increase, but even produced a significant diminution in peripheral GABA content. Gabaculine did not change GABA levels. In conclusion, it has been demonstrated for the first time that a peripheral organ, i.e. fallopian tube, contained higher GABA concentrations than the CNS. On the other hand, in the organs examined GABA seemed to be synthesized similarly, but metabolized by a pathway different from that in the brian.
Metabolism and Brain Uptake of γ‐Aminobutyric Acid in Galactosamine‐Induced Hepatic Encephalopathy in Rats
Authors: Zeneroli M. L. , Iuliano E. , Racagni G. , Baraldi M.
Published: October 2006

Abstract: Kinetic studies of [3H]γ‐aminobutyric acid ([3H]GABA) after an intravenous injection were performed in normal rats and in rats with severe degree of hepatic encephalopathy due to fulminant hepatic failure induced by galactosamine. Moreover, plasma and brain GABA levels, and GABA and glutamic acid decarboxylase activity were studied in some brain areas. After intravenous injection, [3H]GABA disappeared very rapidly in the blood of normal rats, with a prompt increase of 3H metabolites. In comatose rats, a delayed disappearance of [3H]GABA.as parallelled by a lower amount of metabolites, indirectly indicating a peripheral decrease of GABA‐transaminase activity. The amount of [3H]GABA in brain was lightly but constantly lower in comatose rats than in controls, indicating that the change in permeability of the blood‐brain barrier in hepatic encephalopathy does not affect the [3H]GABA uptake of the brain. Furthermore, the assay of endogenous GABA in blood, whole brain, and brain areas did not show any significant difference in any of the two groups. The finding that glutamic acid decarboxylase activity in brain was reduced, together with the indirect evidence of a reduction in GABA‐transaminase, may account for the steady state of GABA in hepatic encephalopathy. However, the reduction in glutamic acid decarboxylase activity is in favor of a functional derangement at the GABA‐ergic nerve terminals in this pathological condition.
Immunochemical Studies of Bovine and Human Choline‐O‐Acetyltransferase Using Monoclonal Antibodie
Authors: Levey A. I. , Rye D. B. , Wainer B. H.
Published: October 2006

Abstract: Immunochemical properties of bovine and human choline acetyltransferase (ChAT, EC 2.3.1.6, acetyl‐CoA:choline‐O‐acetyltransferase) were studied using six monoclonal antibodies (AB1, AB5, AB6, AB7, AB8, and AB9) reactive with the enzyme. All antibodies except AB1 bound specifically to two proteins of 68,000 and 70,000 MW on “Western” blots of sodium dodecyl sulfate‐polyacrylamide gels containing human or bovine ChAT. The enzyme was specifically absorbed to immobilized antibody and could not be eluted by low pH and/or high salt concentrations, although the enzyme retained activity on the immunoabsorbent. Pure bovine enzyme consisting of the same two proteins as seen in the Western blotting studies was eluted from immobilized AB1 in the presence of sodium dodecyl sulfate. Although active enzyme could not be eluted from immobilized antibodies by standard conditions, various combinations of free and immobilized antibodies were effective in competing off bound enzyme. Free antibody AB1 quantitatively eluted the active enzyme from immobilized AB1. The different capacities of the antibodies to elute enzyme from various immunoabsorbents reflect interesting properties of both the enzyme and the antibodies.
Thyrotrophin‐Releasing Hormone Analogues Increase Dopamine Release from Slices of Rat Brain
Authors: Sharp Trevor , Bennett Geoffrey W. , Marsden Charles A.
Published: October 2006

Abstract: Rat brain slices were incubated with a high concentration of K+, thyrotrophin‐releasing hormone (TRH), or one of two biologically stable TRH analogues (CG 3509 or RX 77368). Basal release of endogenous dopamine, measured by electrochemical detection, was increased by K+ (30 mM) from slices of hypothalamus, septum, nucleus accumbens, and striatum. CG 3509 (105–10−3M) increased the release of dopamine from slices of nucleus accumbens, septum, and hypothalamus in a dose‐dependent fashion, whereas RX 77368 (10−4M) increased the release of dopamine from the septum only. Neither analogue increased the release of striatal dopamine. The results provide further evidence for specific regional interactions between TRH and dopamine in rat brain.
The Ontogeny of Acetylcholinesterase Activities in Rat Brain Regions and the Effect of Chronic Treatment with Manganese Chloride
Authors: Lai James C. K. , Leung Thomas K. C. , Lim Louis
Published: October 2006

Abstract: Acetylcholinesterase activities were determined in the rat cerebral cortex, striatum, midbrain, pons and medulla, hypothalamus, and cerebellum at 5, 12, 20, 30, and 60 days after birth. The ontogeny of the enzyme differed in the various regions, occurring earlier in the more caudal regions, except in the cerebellum where there was no increase. Chronic manganese treatment from conception did not influence the developmental profile of this cholinergic marker.
Characteristics of Protein Carboxyl Methylation in the Rat Hypothalamus
Authors: Eiden Lee E. , Borchardt Ronald T. , Rutledge Charles O.
Published: October 2006

Abstract: The formation of methyl‐labeled S‐adenosylmethionine (AdoMet) and methyl esters of endogenous methyl‐acceptor proteins (MAPs) was studied in a synaptosomal preparation from the rat hypothalamus labeled with L‐[methyl‐3H]methionine. Incubation of synaptosomes with l‐[methyl‐3H]methi‐onine resulted in a rapid labeling of the AdoMet pool and a less rapid formation of 3H‐methyl‐MAPs. Accumulation of 3H‐methyl‐MAPs was linear over a 30‐min period. The effects of various inhibitors of AdoMet‐dependent trans‐methylation reactions on the formation of carboxylmethylated MAPs were examined. When hypothalamic synaptosomes were preincubated with l‐[methyl‐3H]methionine and subsequently incubated for 30 min in the presence of S‐adenosyl‐l‐homocysteine (AdoHcy, 100 μm), 3H‐methyl‐MAP formation was inhibited by approximately 70%. 100 μm‐l‐homocysteine thiolactone (HTL) as well as 100 μm‐3‐deazaadenosine (c3Ado) also caused a 60–70% inhibition of 3H‐methyl‐MAP formation; the combination of both c3Ado and HTL produced a slightly but not significantly greater inhibition than either agent alone. 10 μm‐adenosine or 10 μm‐HTL each produced an approximately 40% inhibition of 3H‐methyl‐MAP formation: the inhibitory effect of the two agents in combination was additive. Sinefungin and A9145C, potent inhibitors of bovine adrenomedullary protein carboxyl methylase, had no effect on 3H‐methyl‐MAP formation in hypothalamic synaptosomes at concentrations up to 1 mM. However, these compounds were potent inhibitors of 3H‐methyl‐MAP formation in lysed synaptosomes incubated with [3H‐methyl]AdoMet. These results demonstrate that hypothalamic synaptosomes are capable of methio‐nine activation and protein carboxyl methylation.
Glycine Receptors in the Human Substantia Nigra as Defined by [3H]Strychnine Binding
Authors: de Montis G. , Beaumont K. , Javoy‐Agid F. , Agid Y. , Constandinidis J. , Lowenthal A. , Lloyd K. G.
Published: October 2006

Abstract: Specific [3H]strychnine binding was used to identify the glycine receptor macromolecular complex in human spinal cord, substantia nigra, inferior olivary nucleus, and cerebral cortex. In material from control patients a high‐affinity Kd (3–8 nm) was observed in the spinal cord and the substantia nigra, both the pars compacta and the pars reticulata. This is very similar to the values observed in the rat and bovine spinal cord (8 and 3 nm, respectively) and rat substantia nigra (12 nm). In the human brain the distribution of [3H]strychnine binding (at 10 nm) was: spinal cord – substantia nigra, pars compacta > substantia nigra, pars reticulata = inferior olivary nucleus > cerebral cortex. The binding capacity (Bmax) of the rat brain (substantia nigra or spinal cord) was approximately 10‐fold that of the human brain. [3H]Strychnine binding was significantly decreased in the substantia nigra from Parkinson's disease patients, both in the pars compacta (67% of control) and the pars reticulata (50% of control), but not in the inferior olivary nucleus. The results were reproduced in a preliminary experiment in rats with unilateral 6‐hydroxydopamine lesions of the medial forebrain bundle. In the substantia nigra from patients who died with Huntington's disease, [3H]strychnine binding tended to be high (150% of control, NS) in both the pars compacta and the reticulata. [3H]Strychnine binding was unaltered in the substantia nigra of patients with senile dementia. Together with previous neurophysiological and neuropharmacological findings, those results support the hypothesis of glycine receptors occurring on dopamine cell bodies and/or dendrites in the substantia nigra.
Adenylate Cyclases in the Vertebrate Retina: Distribution and Characteristics in Rabbit and Ground Squirrel
Authors: De Vries G. W. , Campau K. M. , Ferrendelli J. A.
Published: October 2006

Abstract: Adenylate cyclase activity and the effects of EGTA, 5′‐guanylylimidodiphosphate (GPP(NH)P), and dopamine were measured in microdissected layers of rod‐dominant (rabbit) and cone‐dominant (ground squirrel) retinas. The distribution of basal enzyme activity was similar in both species, with the highest levels found in the inner plexiform and photoreceptor cell inner segment layers. EGTA inhibited adenylate cyclase in the inner retina of both species and stimulated activity in rabbit outer and inner segment layers, but had no effect in these layers from ground squirrel. Enzyme activity was stimulated in all regions by GPP(NH)P, except in the outer segments of the photoreceptors. Dopamine stimulated the enzyme in the outer and inner plexiform and inner nuclear layers in rabbit, but only in the inner plexiform layer in ground squirrel. These data demonstrate that the enzymatic characteristics of adenylate cyclase vary extensively from region to region in vertebrate retina and suggest that cyclic AMP may have multiple roles in this tissue. A model for the distribution of the different forms of adenylate cyclase in retina is proposed.
Regional Localization and Subcellular Compartmentalization of Thyrotropin‐Releasing Hormone in Adult Human Brain
Authors: Parker C. Richard , Porter John C.
Published: October 2006

Abstract: In the current study, we sought to define the subcellular compartmentalization of thyrotropin‐releasing hormone (TRH) in adult human brain tissues. Upon evaluating tissues (3–24 h post mortem) from 62 humans, ranging in age from 5 to 75 years, we found that TRH was widely distributed throughout the brain. The highest TRH concentration (ng/mg protein) was in the stalk‐median eminence region of the hypothalamus (19.3 ± 3.3, mean ± SE); the TRH concentration in the hypothalamus, exclusive of the stalk‐median eminence, was much lower (1.7 ± 0.2). Substantial quantities of TRH also were detected in the medulla oblongata (0.26 ± 0.08), mammillary bodies (0.33 ± 0.25), and optic chiasm (0.14 ± 0.07). Lower levels of TRH were found in the amygdala (0.060 ± 0.015) and the corpus striatum (0.033 ± 0.010). TRH was near or below the limits of detection in tissues of the cerebral and cerebellar cortices, the olfactory bulbs, the pons, and the hippocampus. When homogenates of medial basal hypothalamic tissue (preparedin 0.32 M sucrose‐10 μM CaCl2) were fractionated by means of differential centrifugation, most of the TRH was recovered in subcellular particles which were pelleted at 10,000 ×g and which contained the highest amounts of occluded LDH activity. When the nuclei‐free supernatant fluid (900 ×g S) was fractionated on discontinuous sucrose density gradients or continuous sucrose density gradients, most of the TRH was recovered in subcellular fractions containing synaptosomes. The subcellular distribution of TRH appeared to be stable for up to 24 h post mortem in rat and human brain tissue. In contrast to findings with homogenates of rat hypothalami, exposure of subcellular particles of human hypothalamic origin to hypoosmotic shock caused only minor release of particle bound TRH, which was recovered in the cytosolic fraction. Also in contrast to findings with rat hypothalamic tissue, we were unable to demonstrate convincingly the presence of TRH containing subsynaptosomal particles in homogenates of human hypothalami which were resistant to hypoosmotic shock.
Heterogeneity of Benzodiazepine Receptors: Experimental Differences Between [3H]Flunitrazepam and [3H]Ethyl‐β‐carboline‐3‐carboxylate Binding Sites in Rat Brain Membranes
Authors: Medina Jorge H. , Novas María Laura , De Robertis Eduardo
Published: October 2006

Abstract: This study was designed to analyze possible differences in the binding of [3H]flunitrazepam ([3H]FNZP) and [3H]ethyl ‐ β ‐ carboline ‐ 3 ‐ carboxylate ([3H]β‐CCE), to rat brain membranes, in various experimental conditions. In cerebral cortex, hippocampus, cerebellum, and orain stem the number of binding sites for [3H]β‐CCE was higher than for [3H]FNZP; both were displaced by clonazepam. Until the 7th day of postnatal brain development the numbers of [3H]FNZP and [3H]β‐CCE sites were equivalent; but later on, the β‐carboline sites increased to a higher level. Noradrenergic denervation by 6‐hydroxydopamine was followed in the hippocampal formation. Already after 2 days, there was a decrease in [3H]FNZP sites, which reached 70% of control after 14 days. Similar results were obtained with DSP‐4 denervation. This change was only in Bmax and not in KD, In contrast, the [3H]β‐CCE sites did not change with denervation. Neonatal injection of l ‐ 2,4,5 ‐ trihydroxyphenylalamine or DSP‐4 produced in the adult a decrease in [3H]FNZP sites in the cerebral cortex, in parallel with the noradrenergic denervation. On the other hand, there was an increase in the cerebellum and brain stem, in correspondence with the hyperinnervation by sprouting. In these rats, the number of sites for [3H]β‐CCE did not change in the different brain regions. With 0.1% Triton X‐100, applied to synaptosomal membranes, [3H]FNZP binding was reduced by 35%, while that of [3H]β‐CCE was not significantly changed. These results suggest that there is heterogeneity of binding sites for benzodiazepine receptors in rat brain. A tentative interpretation of the experiments involving noradrenergic denervation and hyperinnervation, as well as those with Triton X‐100, is that [3H]FNZP binds to pre‐ and postsynaptic receptors, while [3H]β‐CCE binds mainly to postsynaptic benzodiazepine receptors.
Effects of a 4‐Aminopyridine in Calcium Movements and Changes of Membrane Potential in Pinched‐Off Nerve Terminals from Rat Cerebral Cortex
Authors: Agoston D. , Hargittai P. , Nagy A.
Published: October 2006

Abstract: A study was conducted on the influence of 4‐aminopyridine (4‐AP) on the radiocalcium uptake and membrane potential of rat cortical synaptosomes previously depolarized by biochemical procedures. The initial calcium entry into isolated nerve terminals was substantally enhanced in the presence of 10‐−4M 4‐AP in potassium‐rich media (60 mM). The fast initial phase of potassium‐stimulated calcium entry involves different kinetic characteristics in the presence and the absence of 4‐AP. In the presence of 4‐AP, the fast component of calcium entry reached a peak during the first 15 s immediately following depolarization. The potassium‐stimulated synaptosomal calcium influx at 15 s was 3.5 ± 0.17 times higher in the presence of 4‐AP (10‐−4M) and 2.6 ± 0.11 times higher in the absence of 4‐AP as compared with the respective unstimulated uptake values. In the absence of 4‐AP, however, the calcium entry did not show a peak until 45 s after depolarization. The total amount of calcium accumulated in the synaptosome treated and untreated by 4‐AP is equal at the end of this initial uptake period. The effect of 4‐AP on membrane potential during depolarization of synaptosomes evoked by potassium‐rich medium was also determined by means of a potential sensitive fluorescent dye. It was found that 4‐AP had no effect on the final level of membrane potential, induced by high [K+]o; however, the kinetics showed significant differences. The initial phase of synaptosomal membrane depolarization was slower in the presence of 4‐AP: τ‐4‐AP: 42 ± 4.0 s, τ‐control: 18 ± 2.5 s. The initial calcium entry into the nerve terminals was increased during the 4‐AP treatment in potassium‐rich media—as resulted from our calcium uptake studies—and thus, during this higher calcium influx, the degree of membrane depolarization was decreased. This faster initial calcium influx constitutes a feasible explanation for the unique effect of 4‐AP termed “chemical potentiation” on transmitter release at chemical synapses.
Development and Applications of a Solid‐Phase Radioimmunoassay for the P0 Protein of Peripheral Myelin
Authors: Fryxell Karl J. , Balzer David R. , Brockes Jeremy P.
Published: October 2006

Abstract: This is the first report of a quantitative radioimmunoassay for P0. The assay uses antigen‐coated plastic microwells, with antibody binding detected by 125I‐labeled protein A. Either peripheral myelin proteins or purified P0 may be used as the antigen. Optimal extraction of tissue samples for P0 immunoassay requires careful attention to the sodium dodecyl sulfate‐to‐protein ratio. Sodium dodecyl sulfate interference with antibody binding can be minimized by adding an excess of nonionic detergent and carrier protein to the incubation buffer. This method allows the detection of 0.8 ng of P0 (20 ng/ml). Results from this assay showed little or no im‐munoreactivity in extracts of brain, central myelin, liver, purified myelin basic proteins, cultured, purified second ary Schwann cells, or membrane preparations from these cells. P0 was clearly detectable in Schwann cell cultures from 3‐ to 4‐day‐old rats at 12–18 h after dissociation (4% of the level in adult sciatic nerve) and in extracts of one‐day‐old rat sciatic nerve (2% of the level in adult nerve). Myelin basic protein radioimmunoassays showed that the ratio of P0 to myelin basic protein is essentially constant in extracts of sciatic nerve from one‐day‐old, four‐day‐old and young adult rats. Another result was that P0 levels are reduced in the trembler mouse sciatic nerve.
Biochemical Assessment of Serotonergic and Cholinergic Dysfunction and Cerebral Atrophy in Alzheimer's Disease
Authors: Bowen D. M. , Allen S. J. , Benton J. S. , Goodhardt M. J. , Haan E. A. , Palmer A. M. , Sims N. R. , Smith C. C. T. , Spillane J. A. , Esiri M. M. , Neary D. , Snowdon J. S. , Wilcock G. K. , Davison A. N.
Published: October 2006

Abstract: Markers of serotonin synapses in entire temporal lobe and frontal and temporal neocortex were examined for changes in Alzheimer's disease by use of both neurosurgical and autopsy samples. Uptake of [3H]sero‐tonin, binding of [3H]imipramine, and content of indola‐mines were all significantly reduced, indicating that serotonin nerve terminals are affected. Binding of [3H]serotonin was also reduced, whereas that of [3H]qui‐nuclidinyl benzilate, [3H]muscimol, and [3H]dihydroal‐prenolol were unaltered. When the Alzheimer's samples were subdivided according to age, the reduction in [3H]serotonin binding was a feature of only autopsy samples from younger patients. In contrast, presynaptic cholinergic activity was reduced in all groups of Alzheimer's samples, including neurosurgical specimens. Five markers, thought to reflect cerebral atrophy, cytoplasm, nerve cell membrane, and neuronal perikarya were measured in the entire temporal lobe. In Alzheimer's disease the reductions (mean 25%, range 20–35%) were thought to be too large to be due only to loss of structures associated with the presumed cholinergic perikarya in the basal forebrain and monoamine neurones in the brain stem.
[3H]Xylamine Accumulation by Two Sympathetically Innervated Tissues
Authors: Ransom Richard W. , Waggaman Lorene A. , Cho Arthur K.
Published: October 2006

Abstract: The accumulation of [3H]xylamine ([ring‐G‐3H]‐N‐2′‐chloroethyl‐N‐ethyl‐2‐methylbenzylamine; [3H]XYL) by rabbit thoracic aorta and rat vas deferens was studied in vitro. [3H]XYL uptake in both tissues saturated at the same concentrations and displayed the same time course as that for maximal inhibition of [3H]noradrenaline ([3H]NA) accumulation by XYL. Hydrolysis of [3H]XYL or coincubation with Na2S2O3 reduced tissue accumulation of tritium by 80%. In aorta and vas deferens, about 45% and 65%, respectively, of the total [3H]XYL accumulation was blocked by 100 μmol/L desmethylimipramine (DMI), l‐NA, or bretylium. In the absence of sodium ion, the uptake of [3H]XYL was reduced by approximately these same amounts. In both tissues nearly 70% of the [3H]XYL taken up was protein bound when measured as trichloroacetic acid‐precipitated tritium. Of this radioactivity, the same proportion was sensitive to 100 μmol/L DMI or the absence of sodium ion as found for total tissue accumulation of [3H]XYL. Hydrolysis of [3H]XYL prior to incubation with vas deferens reduced the protein‐bound tritium by more than 95%. The studies indicate that [3H]XYL interacts with NA uptake carrier in both rabbit aorta and rat vas deferens and that a substantial portion of the resulting protein‐bound radioactivity is carrier‐dependent.
Functional Significance of the Effects of Neurotransmitters on the Na+,K+‐ATPase System
Authors: Kometiani Z. P. , Tsakadze L. G. , Jariashvili T. Ya.
Published: October 2006

Abstract: The aim of the present experiments was to study the effects of the neurotransmitters acetylcholine, noradrenaline, 5‐hydroxytryptamine, and dopamine on the Na+,K+‐ATPase of rat brain synaptosomal fractions. It is shown that dopamine at low concentrations specifically inhibits the Na+,K + ‐ATPase of synaptic membranes from the brain regions rich in dopaminergic endings, but has no effect on the synaptosomal Na+,K+‐ATPase from the other parts of brain. Acetylcholine and noradrenaline have similar specific effects on Na+,K+‐ATPase from cholinergic and adrenergic synaptosomes. The Na+,K+‐ATPases of synaptic membranes from the different brain regions, characterised by different distributions of cholinergic, adrenergic, and 5‐hydroxytrypta‐minergic endings, show different reactions with neurotransmitters. These data indicate a functional significance of the effects of the neurotransmitters on the synaptosomal Na+,K+‐ATPase.
Two Populations of Axonally Transported Tubulin Differentiated by Their Interactions with Neurofilaments
Authors: Tashiro Tomoko , Kurokawa Masanori , Komiya Yoshiaki
Published: October 2006

Abstract: In the sensory fibers of the rat sciatic nerve (fibers of the dorsal root ganglion cells), two components of tubulin transport were observed that differed in the rate of transport, solubility in Triton, and subunit composition. The faster component, migrating ahead of the neurofilament proteins, was soluble in 1% Triton. The slower component, migrating with the neurofilament proteins, was insoluble in 1% Triton and contained a unique polypeptide, “NAP,” in the tubulin region that was not present in the faster component. “NAP” was not a subspecies of tubulin as evidenced by peptide mapping. It seems to be a neurofilament‐associated protein. When a complete separation of the main tubulin wave from the neurofilament wave was achieved in the motor axons of the same nerve (axons of the ventral motoneurons) under the effect of ββ‐iminodipropionitrile, a portion of tubulin was still found associated with the retarded neurofilament wave. The subunit composition of this portion was similar to the slower, neurofilament‐associated component in the sensory fibers under normal conditions, i.e., enriched in “NAP” and the most acidic subtype of (β‐tubulin. It is suggested that two populations of transported tubulin exist that are differentiated by the extent of their interaction with neurofilaments.
Hormonal Regulation of Adipose S‐100 Protein Release
Authors: Suzuki Fujiko , Kato Kanefusa , Nakajima Takashi
Published: October 2006

Abstract: The release of S‐100 protein from epididymal fat pads was enhanced by epinephrine in vitro, and about 50% of S‐100 protein in the tissue was released into the medium after 2‐h incubation at 37°C with 10 μM epinephrine. Similar results were obtained with the incubation of isolated adipocytes. The S‐100 protein release was also enhanced by isoproterenol, norepinephrine, ACTH, and dibutyryl cyclic AMP, which all increase the lipolysis by increasing cyclic AMP levels in the tissue. Propranolol, a β‐adrenergic blocker, could block the increase of S‐100 protein release by catecholamines, indicating that the release was mediated by the β‐adrenergic effect of catecholamines. However propranolol had no suppressive effect on the enhancement of S‐100 protein release by ACTH or dibutyryl cyclic AMP. Insulin had an inhibitory effect on the epinephrine‐enhanced S‐100 protein release. Epinephrine or ACTH could not stimulate the S‐100 protein release in the absence of Ca2+, whereas the epinephrine‐enhanced glycerol release was not affected under the same conditions. The increase in S‐100 protein release was induced by only a pretreatment of the tissue with epinephrine. However, the lipolysis in the tissue was not enhanced by the pretreatment alone. These results indicate that the release of S‐100 protein from adipocytes is regulated by the hormones that have been known to control the lipolysis with a manner slightly different from that of lipolysis.
Studies on the Bovine Brain Pyruvate Dehydrogenase Complex Using the Antibodies Against Kidney Enzyme Complex
Authors: Sheu Kwan‐Fu Rex , Kim Young Tai
Published: October 2006

Abstract: Pyruvate dehydrogenase complex (PDHC) was purified from bovine kidney with a specific activity of 12–16 μmol of NADH or acetyl‐CoA formed/min/mg protein. The four peptides comprising its three catalytic components were separated by sodium dodecylsulfate‐polyacrylamide gel electrophoresis (SDS‐PAGE). Rabbit antibodies against this highly purified PDHC (anti‐PDHC) exhibited similar binding affinity to the phospho‐PDHC as it did to the PDHC antigen. To test whether there exist brain isozymes of PDHC differing from kidney enzyme, which has been extensively characterized, the PDHCs in bovine brain and kidney were compared using this anti‐PDHC. The PDHC activities in the brain and kidney mitochondrial extracts were inhibited to the same degree by varying amounts of anti‐PDHC. Brain PDHC was precipitated with the anti‐PDHC and resolved by SDS‐PAGE. The four brain PDHC peptides isolated immunochemically with anti‐PDHC had the same sizes as the kidney PDHC peptides. These PDHC peptides from kidney and brain were further compared by their peptide fragment patterns, which were generated by partial proteolysis with Staphylococcus aureus V8 protease or by CNBr and resolved by SDS‐PAGE. The peptide patterns generated with the former method indicated that the α and β peptides of the pyruvate dehydrogenase (E1) component and the peptide of dihydrolipoyl transacetylase (E2) component of kidney PDHC were very similar to the corresponding peptides immunologically isolated from brain. The peptide patterns generated with CNBr further confirmed that the βE1 and E2 peptides of kidney PDHC were similar to the corresponding peptides from brain.
Subcellular Site of Biosynthesis of the Catecholamine Biosynthetic Enzymes in Bovine Adrenal Medulla
Authors: Sabban Esther L. , Goldstein Menek
Published: October 2006

The subcellular site of biosynthesis of the catecholamine biosynthetic enzymes was examined. Free and membrane‐bound polysomes were prepared from bovine adrenal medulla and mRNA was isolated from these polysomes. Both were active in directing cell‐free translations. Immunoprecipitation of cell‐free products with specific antisera localized the biosynthesis of the subunits of tyrosine hydroxylase (TH) (apparent Mr= 61,000) and of phenylethanolamine N‐methyltransferase (PNMT) (apparent Mr= 32,000) on free polysomes, compared with biosynthesis of subunits of dopamine β‐hydroxylase (DBH) (apparent Mr= 67,000) on membrane‐bound polysomes. Cross‐reactivity between translation products was observed. Antibodies for DBH recognized a poly‐peptide with electrophoretic mobility identical to newly synthesized PNMT. However increasing concentrations of antibodies to DBH recognized at most 1/2 of the PNMT formed. The results of this study show the subcellular distribution of the catecholamine synthesizing enzymes is determined by their site of biosynthesis.
High‐Affinity Uptake of Neurotransmitters in Rat Neostriatum: Effects of Aging
Authors: Strong Randy , Samorajski T. , Gottesfeld Zehava
Published: October 2006

High‐affinity uptake of dopamine (DA), glutamate, and γ‐aminobutyric acid (GABA) was determined in crude synaptosomal preparations from neostriatal regions of rats 7, 17, and 27 months of age. Dopamine uptake was highest in rostral neostriatum, but no age‐related differences were detected. On the other hand, the high‐affinity uptake of both GABA and glutamate was increased with age. This may reflect astrocytic hypertrophy or hyperplasia, which have been reported to occur in the neostriatum during the aging process.
Effect of Adrenocorticotropin Administration on β‐Adrenergic Receptor Adaptations in Rat Brain Cerebral Cortex
Authors: Duman Ronald S. , Andree Terrance , Kendall D. A. , Enna S. J.
Published: October 2006

Abstract: It has been reported that adrenocorticotropin (ACTH) administration reduces the time necessary for observing the imipramine‐induced decline in β‐adrenergic receptor binding and function in rat brain frontal cortex. This interaction was examined in the present study following the destruction of the dorsal noradrenergic bundle in an attempt to determine whether the hormone treatment influences pre‐ or postsynaptic activity to facilitate the receptor response. Lesioning completely prevented the decline in β‐receptor binding normally observed following treatment with the drug combination. In fact, the number of cerebral cortical p‐adrenergic receptor binding sites was significantly greater in lesioned animals receiving ACTH than in lesioned controls. Lesioning significantly increased the amount of cyclic AMP produced in response to a saturating concentration of norepineph‐rine, an effect that was not influenced by ACTH treatment. These findings suggest that ACTH administration modifies the norepinephrine‐stimulated cyclic nucleotide system indirectly, perhaps through an action on presyn‐aptic neurons, whereas the effect on receptor recognition site number may be due to a direct action on the postsynaptic cell.
Adrenalectomy of Rats Results in Hypomyelination of the Central Nervous System
Authors: Preston Sheryl L. , McMorris F. Arthur
Published: October 2006

Abstract: The effect of adrenalectomy on CNS myelin accumulation was investigated to determine whether glucocorticoids play a role in regulating myelination. When 14‐day‐old rats were adrenalectomized and sacrificed 7–8 days later, the amount of bulk‐isolated myelin in whole brain, as expressed per gram wet weight of brain or per milligram DNA‐phosphate, was reduced to about 75% that of sham‐operated controls. Both brain weight and DNA content were unchanged by adrenalectomy. Examination of individual brain regions also revealed decreased amounts of myelin in adrenalectomized animals. Brain glycerol 3‐phosphate dehydrogenase specific activity was reduced in adrenalectomized animals to 40–60% that of controls, and serum corticosterone levels were less than 0.6% of control levels. The amount of cerebral myelin in animals adrenalectomized on day 21 and sacrificed 9 days later was not significantly reduced. This suggests a possible role of glucocorticoids during the early period of rapid myelination.
Differential Effects of Bromocriptine on Dopamine and Acetylcholine Release Modulatory Receptors
Authors: Hoffmann Irene S. , Cubeddu Luigi X.
Published: October 2006

Abstract: Rabbit neostriatal slices were prelabeled with [3H]dopamine (DA) and [14C]choline and then superfused. The electrical stimulation‐evoked release of DA and of acetylcholine (ACh) was abolished by 0.33 μM tetrodotoxin and by low calcium concentrations (0.13 μM). Bromocriptine, a selective D2‐DA receptor agonist, inhibited in a concentration‐dependent manner the evoked overflow of DA and ACh, without affecting the basal efflux of both transmitters. The effects of bromocriptine were antagonized by sulpiride, a specific antagonist of D2‐DA receptors. With stimulation at 0.3 Hz and 120 pulses, bromocriptine was eight times more potent in inhibiting the evoked overflow of DA (IC50: 11 nM) than that of ACh (IC50: 83 nM). Stimulations at 3 Hz and 360 pulses markedly reduced the potency of bromocriptine in inhibiting DA and ACh release, and diminished its selectivity for presynaptic receptors. These results indicate that DA receptors that modulate the release of DA and ACh are of the D2 subtype. The greater potency of bromocriptine at pre‐ than at postsynaptic sites suggests that these receptors may be different in quantity and/or quality [D2‐α (presynaptic) versus D2‐β (postsynaptic)]. Finally, marked differences in the potency and efficacy of DA agonist actions on DA and ACh release modulatory receptors are obtained, depending on the parameters of stimulation used.
Interaction of Opiates with Opioid Binding Sites in the Bovine Adrenal Medulla: II. Interaction with K Sites
Authors: Castanas E. , Bourhim N. , Giraud P. , Boudouresque F. , Cantau P. , Oliver C.
Published: October 2006

Abstract: In this study we examined the interaction of opiates with K binding sites in the bovine adrenal medulla. [3H]Ethylketocyclazocine (EKC), [3H]etorphine, and [3H]bremazocine stereoselective bindings were used to assay these interactions. The K sites were found to be heterogeneous: [3H]bremazocine identified with high affinity all subtypes of these sites. [3H]EKC, in the presence of saturating concentrations of [D‐Ala2, D‐Leut]‐enkephalin (DADLE) (5μM), was used to identify K1 sites, on which dynorphin A (1–13) bound with high affinity. Either [3H]EKC or [3H]etorphine in the presence of 5μM DADLE identified the K2 subtype. This subtype was found to interact with β‐endorphin and especially with the octapeptide Met5‐enkephalyl‐Arg6‐Gly7‐Leu8. Furthermore, [3H]etorphine identified in the bovine adrenal medulla a third high‐affinity component, in the presence of 5 μM DADLE. This residual interaction was found to be equally stereoselective and presenting K selectivity. Met5‐enkephalyl‐Arg6‐Phe7 interacted preferentially with this site. The three K subtypes interacted differentially with monovalent (Na+, K+, and Li+) and divalent (Ca2+, Mg2+, and Mn2+) ions by modification of the apparent concentration of the accessible sites and/or by changes of the apparent KD for radioligands. Modifying agents (proteolytic enzymes, thiol‐modifying reagents, and A2‐phospholipase) produced different effects on each subtype of the K site, suggesting a different protein (or protein‐lipid?) composition.
γ‐Aminobutyric Acid Metabolism and Behavioral Effects After Intraventricular Injection of Spermine in Chicks
Authors: Di Giorgio R. M. , De Luca G. , Nisticò G. , Ientile R.
Published: October 2006

Abstract: Effects of intraventricularly injected spermine on behavior and electrocortical activity and γ‐aminobutyric acid (GABA) metabolism after a single dose of 1.13 μmol/animal were studied. Decrease in locomotor activity, sedation or sleep, and electrocortical synchronization that lasted approximately 2 h were observed. In addition spermine caused a significant increase in GABA content in diencephalon and brainstem, 30 min after administration. Concomitantly a significant increase of glutamate decarboxylase (GAD) activity was observed in cerebral hemispheres, diencephalon, and brainstem. Reduction in γ‐aminobutyrate:α‐oxoglutarate aminotransferase (GABA‐T) levels occurred in the diencephalon along with a significant increase of GABA‐T in the brainstem. The present results demonstrate that spermine has the capacity to affect GABA metabolism and are in favor of the suggestion that endogenous polyamines may modulate GABAergic mechanisms.
Altered Axonal Transport of Cytoskeletal Proteins in the Mutant Diabetic Mouse
Authors: Vitadello M. , Filliatreau G. , Dupont J. L. , Hassig R. , Gorio A. , Di Giamberardino L.
Published: October 2006

Abstract: Polypeptides in the motor axons of the sciatic nerve in 120‐day‐old normal and diabetic mice C57BL/Ks (db/db) were labeled by injection of [35S]methionine into the ventral horn of the spinal cord. At 8, 15, and 25 days after the injection, the distribution of radiolabeled polypeptides along the sciatic nerve was analyzed by sodium dodecyl sulfate‐polyacrylamide gel electrophoresis. Four major radiolabeled polypeptides, tentatively identified as actin, tubulin, and the two lightest subunits of the neurofilament triplet, were studied in both diabetic and control mice. In the diabetic animals, the two polypeptides identified as actin and tubulin showed a reduction of average velocity of migration along the sciatic nerve, resulting in a higher fraction of radioactivity in the proximal part of the sciatic nerve, whereas the front of radioactivity (advancing at maximal velocity) moved at a normal rate. In contrast, both the average and maximal velocities of the two neurofilament subunits were slower in the diabetic mice than in the control mice. These results indicate that the axonal transport of the cytoskeletal proteins is differentially affected in the course of diabetic neuropathy, and may suggest that the impairment concerns mainly the proteins carried by the slowest component of axonal transport.