bluuebunny/crossref_metadata_2025 huggingface dataset:
Pyruvate Metabolism in the Brain of Young Rats Intoxicated with Organic and Inorganic Lead
Authors: Regunathan S. , Sundaresan R.
Published: October 2006

Abstract: The activities of lipoyl dehydrogenase, aspartate transaminase, and alanine transaminase, and levels of lactate were estimated in cerebral cortex, cerebellum, and brainstem of rats intoxicated acutely with tetraethyl lead and chronically with lead acetate. A significant inhibition of lipoyl dehydrogenase was observed in both groups of animals, whereas transaminase activities were increased in inorganic lead toxicity. Oxidative decarboxylation and anaplerosis of pyruvate was assessed in brain slices using [l‐14C]pyruvate. Pyruvate dehydrogenase activity was decreased in both organic and inorganic lead toxicity, whereas labelling of aspartate and alanine was increased in inorganic lead toxicity. In studies in vitro, lead acetate showed a more significant effect than tetraethyl lead. The higher anaerobic metabolism in inorganic lead toxicity, as evidenced by increased anaerobic lactate production by brain slices, could either be an adaptive mechanism or be due to the delayed maturation of brain in the developing rat. Such a mechanism does not occur in acute organic lead toxicity, as the compound brings about massive and rapid degenerative changes in brain, resulting in convulsive seizures and death of the animals.
Insensitivity of Ubiquinone Biosynthesis in Glioblastoma Cells to an Epileptogenic Drug, U18666A
Authors: Jeng Ingming , Klemm Nancy , Proctor Barbara
Published: October 2006

Abstract: To investigate the perturbation of ubiquinone biosynthesis by a hypocholesterolemic drug, 3β‐(2‐di‐ethylaminoethoxy)androst‐5‐en‐17‐one hydrochloride (U18666A), we measured the incorporation of radioactive mevalonate, methionine, tyrosine, and 4‐hydroxybenzoic acid into ubiquinone in glioblastoma cells. These four precursors unanimously showed that ubiquinone biosynthesis was not significantly altered by U18666A, which blocked cholesterol biosynthesis at steps beyond mevalonate formation. The fluctuation of the endogenous mevalonate level had little effect on ubiquinone biosynthesis, implying the relative stability of cellular ubiquinone biosynthesis. Furthermore, exogenously added mevalonate did not have an appreciable effect on ubiquinone biosynthesis. The major ubiquinone produced in rat glioblastoma cells was identified as ubiquinone‐9. The mevalonate‐derived products accumulated in the U18666A‐treated cells differed significantly from those reported in a broken cell study, suggesting the existence of delicate mechanisms regulating the formation of cholesterol intermediates.
Immunoblot Identification of 13.5 Kilodalton Myelin Basic Protein in Goldfish Brain Myelin
Authors: Roots B. I. , Agrawal D. , Weir G. , Agrawal H. C.
Published: October 2006

Abstract: Myelin isolated from goldfish brain shows a multilamellar structure with a major dense line and two intraperiod lines. Sodium dodecyl sulfate gel electrophoresis revealed that the protein profile of goldfish brain myelin is distinctly different from that of rat brain myelin. No protein migrating to the position of proteolipid protein or DM‐20 was seen in goldfish myelin. Goldfish acclimated to 5°, 15°, and 30°C showed no qualitative differences in myelin proteins. The 13.5 kD protein in goldfish brain myelin and brain homogenate was intensely immunostained with the antiserum to human basic protein by the immunoblot technique. In contrast, none of the proteins of goldfish myelin were immunostained with an‐tiproteolipid protein serum; however, both proteolipid protein and DM‐20 of rat brain myelin were immunostained. The significance of the synthesis of myelin proteins by astrocytes in the goldfish brain is discussed.
Regionally Selective Inhibition of Cerebral Protein Synthesis in the Rat During Hypoglycemia and Recovery
Authors: Kiessling Marika , Xie Yaxia , Kleihues Paul
Published: October 2006

Regional cerebral protein synthesis was investigated in anesthetized, mechanically ventilated rats during progressive insulin‐induced hypoglycemia and the recovery period following glucose infusion. Polysome profiles from precomatose animals with slow wave/polyspike EEG revealed a slight reduction of polyribosomes and a concurrent increase in monoribosomes, but autoradiographs showed a pattern of l‐[3‐3H]tyrosine incorporation indistinguishable from that of control rats. During the initial 30 min of insulin‐induced isoelectric EEG (“coma”), autoradiographs showed a selective inhibition of protein synthesis in neurons and glial cells of the hippocampus and cerebral cortex, i.e., regions with high susceptibility for the development of hypoglycemic brain damage. Basal ganglia were less affected and areas with low vulnerability (hypothalamus, brainstem, and cerebellum) exhibited a normal pattern of amino acid incorporation. Using a flooding dose of l‐[1‐14C]valine (7.5 mmol/kg; 15 μCi/mmol), the rate of incorporation in cerebral cortex and cerebellum was found to be reduced to 2% and 80% of control values, respectively. Inhibition of protein synthesis was paralleled by a breakdown of polyribosomes and a concomitant increase in ribosomal subunits, indicating a block in peptide chain initiation. After 90 min of isoelectric EEG all brain structures with the exception of hypothalamus and area postrema showed an almost complete lack of amino acid incorporation. Glucose infusion after a 30‐min period of hypoglycemic coma led to a partial restoration of cortical and hippocampal protein synthesis. Within 70–90 min of recovery, l‐[1‐14C]valine incorporation into neocortical and cerebellar proteins amounted to 47% and 125% of fasted controls. The very specialized patterns of regional impairment of cerebral protein synthesis during progressive hypoglycemia may reflect a selective vulnerability of some neuronal cell populations to hypoglycemic cell injury. Alternatively, the striking resistance of hypothalamus, brainstem, and cerebellum may be due to a more efficient glucose uptake at very low blood glucose levels.
DNA Changes in Spinal Cords of Rats with Experimental Allergic Encephalomyelitis
Authors: Smith Marion Edmonds , Somera F. Paul , Saldivar Robert , Massacesi Luca , Trotter Jacqueline
Published: October 2006

DNA levels were measured in the spinal cords of Lewis rats during the development of and recovery from experimental allergic encephalomyelitis (EAE). Spinal cord DNA was first increased 11 days after immunizing the rats with guinea pig myelin and rose to levels four times that of the Freund's adjuvant controls at day 14, then subsided after day 22. Spinal cord DNA was still 150% of control levels 60 days after immunization. These DNA changes were compared with fluctuations in spinal cord acid proteinase in the same animals. Acid proteinase activity in EAE spinal cord increased later than the rise in DNA and attained a level of 170% of control at days 15–17, then subsided. Spinal cord DNA was higher in rats immunized with whole myelin than in those administered equivalent amounts of purified myelin basic protein. Furthermore DNA was higher in spinal cords of rats immunized with a larger dose of myelin (1.0 mg) than with a lower amount (0.5 mg). Various protease inhibitors including pepstatin, nitrophenyl p‐guanidino benzoate, polylysine, and dipropionyl rhein, previously shown to protect Lewis rats against EAE, suppressed the increase of DNA in the spinal cord. Measurement of DNA increases in the spinal cord of EAE animals provides a convenient reproducible measurement of the severity of inflammation in the CNS and provides an objective criterion for assessment of the efficacy of various agents screened as possible therapeutic treatment for multiple sclerosis.
Decreased G4 (10S) Acetylcholinesterase Content in Motor Nerves to Fast Muscles of Dystrophic 129/ReJ Mice: Lack of a Specific Compartment of Nerve Acetylcholinesterase?
Authors: Gisiger Victor , Stephens Heather R.
Published: October 2006

Abstract: Acetylcholinesterase (AChE; EC 3.1.1.7) acivity and the distribution of its molecular forms were studied in the nervous system of normal and dystrophic 129/ReJ mice, including the sciatic‐tibial nerve trunk and motor nerves to slow‐ and fast‐twitch muscles. In normal mice, motor nerves to the slow‐twitch soleus exhibited a low AChE activity together with a low level of G4 (10S form) as compared with nerves of the predominantly fast‐twitch plantaris and extensor digitorum longus. In contrast, in dystrophic mice, the AChE activity as well as the G4 content of nerves to the fast‐twitch muscles were low, displaying an AChE content similar to that of the nerve of the soleus muscle. In the sciatic‐tibial nerve trunk, the AChE activity decreased along the nerve in an exponential mode, at rates that were similar in both conditions. However, in dystrophic mice, the AChE activity was reduced throughout the nerve length by a constant value of approximately 180 nmol/h/mg protein. Further analyses indicated that AChE in this nerve trunk was distributed among two compartments, a decaying and a constant one. The decay involved exclusively the globular forms. The activity of A12 (16S form) remained constant along the nerve and was similar in both normal and dystrophic mice. In addition, according to the equation describing the decay of AChE, the reduction in enzymatic activity observed in the dystrophic mice affected mainly G4 in the constant compartment. Brain, spinal cord, sympathetic ganglia, and serum, which were also examined, showed no remarkable differences between the two conditions in their G4 content. The AChE abnormalities that we found in nervous tissues of 129/ReJ dystrophic mice were confined to the motor system.
Comparison of Two Superfusion Systems for Study of Neurotransmitter Release from Rat Cerebral Cortex Slices
Authors: Korpi E. R. , Oja S. S.
Published: October 2006

Abstract: Depolarization‐induced release of [3H]γ‐aminobutyric acid (GABA) and [3H]noradrenaline (NA) from rat cerebral cortex slices was studied in two superfusion systems: one with stationary and the other one with continuously shaken slice compartments. Calcium‐dependent depolarization‐induced release of GABA and NA could be demonstrated only with shaken slices. GABA, but not NA, could also be released by high K + media and veratridine from stationary slices. Synaptic transmitterreleasing mechanisms are apparently damaged in stationary slices, possibly due to impaired energy metabolism.
Photoaffinity Labelling of the Benzodiazepine Receptor Compromises the Recognition Site but Not Its Effector Mechanism
Authors: Brown Chloë L. , Martin Ian L.
Published: October 2006

Abstract: When subjected to ultraviolet light flunitrazepam could be irreversibly attached to brain membrane preparations such that the affinity of the benzodiazepinereceptor for certain ligands, such as diazepam, becomes markedly reduced. However, the apparent affinity for diazepam is increased by the addition of y‐aminobutyricacid and sodium chloride, to the same extent in this membrane preparation as in membranes not so pretreated. This observation suggests that photoaffinity labelling ofthe benzodiazepine receptor with flunitrazepam modifiesthe recognition characteristics of the receptor but not itseffector mechanism.
Determination of Daily Variations of Brain 5‐Hydroxytryptamine and Dopamine Turnovers and of the Clearance of Their Acidic Metabolites in Conscious Rats by Repeated Sampling of Cerebrospinal Fluid
Authors: Hutson P. H. , Sarna G. S. , Curzon G.
Published: October 2006

Abstract: Central 5‐hydroxytryptamine (5‐HT) and dopamine (DA) turnovers were estimated simultaneously in conscious freely moving rats kept on a 12‐h dark/l2‐h light cycle by sampling cisternal CSF of each animal before and after giving probenecid and determining the accumulation of the acidic metabolites of the two amines. The turnovers of both transmitters and the clearances of their acid metabolites from the brain were shown to be significantly greater during the dark (red light) period than during the white light period.
Spontaneous and Experimental Neuritis and the Distribution of the Myelin Protein P2 in the Nervous System
Authors: Kadlubowski M. , Hughes R. A. C. , Gregson N. A.
Published: October 2006

Abstract: The P2 contents of nervous tissues from the human, rabbit, guinea pig, and Lewis rat were measured by radioimmunoassay. The ventral spinal roots contained more P2 than any other tissue. Human dorsal roots and peripheral nerves contained 41–65% of the amount in human ventral roots. Human olfactory and optic nerves and brain contained 1.1–2.7%, spinal cord, 2.8%, cranial nerve VIII, 11%, and cerebral grey matter, 0%. The relative amounts in the rabbit nervous system were similar except that the spinal cord contained 20% of the amount in the ventral roots. Qualitative estimates in the guinea pig showed that the spinal roots and peripheral nerves contained more P2 than the spinal cord, and that none was present in the brain. In the Lewis rat, P2 could be detected in the spinal roots and peripheral nerves but not in the CNS. The distribution of P2 in the human nervous system parallels the incidence and severity of lesions in acute polyradiculoneuritis. It also explains the absence of any lesions in the CNS when experimental allergic neuritis is induced in the Lewis rat.
Growth Kinetics, Cell Shape, and the Cytoskeleton of Primary Astrocyte Cultures
Authors: Goldman James E. , Chiu Fung‐Chow
Published: October 2006

Abstract: We examined correlations among growth kinetics, cell shape, and cytoskeletal protein content in rat astrocytes grown in primary culture. Cell suspensions from brains of newborn rats were seeded at densities from 0.2 to 3 × 105/cm2. At initial densities above 1 × 105 the population increased to reach confluency by 10–12 days, after which cell number remained stable for many weeks. At low initial densities, 0.2–0.4 × 105/cm2, cells did not increase in number. Final density increased with increasing plating densities. High‐density cells had small perikarya and several long cytoplasmic processes; low‐density cells appeared flat and polygonal. All cultures were almost entirely astrocytic, as judged by immunofluorescent staining with antiserum against glial fibrillary acidic protein (GFAP). Cytoskeletal proteins were analyzed by gel electrophoresis after extraction from cells with nonionic detergent. Relative amounts of the proteins differed, in that low‐density cells contained large amounts of cytoskeletal actin relative to the intermediate filament (IF) proteins vimentin and GFAP, whereas high‐density cells contained relatively less actin and more IF proteins. Such differences in cytoskeletal proteins between the high‐ and low‐density cultures were mirrored in the relative rates of synthesis of the cytoskeletal proteins. In the low‐density cells amino acid incorporation into cytoskeletal‐associated actin was more active than that into the IFs, whereas in the high‐density cells higher rates of IF protein synthesis were observed.
Histidine Transport into Rat Brain Synaptosomes
Authors: Hegstrand Linda R. , Simon Jay R.
Published: October 2006

Abstract: Histidine transport and metabolism in rat brain synptosomes were investigated to study the possible role of histidine uptake in the synthesis of the putative neurotransmitter histamine (HA). Histidine uptake was found to be regionally distributed and temperature sensitive and was not totally independent of sodium or possium ions. Transport was inhibited by metabolic inhibitors, as well as by promethazine and quinacrine. A number of other HA‐related agents and several histidine metabolites had no effect. Kinetic analyses of histidine transport revealed the presence of both high‐ and lowaffinity systems in cerebral cortex. Histidine uptake increased following preexposure of synaptosomes to depolarizing concentrations of potassium. This effect was dependent on the presence of calcium ions during the preincubation. No newly formed [3H]HA was detectable in rat brain synaptosomes following [3H]histidine transport. Lesions of the medial forebrain bundle did not alter histidine uptake in the hippocampus or cerebral cortex. Ontogenic studies indicated that the histidine uptake system developed rapidly and reached a peak during postnatal days 12–17. Overall, the present findings do not support a role for histidine transport in the regulation or maintenance of neurotransmitter pools of HA in rat brain.
Tryptamine Concentrations in Areas of 5‐Hydroxytryptamine Terminal Innervation After Electrolytic Lesions of Midbrain Raphe Nuclei
Authors: Juorio A. V. , Greenshaw A. J.
Published: October 2006

Abstract: The possible existence of tryptamine‐containing neurons originating in the midbrain raphe is suggested by several reports of tryptamine‐mediated responses to electrical stimulation of the raphe nuclei. To assess this hypothesis, we have investigated the effects of electrolytic lesions of the median and dorsal raphe nuclei on striatal, hypothalamic, and hippocampal concentrations of tryptamine, 5‐hydroxytryptamine (5‐HT), and 5‐hydroxyindoleacetic acid. In addition, the rat striatal tryptophan concentrations were also determined. No changes in the concentrations of tryptamine were observed at 1 or 2 weeks after lesioning the dorsal and median raphe nuclei, at which time the other 5‐hydroxyindoles were markedly reduced; furthermore, no reductions were observed in tryptamine concentrations in the striatum, hypothalamus, or hippocampus of rats pretreated with a monoamine oxidase inhibitor. The only change observed in these rats was a limited increase in striatal tryptamine and tryptophan observed at 1 day after lesioning. The results indicate that tryptamine concentration is independent of the integrity of 5‐HT‐containing neurons of the midbrain raphe nuclei. Furthermore, if tryptamine‐containing neurons that have terminal projections to the striatum, hypothalamus, and hippocampus exist, their cell bodies are located in regions outside the dorsal and median raphe nuclei. Another possibility could be that tryptamine is located in glial cells.
Indoleamine‐Sensitive Adenylate Cyclase in Rabbit Retina: Characterization and Distribution
Authors: Blazynski Christine , Ferrendelli James A. , Cohen Adolph I.
Published: October 2006

Abstract: Previous histological, electrophysiological, and biochemical reports have addressed the hypothesis that serotonin functions as a neurotransmitter in mammalian retinas. We have tested the effect on the levels of cyclic AMP of the application of exogenous serotonin, 5‐methoxytryptamine, melatonin, and 5‐methoxydimethyl‐tryptamine to isolated, incubated rabbit retinas. All indoleamines tested significantly elevated intracellular levels of cyclic AMP in both light‐ and dark‐adapted, incubated, intact retinas, provided a phosphodiesterase inhibitor was present. In homogenates of rabbit retina, all indoleamines tested also markedly increased adenylate cyclase activity over basal levels. Maximal activity was observed with 50 μM indoleamine; addition of GTP augmented this increase. The increase in enzyme activity persisted in the presence of known antagonists of dopamine and serotonin 5‐HT2‐receptors, but was blocked by the mixed 5‐HT1, 5‐HT2‐antagonist lysergic acid diethylamide. The retinal locations of this response have also been identified using layer microdissection techniques on freeze‐dried samples obtained from rabbit eyecups suprafused with indoleamine plus phosphodiesterase inhibitor. Cyclic AMP levels were measured in discrete retinal layers of both light‐ and dark‐adapted suprafused eyecups, and increased levels were observed primarily in the inner and outer plexiform layers, which contain the synapses of the retinal neurons.
Quantitative Autoradiography of [3H]Indalpine Binding Sites in the Rat Brain: I. Pharmacological Characterization
Authors: Bénavidès J. , Savaki H. E. , Malgouris C. , Laplace C. , Margelidon C. , Daniel M. , Courteix J. , Uzan A. , Guérémy C. , Le Fur G.
Published: October 2006

Abstract: The binding of [3H]indalpine {4‐[2‐(3‐indolyl)]ethyl piperidine} to slide‐mounted sections of rat brain has been characterized. This 5‐hydroxytryptamine (5‐HT) uptake blocker binds to sections with high affinity (KD∼ 1 nM). The binding is saturable, and can be displaced by the addition of clomipramine (1 μM). Other drugs inhibiting the uptake of 5‐HT also have the capacity to inhibit the binding of [3H]indalpine. A significant correlation (r = 0.86) was found between the capacity of these compounds to inhibit the uptake of 5‐HT and their potencies as inhibitors of [3H]indalpine binding. Binding was Na+‐ and Cl−dependent and was inhibited competitively by 5‐HT. Furthermore, electrolytic lesions of the dorsal raphe or medial forebrain bundle, which cause a degeneration of 5‐HT cell bodies and fibers, respectively, resulted in a 30–40% reduction in the binding of [3H]indalpine. [3H]Indalpine binds to the 5‐HT uptake recognition sites in a different manner from imipraminelike compounds.
Metabolic Turnover of Fatty Acids and Acylglycerols in Rat Sciatic Nerve
Authors: Yao Jeffrey K.
Published: October 2006

Abstract: To explain the discrepancy between the low level and high metabolic activity of endoneurial free fatty acids (FFAs) and triacylglycerol (TG), levels of de novo synthesized FFA and acylglycerols were measured in rat sciatic endoneurium at various intervals after endoneurial microinjection of [14C]acetate. Soon after injection (< 10 min), the [14C]acetate was metabolized to FFA and incorporated into diacylglycerol (DG), TG, sterols, ceramides, and various phospholipids. The proportions of 14C‐labeled FFA, DG, TG, and ceramides to total 14C‐labeled lipids decreased, whereas those of phospholipids and cerebrosides increased with time after injection. These findings suggest that rapid turnover of FFA and TG may contribute to their low level in sciatic endoneurium. The de novo synthesized fatty acids were largely incorporated into phosphatidylcholine (∼50% of total 14C‐labeled phospholipids), probably via the cytidine nucleotide pathway using 1,2‐DG as a metabolic intermediate. Hydrolysis of [14C]phosphatidylcholine revealed that fatty acids were labeled at both the C‐1 (∼43%) and C‐2 (∼57%) positions. On the other hand, a temporal association between decreased amounts of 14C‐label in ceramides and increased amounts of 14C‐label in sphingomyelin and galactocerebrosides supports the hypothesis that peripheral nerve galactocerebroside is derived, in vivo, from ceramide via acylation of sphingosine. This exclusive labeling of endoneurial lipids by endoneurial microinjection of labeled precursor provides a unique model for studying synthesis and metabolic turnover of membrane lipids in experimental neuropathies.
Nature of Extrasynaptosomal Accumulation of Endogenous Adenosine Evoked by K+ and Veratridine
Authors: MacDonald W. F. , White T. D.
Published: October 2006

Abstract: When rat brain synaptosomes were incubated for 10 min at 37°C, basal accumulation of adenosine in the medium was 66 pmol/mg of protein. An elevated K+ level (24 mM) evoked an additional accumulation of 200 pmol/mg of protein, and 50 μM veratridine evoked 583 pmol of adenosine accumulation/mg of protein. K+‐ and veratridine‐evoked accumulation of adenosine did not arise from microsomal or mitochondrial contaminants of the synaptosomal preparation, because purified microsomes and mitochondria did not exhibit evoked accumulation of adenosine in the medium. K+‐evoked accumulation of extrasynaptosomal adenosine was Ca2+‐dependent, whereas veratridine‐evoked accumulation of adenosine was increased in Ca2+‐free medium. In the presence of α,β‐methylene ADP and GMP, which inhibit ecto‐5′‐nucleotidase, conversion of added ATP and AMP to adenosine was inhibited by 90% in synaptosomal suspensions. However, inhibition of ecto‐5′‐nucleotidase only reduced basal extrasynaptosomal accumulation of adenosine by 74%, veratridine‐evoked accumulation of adenosine by 46%, and K+‐evoked accumulation by 33%. Most of the basal accumulation of extrasynaptosomal adenosine appears to be derived from released nucleotide, probably ATP, but about half of the veratridine‐evoked accumulation of adenosine and most of the K+‐evoked accumulation may arise from adenosine released in its own right, rather than from a released nucleotide.
Temporal Expression of Mouse Glial Fibrillary Acidic Protein mRNA Studied by a Rapid In Situ Hybridization Procedure
Authors: Lewis Sally A. , Cowan Nicholas J.
Published: October 2006

Abstract: A rapid and sensitive in situ hybridization technique is described for the detection of mRNA sequences in 6–8‐μm cryostat sections. The method incorporates the use of α‐thio‐35S‐labelled nucleoside triphosphates for the generation of high‐specific‐activity DNA probes and a high‐stringency washing procedure that virtually eliminates background without unduly compromising histological integrity. Whereas signal resolution is less than that observed using 3H probes, 35S‐labelled probes are well‐suited for experiments where resolution at the cellular level is required. The method has been applied to a study of the developmental regulation of glial fibrillary acidic protein (GFAP) mRNA expression in developing mouse brain. GFAP‐specific sequences are first detectable after the second postnatal day, and thereafter rise to a level that is maintained throughout development and into adulthood. The distribution of GFAP‐encoding sequences broadly reflects the known distribution of astrocytes, but the levels of mRNA within these cells vary by a surprisingly large amount depending on their location. For example, in adult animals, the astrocytes of the glial limitans contain an abundance of GFAP‐specific mRNA that is higher than corresponding levels in astrocytes in the cerebellar white matter, whereas these cells in turn contain considerably more GFAP‐specific mRNA than astrocytes in the gray matter of the cerebrum. Unexpectedly, parallel RNA blot transfer experiments show the existence of some GFAP‐encoding mRNA size heterogeneity that is restricted to the first postnatal week.
Dibutyryl‐Cyclic GMP Stimulation of Ca2+‐ATPase Activity in Rat Brain Synaptic Membranes
Authors: Stauderman Kenneth A. , Jones David J. , Ross David H.
Published: October 2006

Abstract: The effects of dibutyryl cyclic AMP (dbcAMP) and dibutyryl cyclic GMP (db‐cGMP) were tested on Ca2+‐ATPase, Mg2+‐ATPase, and (Ca2++ Mg2+)‐ATPase activities in lysed synaptosomes prepared from whole rat brains (minus cerebellum). At concentrations from 0.1 to 2.0 mM, db‐cGMP produced a selective, concentration‐dependent increase in Ca2+‐ATPase activity. Both db‐cGMP and db‐cAMP slightly reduced Mg2+‐ATPase activity, whereas neither compound had concentration‐dependent effects on (Ca2++ Mg2+)‐ATPase activity. These findings suggest that the Mg2+‐independent, Ca2+‐ATPase activity in rat brain is regulated by a cyclic GMP‐dependent process. Further, the data provide evidence that the Ca2+‐ATPase activity in lysed synaptosomal membranes represents an enzyme that is distinguishable from both the Mg2+‐ and (Ca2++ Mg2+‐ATPase.
Reduced Axoplasmic Somatostatin Transport in Hypothyroid Rats
Authors: Rasool C. G. , Bradley W. G. , Reichlin S.
Published: October 2006

Abstract: The effect of hypothyroidism on neuronal function was studied by measuring axoplasmic transport of immunoreactive somatostatin in rat sciatic nerve by the ligation technique. Accumulation of immunoreactive somatostatin proximal to a ligature was linear up to 8 h in normal, in thyroidectomized, and in parathyroidectomized rats. The transport rate was decreased by 38% in thyroidectomized rats as compared to normal rats and was unchanged in parathyroidectomized rats. Sciatic nerve content of somatostatin in hypothyroid rats did not differ from control. Reduced accumulation of immunoreactive somatostatin in hypothyroid rats may be due to a decrease in somatostatin synthesis or in axoplasmic transport, or to an increase in the degradation rate of the peptide.
Differential Distribution of Carboxypeptidase‐Processing Enzyme Activity and Immunoreactivity in Membrane and Soluble Components of Chromaffin Granules
Authors: Hook Vivian Y. H.
Published: October 2006

Abstract: A comparison of carboxypeptidase‐processing enzyme activity and immunoreactivity showed that they were differentially distributed in soluble and membrane components of bovine adrenal medulla chromaffin granules. The majority of enzyme activity (80% of total activity in the granules) was present in the soluble fraction, but the number of enzyme molecules was distributed equally between the soluble and membrane fractions. When equivalent amounts of carboxypeptidase enzyme (ng of immunoreactivity) in each fraction were compared, the carboxypeptidase in the soluble component appeared to be five to six times more active than the membranebound form of the enzyme. The soluble and membrane components of the granules may represent populations of enzyme at different states of activation. This finding could have important implications for the regulation of the carboxypeptidase‐processing enzyme.
Tryptamine, a Substrate for the Serotonin Transporter in Human Platelets, Modifies the Dissociation Kinetics of [3H]Imipramine Binding: Possible Allosteric Interaction
Authors: Segonzac A. , Raisman R. , Tateishi T. , Schoemaker H. , Hicks P. E. , Langer S. Z.
Published: October 2006

Abstract: Tricyclic antidepressants and nontricyclic serotonin (5‐hydroxytryptamine) uptake blockers monophasically inhibit [3H]imipramine binding in human platelets. Similarly, serotonin and tryptamine inhibit the binding of [3H]imipramine in the low micromolar range and with a pseudo‐Hill coefficient near unity. Dissociation of the [3H]imipramine receptor complex in the presence of uptake inhibitors follows first‐order kinetics with a half‐life of approximately 60 min. Although serotonin and tryptamine do not decrease [3H]imipramine binding when added under equilibrium conditions, simultaneous addition of serotonin or tryptamine with serotonin uptake inhibitors decreases the rate of ligand‐receptor dissociation in a concentration‐dependent manner. These data suggest a common site of action for serotonin, which is the substrate of the transporter system, and of tryptamine, its nonhydroxylated analog. This hypothesis is supported by the identification of a high‐affinity (Km= 0.55 μM), saturable, and temperature‐dependent uptake of [3H]tryptamine in human platelets. Uptake of [3H]tryptamine was inhibited potently by imipramine and nontricyclic serotonin uptake inhibitors with a potency similar to that observed for [3H]serotonin uptake. These data support the hypothesis that in platelets, [3H]imipramine, tricyclic, and nontricyclic serotonin up‐take inhibitors bind to a common recognition site that is associated with the serotonin transporter but that differs from the substrate recognition site of the carrier through which serotonin and tryptamine exert a heterotropic allosteric modulation on [3H]imipramine binding.
Denervation Supersensitivity of the Rat Pineal to Norepinephrine‐Stimulated [3H]Inositide Turnover Revealed by Lithium and a Convenient Procedure
Authors: Zatz Martin
Published: October 2006

Abstract: A convenient procedure for the assay of myo‐[2–3H(N)]inositol ([3H]inositol) metabolites in cells or small amounts of tissue was developed. The procedure is a composite of modifications of published methods. After preincubation with [3H]inositol, rat pineal glands were disrupted in an acidified organic solvent mixture. Lipids were separated from the hydrophilic products and precursor using Sephadex G‐25 columns and further analyzed by TLC. Hydrophilic products were further analyzed by anion‐exchange column chromatography using Dowex AG1‐X8 (formate form). In the presence of lithium, increases in inositol phosphates consequent to stimulation of the glands by norepinephrine were apparent within 10 min. The response in denervated glands was considerably greater than in intact pineals.
Acetylcholine Receptors in Small Cell Carcinomas
Authors: Cunningham Julie M. , Lennon Vanda A. , Lambert Edward H. , Scheithauer Bernd
Published: October 2006

Abstract: The presence of muscarinic and nicotinic acetylcholine receptors (m‐ and nAChR, respectively) in small cell carcinomas (SCC) of the lung was assessed by measurement of specific binding of (—)[3H]quinuclidinyl benzilate ([3H]QNB) and 125I‐α‐bungarotoxin, respectively. Of five SCC studied, four were originally derived from patients with the Lambert‐Eaton myasthenic syndrome, an autoimmune disease of neuromuscular transmission, and one was from a patient without evidence of neurological disease. There was no evidence of nAChR, but all tumors bound (‐)[3H]QNB in a saturable and specific manner. Dissociation constants derived from saturation isotherms ranged between 0.13 and 0.3 nM and maximum binding capacities between 35.4 and 181.7 fmol/mg protein (mean values for the five SCC). Competition studies revealed a pharmacological profile consistent with previous descriptions of mAChR. Competition by pirenzepine revealed only one class of binding sites, that with a relatively low affinity for pirenzepine. In the presence of the guanine nucleotide analogue 5′‐guanylyl imidodiphosphate, a decrease in affinity of the mAChR of SCC for oxotremorine was observed, with an increase in the pseudo‐Hill coefficient, but there was no change in the binding of atropine. The expression on SCC of mAChR, apparently of the M2 subclass, represents yet another neural differentiation marker of SCC. It is noteworthy that the expression of this marker is not restricted to patients with an autoimmune paraneoplastic syndrome involving cholinergic neurons.
High‐ and Low‐Affinity Binding Components for [3H]Imipramine in Rat Cerebral Cortex
Authors: Conway P. G. , Brunswick D. J.
Published: October 2006

Abstract: The present study demonstrates that [3H]imipramine binds to both high‐ and low‐affinity imip‐ramine binding components on membranes prepared from rat cerebral cortex. Scatchard and computer analyses of saturation experiments using a wide range of [3H]imipramine concentrations (0.5 nM‐50 nM) revealed the presence of two binding components. Inhibition experiments in which membranes were incubated with [3H]imipramine and various concentrations of unlabelled imipramine gave shallow inhibition curves with a Hill coefficient of 0.60 ± 0.04. When dissociation rates of imipramine were studied, biphasic dissociation curves were obtained with apparent half‐times of dissociation of 2.5 ± 0.4 min and 18.5 ± 2.5 min. Thus analysis of saturation, competition, and dissociation experiments indicate that [3H]imipramine binds to low as well as high‐affinity binding sites in rat cortex.
Parvalbumin in Human Brain
Authors: Berchtold Martin W. , Celio Marco R. , Heizmann Claus W.
Published: October 2006

Abstract: Parvalbumin was isolated from human cerebral cortex and biceps and triceps muscles by HPLC. The immunological properties of the human protein and the mobility in two‐dimensional polyacrylamide gels were similar to that of parvalbumin isolated from the muscles of rat, mouse, rabbit, and chicken. The tryptic peptide maps of the human parvalbumin, however, differed considerably from all other parvalbumins, indicating a distinct primary structure. The immunolabeled cells in the hippocampus of the human brain were of different sizes and forms; they occurred in all subfields and probably represent interneurons.
5‐Methoxytryptoline, a Competitive Endocoid Acting at [3H]Imipramine Recognition Sites in Human Platelets
Authors: Segonzac A. , Schoemaker H. , Tateishi T. , Langer S. Z.
Published: October 2006

Abstract: 5‐Methoxytryptoline potently inhibits [3H]‐imipramine binding to membranes from the cerebral cortex and platelets. Since 5‐methoxytryptoline, which appears to occur endogenously with particularly high levels in the human pineal gland, also inhibits 5‐hydroxy‐tryptamine (5‐HT, serotonin) uptake, it should be considered as a putative endogenous ligand modulating 5‐HT transport. As the 5‐HT transporter complex comprises the imipramine and the substrate recognition sites, which interact allosterically, it was essential to define the mechanism of inhibition of [3H]imipramine binding by 5‐methoxytryptoline. Human platelets show an active and saturable uptake of 5‐HT and tryptamine. The uptake of both substrates appears to be mediated by the same carrier and it is inhibited by 5‐methoxytryptoline at submicromolar concentrations. 5‐HT and tryptamine inhibit [3H]imipramine binding in human platelets with a Hill slope for inhibition close to unity and IC70 values of 3,265 and 3,475 nM, respectively. This inhibition is, however, not competitive because both 5‐HT and tryptamine significantly decrease the rate of [3H]imipramine‐receptor dissociation. Although 5‐methoxytryptoline potently inhibits [3H]imipramine binding (IC50= 44 nM) in human platelets with a Hill slope of unity, it does not affect the receptor‐ligand dissociation rate of [3H]imipramine even at concentrations up to 100 μM. The present experiments show that 5‐methoxytryptoline, in spite of its chemical similarity to the indoleamine transporter substrates, interacts with the imipramine receptor through a mechanism of competitive inhibition. This conclusion is supported by a selective effect of 5‐methoxytryptoline on the Kd of [3H]imipramine binding. These data are compatible with the hypothesis that 5‐methoxytryptoline may be an endogenous modulator that interacts competitively with the imipramine receptor associated with the 5‐HT uptake complex.
A Low Repeat Length in Oligodendrocyte Chromatin
Authors: Di Liegro Italia , Salemi Giuseppe , Cestelli Alessandro
Published: October 2006

Abstract: The behavior of oligodendrocyte chromatin after micrococcal nuclease digestion of nuclei was assayed in brains of rats of four different ages. During oligodendrocyte differentiation, a decreasing sensitivity of the chromatin to enzymatic attack was observed. On the other hand, the nucleosomal repeat length showed a slight tendency to increase during development. It is worth noting that even the highest values reported here for “oligodendrocyte’ chromatin repeat lengths are significantly lower than 200 base pairs, the value previously reported by others for “non‐astrocytic glia.”
Is Propionylcholine Present in or Synthesized by Electric Organ?
Authors: Kosh Joseph W. , Whittaker Victor P.
Published: October 2006

Abstract: When small blocks comprising four columns of electrocytes were excised from electric organs of Torpedo marmorata after stimulation in vivo via the electric lobe at 1 Hz for 1 h and allowed to recover at 20–22°C for several hours in medium containing 100 μM d4 choline and 500 μM propionate, small quantities of propionylcholine amounting to no more than 1% of the endogenous acetylcholine of the tissue could be detected in tissue extracts by gas chromatography‐mass spectrometry (GCMS). Kinetic studies demonstrated that there was no nonexchangeable propionylcholine in the tissue and in the absence of added propionate, propionylcholine levels were < 0.2% of tissue acetylcholine. Vesicular propionylcholine amounted to < 0.5% of vesicular acetylcholine and the distribution of d0 and d4 propionylcholine suggested that an appreciable proportion (up to one‐third) of this could be an artifact of preparation for GCMS determinations. Propionylcholine formation during extraction and demethylation of an artificial mixture of acetylcholine, choline, and propionate was indeed detected. It is concluded that propionylcholine has no significance as an endogenous or as a false transmitter at this terminal, in conformity with the work of Sheridan et al. [Z. Zellforsch.74, 281–307 (1966)] but in contrast to the report of O]Regan [J. Neurochem.39, 764–772 (1982)].
Structure of Catecholamine Secretory Vesicles from PC12 Cells
Authors: Wagner John A.
Published: October 2006

Abstract: Catecholamine secretory organelles were partially purified from PC12 cells. Measurement of the sedimentation coefficient (540S in 0.32 M sucrose), density in an isoosmotic gradient (1.139 g/cm), and density in an isoosmotic gradient using D2O as a solvent (1.205 g/cm3) have allowed us to calculate the molecular weight (1.17 × 109 daltons), radius (74 nm), and water content (62% vol/vol) of the secretory vesicle. The vesicle appears to contain ATP, but the molar ratio of 3,4‐dihydroxyphenylethylamine (dopamine) to ATP in the particles is high (16.5) and the ATP was frequently asymmetrically distributed in the vesicle fraction. The particle behaves like a true secretory particle in that the dopamine content of the particle is increased by pargyline, diminished by depolarization, and abolished by reserpine. Sequential purification of PC12 lysates on controlled pore glass columns and isoosmotic Ficoll gradients produced a 20–30‐fold purification, but this enrichment is not sufficient to produce a homogeneous population of vesicles. An 82,000‐dalton protein copurifies with secretory granules and appears to be the major secreted protein. At this stage of purification this single protein makes up about 30% of the protein in the vesicle‐containing fractions and so the vesicles must be approaching homogeneity.
Multiple [35S]t‐Butylbicyclophosphorothionate Binding Sites in Rat and Chicken Cerebral Hemispheres
Authors: Tehrani Mohammad H. Jalilian , Clancey Constance J. , Barnes Eugene M.
Published: October 2006

Abstract: Specific binding of [35S]t‐butylbicyclophosphorothionate (TBPS) to membranes from cerebral hemispheres of adult rat and chicken was determined over a range of radioligand concentrations from 0.25 to 500 nM. Scatchard plots of these data were curvilinear and non‐linear regression analysis indicated binding to two sites that differ in affinity. For rat cerebrum, KD(1)= 1.15 nM, Bmax(1) 0.085 pmol/mg; KD(2)= 232 nM, Bmax(2)= 16.9 pmol/mg. For chicken cerebrum, KD(1)= 1.39 nM, Bmax(1)= 0.111 pmol/mg; KD(2)= 166 nM, Bmax(2)= 17.6 pmol/mg. This multiplicity of [35S]TBPS binding was further confirmed when unlabeled TBPS or picrotoxinin displaced radioligand. The displacement curves were biphasic and yielded Hill coefficients from 0.65 to 0.70. These displacement curves were also resolved into two components with distinct IC50 values for unlabeled TBPS (rat, 1.55 and 271 nM; chicken, 2.40 and 224 nM). The IC50 values were similar to the dissociation constants obtained from equilibrium binding measurements.
Neurochemical and Histochemical Characterization of Neurotoxic Effects of 1‐Methyl‐4‐Phenyl‐1,2,3,6‐Tetrahydropyridine on Brain Catecholamine Neurones in the Mouse
Authors: Hallman H. , Lange L. , Olson L. , Strömberg I. , Jonsson G.
Published: October 2006

Systemic administration of 1‐methyl‐4‐phenyl‐1,2,3,6‐tetrahydropyridine (MPTP) caused a rapid and long‐lasting reduction of both 3,4‐dihydroxyphenylalanine (dopamine, DA) and noradrenaline (NA) in mouse brain, as observed histo‐ and neurochemically. The depleting effects were more pronounced after repeated MPTP administration and the most marked reductions were observed after 2 × 50 mg MPTP/kg s.c., when DA in striatum and NA in frontal cortex were reduced by > 90% 1 week after MPTP. Mice with such catecholamine depletions were markedly sedated and almost completely immobilized. The behavioural syndrome after MPTP resembled that seen after reserpine, a monoamine‐depleting drug. MPTP also caused a long‐lasting reduction of catecholamine uptake in striatal DA and cortical NA nerve terminals and reduced tyrosine hydroxylase activity in these regions. There was no evidence that MPTP caused any marked DA and NA cell body death. MPTP given acutely transiently elevated serotonin levels. The results are compatible with a neurotoxic action of MPTP on both DA and NA nerve terminals. The nigro‐striatal DA and the locus coeruleus NA neurone systems appeared to be most susceptible. Synthesis and utilization of residual striatal DA and cortical NA were increased, as often observed in partially denervated monoamine‐innervated brain regions. Both DA and NA showed a gradual recovery, which took months to become complete and may have been related to a regrowth of catecholamine nerve terminals.
Effects of Guanyl Nucleotides on [3H]Flunitrazepam Binding to Rat Hippocampal Synaptic Membranes: Equilibrium Binding and Dissociation Kinetics
Authors: Fung S.‐C. , Fillenz M.
Published: October 2006

The effects of guanyl nucleotides on the binding of [3H]flunitrazepam to rat hippocampal synaptic membranes were studied. In equilibrium binding studies, γ‐amino‐n‐butyric acid (GABA) increased and GTP decreased the binding affinity of [3H]flunitrazepam; GTP also caused a decrease in binding capacity. The effect, however, is variable. In studies of the dissociation kinetics of [3H]flunitrazepam using diazepam and the antagonist Ro 15–1788 as the displacers, there was evidence of two dissociation rate constants. GTP increased both the fast and slow‐dissociation rate constants and increased the ratio of the slow‐dissociation binding state. The effect of GTP was mimickled by its nonhydrolyzable analogue 5′‐guanylylimidodiphosphate but not by ATP and occurred when diazepam, but not when Ro 15–1788, was used as the displacer. GABA antagonized the effect of GTP on the dissociation of [3H]flunitrazepam. The nature of the benzodiazepine recepter, its actions, and the possible role of cyclic AMP as a second messenger are discussed.
Dopamine Autoreceptor Stimulation Increases Protein Carboxyl Methylation in Striatal Slices
Authors: Wolf Marina E. , Roth Robert H.
Published: October 2006

We have investigated the possibility that protein carboxyl methylation is involved in coupling dopamine autoreceptor stimulation to intracellular events such as inhibition of dopamine synthesis or release. The dopamine agonists apomorphine and TL‐99 were found to stimulate methyl ester formation in striatal slices preloaded with [3H]methionine. The stimulatory effects of apomorphine were dose‐dependent, were not due to changes in [3H]methionine uptake or S‐[3H]‐adenosyl‐methionine formation, and were blocked by the stereospecific dopamine antagonist (+)‐butaclamol. Stimulation of methyl ester formation by dopamine agonists is readily observed only when slices are prepared from rats pretreated with reserpine to deplete endogenous brain catecholamines. This suggests that in slices prepared from normal rats endogenous dopamine (DA) released during slice preparation and incubation masks the effects produced by exogenously administered dopamine agonists on protein carboxyl methylase (PCM) activity. Additional experiments suggested that the effects of apomorphine were mediated via an interaction with DA autoreceptors rather than with postsynaptic DA receptors. Destruction of monoamine neurons and their associated autoreceptors by injecting 6‐hydroxydopamine into the area of the medial forebrain bundle abolished the stimulatory effects of apomorphine on methyl ester formation in striatal slices. Furthermore the putative selective DA autoreceptor agonist EMD 23 448 was also found to stimulate methyl ester formation in striatal slices. These findings, discussed in terms of calcium‐dependent functions, support the hypothesis that PCM may be a key component in the biochemical transduction of DA autoreceptor stimulation.
Reversible Dihydropyridine Isothiocyanate Binding to Brain Calcium Channels
Authors: Greenberg David A. , Cooper Edward C. , Carpenter Celia L.
Published: October 2006

Voltage‐dependent calcium channels from ileal smooth muscle can be affinity‐labeled with a [3H]dihydropyridine isothiocyanate radioligand. We examined the binding of this agent to brain membranes, to compare the properties of calcium channel drug binding sites in brain with those previously described in ileum. In brain, the [3H]dihydropyridine isothiocyanate labels sites that correspond in number and pharmacologic characteristics to binding sites for the classic calcium entry blocker, [3H]nitrendipine. However, in contrast to the covalent nature of dihydropyridine isothiocyanate binding in ileum, brain calcium channels are labeled reversibly. This difference in binding properties may reflect structural variations in voltage‐dependent calcium channels in different tissues.