bluuebunny/crossref_metadata_2025 huggingface dataset:
Treatment of vitiligo in children and adolescents by autologous cultured pure melanocytes transplantation with comparison of efficacy to results in adults
Authors: Hong WS , Hu DN , Qian GP , McCormick SA , Xu AE
Published: July 2010

Background  Transplantation of autologous cultured pure melanocytes is a well‐established procedure for the treatment of refractory and stabilized vitiligo. However, there was no report specifically comparing the efficacy with the regard to defined age groups (children‐adolescence‐adult).

Objective  We analysed the efficacy of this procedure in the treatment of vitiligo in children and adolescents and compare it with the results in adults treated during the same period and using identical procedures.

Methods  Melanocytes were isolated from the roof of suction blister, cultured and expanded with Hu16 medium in vitro, and transplanted to laser‐denuded receipt area. A total of 12 children (8–12 years), 20 adolescents (13–17 years) and 70 adults with vitiligo were treated using this procedure.

Results  The patients obtained satisfactory results (repigmentation of 50% or more) results in children, adolescents and adults were 83.3%, 95.0% and 84.0% respectively. The mean extent of repigmentation in children, adolescents and adults was 80.7%, 78.9% and 76.6% respectively. There was no statistical difference in repigmentation among these three groups. After adjusting for all factors (gender, type of vitiligo, period of stability, location of the lesion or transplanted cell density) individually or totally using multiple regression analysis, age still did not correlate to the extent of repigmentation.

Conclusions  The satisfactory results obtained in the treatment of vitiligo in children and adolescents by transplantation of cultured autologous pure melanocytes are comparable with the results in adults. Therefore, this procedure can be considered in refractory and stable vitiligo in children and adolescents, especially in patients with large vitiliginous lesions.
MONOAMINES AS POSSIBLE MEDIATORS IN THE REGULATION OF FAST AXOPLASMIC FLOW
Authors: Boegman R. J. , Wood P. L.
Published: July 2010

—The effect of drugs which have been shown to alter monoamines in the CNS on the rate of fast axoplasmic flow of [3H]leucine labelled material in the rat sciatic nerve was examined. Drugs which deplete monoamines led to a decrease while those which increase the level of monoamines led to an increase in the rate of fast axoplasmic flow. Monoaminergic drugs were also found to alter the amount of [3H]leucine incorporated into spinal cord.
INFLUENCE OF DIVALENT CATIONS ON REGULATION OF CYCLIC GMP AND CYCLIC AMP LEVELS IN BRAIN TISSUE
Authors: Ferrendelli J. A. , Rubin E. H. , Kinscherf Dorothy A.
Published: July 2010

—Depolarizing concentrations of K+elevate levels of both adenosine 3′,5′monophosphate (cyclic AMP) and guanosine 3′,5′monophosphate (cyclic GMP) in incubated slices of mouse cerebellum. Calcium is an essential requirement for the K+‐induced accumulation of cyclic GMP. Barium and Sr2+, but not Mn2+or Co2+, can substitute for Ca2+in this process. Relatively high concentrations of Mg2+inhibit the effect of Ca2+on K+‐induced accumulation of cyclic GMP. In contrast, depolarizing concentrations of K+are capable of elevating cyclic AMP levels in brain slices suspended in media containing Mg2+and no other divalent cations. High concentrations of Ca2+(1 mmor greater) augment this Mg2+‐dependent, K+‐induced accumulation of cyclic AMP, however. Strontium and Mn2+, but not Ba2+or Co2+, can substitute for Ca2+in this process, and high concentrations of Mg2+are not inhibitory. The divalent cation ionophore, A‐23187 (10 μm), in the presence of extracellular Ca2+elevates the level of cyclic GMP, but not cyclic AMP, in incubated mouse cerebellum slices.

The results of this study indicate that intracellular Ca2+concentration is a major factor regulating cyclic GMP levels in brain. In addition the present results suggest that, in brain tissue, depolarization‐induced accumulation of cyclic GMP, but not cyclic AMP, is closely linked to some Ca2+‐dependent mechanism(s) mediating release of intracellular substances.
CHARACTERISTICS OF ACETYLCHOLINE RELEASE BY SUPERFUSED SLICES OF RAT BRAIN1
Authors: Richter Judith A.
Published: July 2010

—A superfusion system has been used to examine the effects of choline and the utilization of [3H]choline during resting and potassium‐stimulated release of ACh from rat cerebrum slices. The rate of ACh release from unstimulated tissue, 0·25 nmol/g per min, increased 8‐fold when the concentration of KCl in the superfusing medium was increased from 5 to 50 mm. This rate was not maintained, however, but gradually declined to one‐half the peak rate after approx. 30 min. After an initial washout period, choline was released at a rate of 2·5‐5 nmol/g per min, which was equal to 1‐2 × 10−6min the superfusate. The addition of 1 × 10−5m‐choline to the superfusing medium was required to maintain the stimulated ACh release at near peak rates for 90 min. When hemicholinium‐3 was added to the 50 mm‐KCl medium, the release of ACh reached a peak as usual but then declined to prestimulation rates.

After introducing a pulse of radioactive choline in the superfusing medium, the specific radioactivity of choline and ACh in the superfusate was determined before and during stimulation with 50 mm‐KCl. The specific radioactivity of released ACh was always greater than that of released choline; it decreased rapidly at the onset of stimulation, and then more gradually as stimulation proceeded. The specific radioactivity of ACh released in the initial minutes of stimulation was higher than that of ACh in the tissue before stimulation. In the last 10‐20 min of stimulation the specific radioactivity of the released ACh was lower than that of the tissue ACh at the end of stimulation. The relative contributions of old and newly synthesized ACh to the releasable transmitter pool are discussed.
Brain Na+, K+‐ATPase: Alteration of Ligand Affinities and Conformation by Chronic Ethanol and Noradrenergic Stimulation In Vivo
Authors: Swann Alan C.
Published: July 2010

Abstract:These experiments examined effects of chronic ethanol, repeated noradrenergic stimulation or inhibition, and ethanol combined with the noradrenergic treatments on regulation of Na+, K+‐ATPase. Chronic treatment with ethanol reduced the sensitivity of K+‐p‐nitrophenyl‐phosphatase to ethanol, increased affinity for K+, reduced the sensitivity of K+affinity to ATP or ethanol, and reduced ΔHand ΔSfor K+activation and for the E1–E2 transition. These effects were all opposite to those of ethanol added in vitro. Treatment with yohimbine had the opposite effects on ethanol sensitivity, K+affinity, K+interactions with ethanol and ATP, and thermodynamic parameters for cation activation or conformational change. These effects were similar to those of norepinephrine in vitro. The effects of yohimbine treatment were eliminated or reduced in rats also treated with ethanol. Depletion of norepinephrine had effects opposite to those of yohimbine. These data are consistent with a reduction in membrane fluidity, at least in the vicinity of Na+, K+‐ATPase, during ethanol tolerance. Exposure to norepinephrine, in vitro or in vivo, had effects on Na+, K+‐ATPase that were similar to those of increased membrane fluidity. Key Words: Na+, K+‐ATPase—Ethanol—Norepinephrine—Membrane effects—Ethanol tolerance—Yohimbine.Swann A. C. BrainNa+, K+‐ATPase: Alteration of ligand affinities and conformation by chronic ethanol and noradrenergic stimulation in vivo.J. Neurochem. 47, 707–714 (1986).
D‐1 Dopaminergic and β‐Adrenergic Stimulation of Adenylate Cyclase in a Clone Derived from the Human Astrocytoma Cell Line G‐CCM
Authors: Balmforth Anthony J. , Ball Stephen G. , Freshney R. Ian , Graham David I. , McNamee H. Brian , Vaughan Peter F. T.
Published: July 2010

Abstract: Clones have been isolated from the human astrocytoma cell line G‐CCM. Homogenates of clone D384 contain an adenylate cyclase that is stimulated by 3,4‐dihydroxyphenylethylamine (dopamine), noradrenaline, and isoprenaline with Ka apparent values of 4, 56, and 2.7 μM, respectively. The Ka apparent value for dopamine was increased by the D‐l antagonist cis‐flupenthixol, 25 and 100 nM, to 23 and 190 μM, respectively, but was unaffected by propranolol (1 μf. Noradrenaline stimulation of adenylate cyclase was only partially inhibited by either propranolol (10 μM) or cis‐flupenthixol (1 μM). Propranolol (10 μM), but not cis‐flupenthixol (1 μM), prevented stimulation by isoprenaline. The stimulation of adenylate cyclase by dopamine and noradrenaline remained unchanged in the presence of phentolamine (1 μM) and sulpiride (1 μM). These results suggest that clone D384 contains both D‐l dopaminergic and β‐adrenergic receptors coupled to adenylate cyclase. Dopamine stimulates D384 adenylate cyclase through D‐1 receptors, isoprenaline via β‐receptors, and noradrenaline through both receptors.
Caffeine Ingestion by Rats Increases Noradrenaline Turnover and Results in Self‐Biting
Authors: Miñana María‐Dolores , Grisolía Santiago
Published: July 2010

Abstract:The effects of caffeine on the activity of central and peripheral catecholaminergic structures have been studied in rats ingesting high doses of caffeine. The activities of the enzymes tyrosine hydroxylase and dopamine‐βP‐hydroxylase were measured as well as 3,4‐dihydrox‐phenylethylamine (dopamine), adrenaline, and noradrenaline concentrations, in brain (striatum and hypothal‐amus), heart, and adrenal gland. At the peripheral level, we observed a significant increase in the dopamine and adrenaline plus noradrenaline content in the heart, but an increase in dopamine content only was found in the adrenal gland. Dopamine‐β‐hydroxylase activity in serum was increased, but the only significant enzymic change in brain was an increase in the dopamine‐p‐hy‐droxylase activity of the hypothalamus. However, an increase in catecholamine content was observed in both structures of the brain. These data suggest that the mechanisms involved in caffeine‐induced self‐biting in rats are not limited to the dopaminergic system, because we have also observed an increase in noradrenaline turnover.
Cell‐Free Desensitization of Opioid Inhibition of Adenylate Cyclase in Neuroblastoma × Glioma NG108–15 Hybrid Cell Membranes
Authors: Louie Alan K. , Law Ping‐Yee , Loh Horace H.
Published: July 2010

Abstract:When membranes from neuroblastoma |MX glioma NG108–15 hybrid cells were incubated in a cell‐free system with opioid agonists, a time‐, temperature‐, and dose‐dependent desensitization to opioid inhibition of adenylate cyclase activity was observed. The composition of the system during the incubation was manipulated to elucidate the biochemical mechanisms of desensitization. Receptor coupling appeared to be a prerequisite for desensitization, because both magnesium and sodium, which are necessary for coupling, were required for desensitization. Removal of ATP and addition of cyclic AMP or cyclic GMP had no effect on desensitization.
Extracellular Fluid Proteins of Goldfish Brain: Evidence for the Presence of Proteases and Esterases
Authors: Shashoua Victor E. , Holmquist Barton
Published: July 2010

Abstract: Preparations of enriched fractions of extracellular fluid (ECF) proteins from goldfish brain were found to contain protease(s) and esterase(s). The N‐substituted furanacryloyl (FA) peptides FA‐Phe‐Gly‐Gly and FA‐Phe‐OMe were used as model substrates for determining protease and esterase activity, respectively, in a spectro‐photometric assay. Studies of the profile of substrate specificity and identification of the types of compounds that were effective as inhibitors showed that these ECF enzymes have some distinctive properties. GSH, but not GSSG, and EDTA inhibited the protease(s) without influencing the esterase(s), whereas l‐1‐tosylamide‐2‐phenylethylchloromethyl ketone blocked both protease and esterase activites of ECF. Most of the protease and esterase properties of ECF could be bound to concanav‐alin A‐Sepharose affinity chromatographic columns in association with ependymin—a brain extracellular protein. These observations indicate that ECF may contain a metalloprotease(s) and raise the possibility that the ependymins might be a substrate for these ECF enzymes.
Tissue Distribution, Metabolism, Anticonvulsant Efficacy, and Effect on Brain Amino Acid Levels of the Glia‐Selective γ‐Aminobutyric Acid Transport Inhibitor 4,5,6,7‐Tetrahydroisoxazolo[4,5‐c]pyridin‐3‐ol in Mice and Chicks
Authors: Schousboe A. , Hjeds H. , Engler J. , Krogsgaard‐Larsen P. , Wood J. D.
Published: July 2010

Abstract: Using tritium‐labelled 4,5,6,7‐tetrahydroisoxazolo[4,5‐c]pyridin‐3‐ol (THPO) its tissue distribution and metabolism were investigated in adult mice and 4‐day‐old chicks after systemic administration of the drug. It was found not to be significantly metabolized in the brain since metabolites of THPO corresponding to only approximately 8% of the parent compound could be detected 30 min after administration of the drug intramuscularly in mice. In the liver, however, THPO was found to be metabolized to a considerable extent. In chicks THPO metabolites were found in the brain but they accounted for < 35% of the radioactivity. The brain concentration of THPO in mice and chicks corresponded to respectively 10 and 50% of the dose injected intramuscularly and the tissue level was essentially constant for at least 3 h after injection. Following systemic administration of THPO to mice and chicks the contents of aspartate, glutamate, glutamine, and γ‐aminobutyric acid (GABA) in whole brain and in synaptosomes was determined. It was found that only GABA contents were affected being increased in synaptosomes from mice and decreased in whole brain in chicks. Doses of THPO. which in chicks but not in mice led to brain levels that were sufficient to inhibit glial GABA uptake, were found to protect chicks but not mice against isonicotinic acid hydrazide‐induced seizures. The findings are compatible with the notion that THPO exerts its anticonvulsant activity by inhibition of astrocytic GABA uptake. Key Words: 4,5,6.7‐Tetrahydroisoxazolo[4,5‐c]pyridin‐3‐ol (THPO)—Anticonvulsant—γ‐Aminobutyric acid—Mice—Chicks—THPO metabolism. Schousboe A. et al. Tissue distribution, metabolism, anticonvulsant efficacy, and effect on brain amino acid levels of the glia‐selective 7‐aminobutyric acid transport inhibitor 4,5,6,7‐tetrahydroisoxazolo[4,5‐c]pyridin‐3‐ol in mice and chicks. J. Neurochem. 47, 758–763 (1986).
Tyrosine Hydroxylase Activation in Mesocortical 3,4‐Dihydroxyphenylethylamine Neurons Following Footshock
Authors: Iuvone P. Michael , Dunn Adrian J.
Published: July 2010

Abstract:Mild electric footshock resulted in activation of tyrosine hydroxylase (TH) in prefrontal cortex of mice and rats. In mice, the activation was also observed following restraint. Shock‐evoked activation of prefrontal cortex TH was characterized by a decrease of apparentKmfor the pterin cofactor 6‐methyl‐5,6,7,8‐tetrahydropterin and an increase ofVmax. Activation of prefrontal cortical TH was also demonstrated in vitro following preincubation under conditions that activate cyclic AMP‐dependent protein kinase. Treatment of mice with the noradrenergic neurotoxinN‐2‐chloroethyl‐N‐ethyl‐2‐bromobenzylamine (DSP‐4) caused a 70% decrease in pre frontal cortex norepinephrine levels but had no significant effect on the activity of TH in that brain region. Footshock resulted in the activation of prefrontal cortex TH of DSP‐4‐treated mice, suggesting that shock‐evoked activation of the enzyme occurs in terminals of mesocortical 3,4‐dihydroxyphenylethylamine neurons.
[3H]Ketanserin (Serotonin Type 2) Binding Increases in Rat Cortex Following Basal Forebrain Lesions with Ibotenic Acid
Authors: Wenk Gary L. , Engisch Kathrin L.
Published: July 2010

Abstract:The response of the serotonergic system following injury to the basal forebrain cholinergic system was investigated in rats. The density of 5‐hydroxytryptamine (serotonin) type 2 (S2) receptor sites in the frontal cortex and hippocampus was determined 1 week and 4 months after production of lesions by injections of ibotenic acid into the medial septum and nucleus basalis magnocellularis. One week later, the number of S2 receptor sites in the frontal neocortex, as defined by [3H]ketanserin binding, was unchanged. Four months later, the number of [3H]ketanserin binding sites (andBmax) was increased and high‐affinity [3H]serotonin uptake was decreased in the frontal neocortex, but not in the hippocampus, relative to unlesioned controls. Choline acetyltransferase (acetyl‐CoA:cholineO‐acetyltrans ferase; EC 2.3.1.6) activity was decreased significantly in the frontal neocortex and hippocampus 1 week and 4 months after surgery. The change in frontal neocortical S2 receptor site density (a) was inversely related to the level of choline acetyltransferase activity, (b) was specific for cholinergic denervation associated with the cortex but not the hippocampus, and (c) may represent a localized denervation supersensitivity due to degeneration of median raphe cortical afferents.
Dopamine Autoreceptors Modulate Dopamine Release from the Prefrontal Cortex
Authors: Talmaciu Reiza K. , Hoffmann Irene S. , Cubeddu L. X.
Published: July 2010

Abstract:Electrical stimulation (at 0.3, 1, or 10 Hz, 120 pulses each) produced a calcium‐dependent overflow of radioactivity from slices of the rabbit prefrontal cortex preloaded with [3H]3,4‐dihydroxyphenylethylamine ([3H]DA, [3H]dopamine) in the presence of desipramine. Flat frequency‐release curves were observed. Apomorphine and LY‐171555 inhibited in a concentration‐dependent fashion the evoked overflow of DA, an effect antagonized by haloperidol. Stimulation frequencies comparable to normal firing rates of mesocortical neurons (10 Hz) drastically reduced apomorphine‐induced inhibition of DA overflow. Haloperidol produced greater facilitation of DA overflow at 10 than at 1 Hz. Nomifensine, a neuronal uptake inhibitor, enhanced DA overflow. These results indicate that mesocortical DA neurons projecting to the prefrontal cortex have release modulatory autoreceptors of the D2 subtype.
Phorbol Esters Increase GTP‐Dependent Adenylate Cyclase Activity in Rat Brain Striatal Membranes
Authors: Olianas Maria C. , Onali Pierluigi
Published: July 2010

Abstract:4β‐Phorbol 12‐myristate 13‐acetate (PMA), added to a lysed mitochondrial fraction of rat striatum, stimulates adenylate cyclase activity with an apparent time lag of ∼30 s. Half‐maximal and maximal enzyme stimulations are obtained with 8 and 200 nMPMA, respectively. The PMA stimulation is GTP dependent, reaching a maximum of ∼60% at 50 μ.MGTP, and is associated with disappearance of the enzyme inhibition induced by micromolar concentrations of GTP. Enhancement of enzyme activity by cholera toxin and 3,4‐dihydroxyphenylethylamine is amplified by PMA only at micromolar concentrations of GTP. PMA does not affect the enzyme stimulation by forskolin but reverses the inhibition of forskolin‐stimulated enzyme by GTP. When guanyl‐5′‐yl‐imidodiphosphate is substituted for GTP, PMA does not modify adenylate cyclase activity. Enzyme inhibition by acetylcholine, Leu‐enkephalin, andR(‐)N6‐(2‐phenylisopropyl)adenosine is magnified by PMA. Stimulation of adenylate cyclase by PMA is markedly reduced following EGTA treatment, is not observed when adenyl‐5′‐yl‐imidodiphosphate is substituted for ATP as substrate for adenylate cyclase, and is enhanced by l‐α‐phosphatidyl‐l‐serine. Like PMA, 4β‐phorbol 12,13‐dibutyrate and 1‐oleoyl‐2‐acetylglycerol stimulate striatal adenylate cyclase, whereas 4β‐phorbol and 4β‐phorbol 13‐acetate are ineffective. The results indicate that phorbol esters increase striatal adenylate cyclase activity by reducing the GTP‐induced inhibition of the enzyme, presumably as a result of protein kinase C activation.
Phenytoin Dephosphorylates the Catalytic Subunit of the (Na+,K+)‐ATPase in C57/BL Mice
Authors: Guillaume Daniel , Grisar Thierry , Delgado‐Escueta Antonio V.
Published: July 2010

Abstract:The effects of phenytoin, a potent antiepileptic drug, on the active transport of cations within membranes remain controversial. To assess the direct effects of phenytoin on the Na+,K+pump, we studied the drug's influence on the phosphorylation of partially purified (Na+,K+)‐ATPase from mouse brain. (Na+,K+)‐ATPase subunits were resolved by sodium dodecyl sulfate‐polyacrylamide gel electrophoresis. Phenytoin, in vitro, decreased net phosphorylation of the (Na+,K+)‐ATPase catalytic subunit in a dose‐dependent manner (∼50% at 10−4M). When the conversion of E1‐P to E2‐P, e.g., the two major phosphorylated conformational states of (Na+,K+)‐ATPase, was blocked by oligomycin orN‐ethylmaleimide, phenytoin had no effect. The results suggest that phenytoin acts on the phosphatasic component of the reaction cycle, decreasing the phosphorylation level of the enzyme.
Immunological Nonidentity of 19K Protein and TP0 in Peripheral Nervous System Myelin
Authors: Smith Marion Edmonds , Perret Virginia
Published: July 2010

Abstract: Peripheral nervous system myelin contains as the major structural protein a glycoprotein known as P0. Another glycoprotein present in smaller amounts, known as the 19K or X protein, has been previously identified as derived from P0 and identical with the main tryptic degradation product of P0 (TP0). Although both P0 and 19K protein incorporated fucose in vitro and stained on polyacrylamide gels with the periodic acid‐Schiff stain for carbohydrate, only the P0 blotted to nitrocellulose paper showed immunoreactivity to an antibody to P0, whereas the 19K protein did not. Furthermore, when P0 was hydrolyzed with trypsin or elastase, the main degradation products reacted with P0 on immunoblots, whereas the 19K protein showed no immunoreactivity. From these studies and those of others, it may be concluded that the 19K protein shows some similarities to TP0, but probably has a different structure. P0 and 19K protein do not appear to be related as shown by lack of cross‐immunoreac‐tivity.
Tetanus Toxin in Dissociated Spinal Cord Cultures: Long‐Term Characterization of Form and Action
Authors: Habig William H. , Bigalke Hans , Bergey Gregory K. , Neale Elaine A. , Hardegree M. Carolyn , Nelson Phillip G.
Published: July 2010

Abstract:The clinical course of tetanus is notable, in addition to its often dramatic clinical presentation, by the long duration of the neuromuscular symptoms. Survivors may have tetanic manifestations for several weeks after the onset of the disease. In this article we correlate the duration of specific electrophysiologic effects produced by tetanus toxin with the degradation of cell‐associated toxin in primary cultures of mouse spinal cord neurons. From these studies we can conclude that the toxin has a half‐life of 5–6 days. Both the heavy and the light chains of tetanus toxin degrade at similar rates. Labeled toxin, visualized by radioautography, is associated with neuronal cell bodies and neurites, and its distribution is not altered during a 1‐week period following toxin exposure. Blockade of synaptic activity persists for weeks at the concentration of radiolabeled toxin used in these studies. This blockade of transmission is reversed as the toxin is degraded, suggesting that degradation of toxin may be a sufficient mechanism for recovery from tetanus.
Uptake and Metabolism of [3H] Adenosine byAplysiaGanglia and by Individual Neurons
Authors: McCaman Marilyn W.
Published: July 2010

Abstract[3H]Adenosine was taken up and metabolized by isolated ganglia of the marine molluscAplysia californica. After 2 h, most of the radioactivity was recovered as metabolites, including ATP, ADP, and AMP, as well as the deaminated products, inosine, hypoxanthine, and uric acid. Little remained in the form of adenosine. These pathways were not uniformly distributed among various tissue elements. In most individual neurons, inosine and its breakdown products were the principal metabolites of [3H]adenosine, whereas ATP and other nucleotides predominated in the connective tissue sheath. Endogenous levels of ATP, ADP, AMP, and adenosine in ganglia, sheath, and individual neurons were also determined using a fluorimetric‐HPLC assay. The concentrations of the nucleotides were quite uniform in sheath and among the individual neurons assayed (1–5 pmol/μg of protein); however, concentrations of adenosine were considerably higher in neurons than in the sheath.
In Vivo Voltammetric Monitoring of Catecholamine Metabolism in the A1 and A2 Regions of the Rat Medulla Oblongata
Authors: Suaud‐Chagny M.‐F. , Steinberg R. , Mermet C. , Biziere K. , Gonon F.
Published: July 2010

AbstractCatecholaminergic metabolism was estimated in A1 and A2 noradrenergic nuclei of the rat medulla oblongata using differential normal pulse voltammetry combined with electrochemically treated carbon fiber microelectrodes. In both areas an oxidation peak appearing at +50 mV was recorded. Electrochemical data and pharmacological experiments indicated that 3,4‐dihydroxyphenylacetic acid (DOPAC) synthesized by noradrenergic neurons was the major contributor to this signal. Indeed, α‐methyl‐p‐tyrosine, by inhibiting tyrosine hydroxylase, and pargyline, by inhibiting monoamine oxidase, both totally suppressed the peak appearing at +50 mV in A1 and A2 areas. Conversely, FLA‐63, an inhibitor of dopamine‐β‐hydroxylase, increased it. Moreover, a local and unilateral injection of catecholaminergic neurotoxin (6‐hydroxydbpamine) in the vicinity of A1 induced a 60% decrease in the peak height. This effect was prevented by pretreatment with desipramine, an inhibitor of noradrenaline reuptake, which is known to protect noradrenergic neurons against the action of 6‐hydroxydopamine. Finally, specific drugs acting on α‐2‐noradrenergic receptors (clonidine and piperoxane) modulated the peak height recorded from both structures. Thus, as previously shown in the locus ceruleus, the variations in the extracellular DOPAC levels reflect the metabolic activity of A1 and A2 noradrenergic neurons.
Effects of Arachidonic Acid on Glutamate and γ‐Aminobutyric Acid Uptake in Primary Cultures of Rat Cerebral Cortical Astrocytes and Neurons
Authors: Yu Albert C. H. , Chan Pak H. , Fishman Robert A.
Published: July 2010

Abstract The effects of arachidonic acid on glutamate and γ‐aminobutyric acid (GABA) uptake were studied in primary cultures of astrocytes and neurons prepared from rat cerebral cortex. The uptake rates of glutamate and GABA in astrocytic cultures were 10.4 nmol/mg protein/min and 0.125 nmol/mg protein/min, respectively. The uptake rates of glutamate and GABA in neuronal cultures were 3.37 nmol/mg protein/min and 1.53 nmol/mg protein/min. Arachidonic acid inhibited glutamate uptake in both astrocytes and neurons. The inhibitory effect was observed within 10 min of incubation with arachidonic acid and reached approximately 80% within 120 min in both types of culture. The arachidonic acid effect was not only time‐dependent, but also dose‐related. Arachidonic acid, at concentrations of 0.015 and 0.03 μmol/mg protein, significantly inhibited glutamate uptake in neurons, whereas 20 times higher concentrations were required for astrocytes. The effects of arachidonic acid were not as deleterious on GABA uptake as on glutamate uptake in both astrocytes and neurons. In astrocytes, GABA uptake was not affected by any of the doses of arachidonic acid studied (0.015–0.6 μmol/mg protein). In neuronal cultures, GABA uptake was inhibited, but not to the same degree observed with glutamate uptake. Lower doses of arachidonic acid (0.03 and 0.015 μmol/mg protein) did not affect neuronal GABA uptake. Other polyunsaturated fatty acids, such as docosahexaenoic acid, affected amino acid uptake in a manner similar to arachidonic acid in both astrocytes and neurons. However, saturated fatty acids, such as palmitic acid, exerted no such effect. The significance of the arachidonic acid‐induced inhibition of neurotransmitter uptake in cultured brain cells in various pathological states is discussed.
Investigation of Dopamine Content, Synthesis, and Release in the Rabbit Retina In Vitro: I. Effects of Dopamine Precursors, Reserpine, Amphetamine, and l‐DOPA Decarboxylase and Monoamine Oxidase Inhibitors
Authors: Ofori Senyo , Bretton Christine , Hof Patrick , Schorderet Michel
Published: July 2010

AbstractThe basal catecholamine content of rabbit retina was determined by liquid chromatography with electrochemical detection (LC‐EC) and 3,4‐dihydroxyphenyl‐ethylamine (dopamine, DA) found to be the major catecholamine. The immediate DA precursor, 3,4‐dihydrophenylalanine (l‐DOPA), and the metabolite, 3,4‐dihydroxyphenylacetic acid (DOPAC), were also detected at about 2.8% and 17% of DA levels, respectively. When added exogenously, l‐tyrosine did not increase the rate of DA synthesis over the basal level. In contrast, exogenous l‐DOPA led to a 3.5‐fold increase in DA, and to a 20‐fold increase in DOPAC content. The monoamine oxidase inhibitors par‐gyline and (‐)‐deprenyl differentially affected the degradation of DA, since 100 μMpargyline was apparently more effective than 100 μM(‐)‐deprenyl. Reserpine and (±)‐amphetamine each induced a Ca2+‐independent decrease of DA stores. The separate actions of reserpine and (±)‐amphetamine in lowering tissue DA levels were additive, suggesting two separate pools of DA available for release from presynaptic stores. The present study demonstrates that the LC‐EC technique may be used to investigate the modulation of the synthesis and release of retinal DA in vitro, without the prior uptake of radiolabelled transmitter.
Investigation of Dopamine Content, Synthesis, and Release in the Rabbit Retina In Vitro: II. Effects of High Potassium, Adenylate Cyclase Activators, andN‐n‐Propyl‐3‐(3‐Hydroxyphenyl) Piperidine
Authors: Ofori Senyo , Magistretti Pierre J. , Schorderet Michel
Published: July 2010

AbstractThe modulation of 3,4‐dihydroxyphenylethylamine (dopamine, DA) synthesis and release in rabbit retina in vitro by high K+; adenylate cyclase activators such as forskolin, 2‐chloroadenosine, vasoactive intestinal polypeptide (VIP); and the putative DA autoreceptor agonistN‐n‐propyl‐3‐(3‐hydroxyphenyl) piperidine (3‐PPP) has been investigated. Incubation of retinas in 50 mMK+resulted in the activation of tyrosine hydroxylase (TH). Activation did not require the presence of extracellular Ca2+. K+50 mMalso induced a Ca2+‐dependent release of DA. Forskolin 50 μMstimulated TH but 100 μM2‐chloroadenosine and 650 μMVIP did not. Individually, (±)‐3‐PPP, (‐)‐3‐PPP, and (±)‐3‐PPP reduced DA synthesis and increased its release. The effects of (+)‐3‐PPP were dose‐dependent and did not require the presence of extracellular Ca2+. The activation of TH induced by 50 mMK+, but not that induced by 50 μMforskolin, was abolished by 100 μM(±)‐3‐PPP.
PC12 Variants Deficient in Catecholamine Transport
Authors: Bitler Catherine M. , Zhang Mao‐bin , Howard Bruce D.
Published: July 2010

AbstractWe have isolated PC12 cell variants deficient in transporter‐mediated uptake of 3,4‐dihydroxyphenylethylamine (dopamine). The variants either were obtained nonselectively, or they were selected by resistance to guanethidine orN‐methyl‐4‐phenyl‐1,2,3,6‐tetrahydropyridine (MPTP). Dopamine uptake into guanethidine‐resistant cells occurred with a decreasedVmax; theKmfor dopamine and inhibition by guanethidine were normal. MPTP‐resistant cells lacked the capacity to take up dopamine. Most of the variants resembled wild‐type PC12 in their response to nerve growth factor and the storage and secretion of dopamine. MPTP‐resistant cells exhibited several deficiencies in addition to dopamine transport, i.e., no measurable storage of dopamine or acetylcholine and no observable response to nerve growth factor. Wild‐type and variant cells were compared with respect to the labeling of cell proteins with [3H]xylamine, which binds covalently to certain proteins apparently only after entering PC12 via the catecholamine transporter. When intact variant cells were used, there was markedly reduced labeling of the proteins by [3H]xylamine. Almost all of these proteins were readily labeled when cell homogenates were exposed to [3H]xylamine. However, MPTP‐resistant cells were missing three of these proteins. Northern blot analysis with cDNA clones revealed that the MPTP‐resistant cells had markedly reduced levels of several specific mRNA species.
Analysis of Cyclic AMP‐Dependent Changes in Intermediate Filament Protein Phosphorylation and Cell Morphology in Cultured Astroglia
Authors: Pollenz Richard S. , McCarthy Ken D.
Published: July 2010

Receptor agonists that increase cyclic AMP levels in cultured astroglia have been shown to increase32P‐labeling of the intermediate filament proteins glial fi‐brillary acidic protein (GFAP) and vimentin in these cells. Experiments were designed to determine (1) if the increase in32P‐labeling resulted from either an increase in the turnover or net number of phosphates associated with the intermediate filament proteins and (2) if the phosphorylation of these proteins causally affected astroglial morphology. Time course experiments indicated that 6–8 h were required to reach steady‐state32P‐labeling of both GFAP and vimentin. Treatment with forskolin (10 μM) after steady‐state32P‐labeling increased GFAP and vimentin phosphorylation fourfold and twofold, respectively, and also induced a morphological change from polygonal to process‐bearing cells within 20–30 min of drug addition. Cells incubated in media containing brain extract (30%) for 24 h at 37°C and then 3 h at 23°C under went changes from polygonal to process‐bearing cells with no apparent increase in the phosphorylation of either GFAP or vimentin. Treatment of process‐bearing cells (induced by brain extract) or polygonal cells with 10 μMforskolin at 23°C resulted in a three‐ to fourfold increase in GFAP phosphorylation without significant morphological changes. These results suggest that forskolin stimulation of GFAP and vimentin increases net number of phosphates associated with these intermediate filament proteins and that the resulting increase in phosphorylation can be dissociated from morphological changes.
Effects of Hypoxia on the Activities of Noradrenergic and Dopaminergic Neurons in the Rat Brain
Authors: Miwa Soichi , Fujiwara Motokazu , Inoue Masafumi , Fujiwara Motohatsu
Published: July 2010

The effects of hypoxia (10% O2, 90% N2) on the content, biosynthesis, and turnover of noradrenaline (NA) and 3,4‐dihydroxyphenyIethylamine (dopamine, DA) in the rat brain were examined. Up to 24 h following exposure to hypoxia, NA content in the whole brain was decreased, whereas DA content remained unchanged. The accumulation of 3,4‐dihydroxyphenylalanine (DOPA) after central decarboxylase inhibition was decreased. The turnover rate of DA after synthesis inhibition was markedly decreased up to 8 h and returned to the control level within 24 h. In contrast, the turnover rate of NA was all but unchanged, except for a 4‐h exposure. The 2‐h exposure to the hypoxic environment resulted in a significant decrease in NA content and DOPA accumulation in all brain regions tested, but no significant change was observed in DA content. The turnover rate of DA was remarkably decreased in all brain regions tested, whereas the rate of NA was slightly decreased only in the cerebral cortex and hippocampus. These results suggest that although hypoxia decreases the biosynthesis of both NA and DA, the effects of oxygen depletion on the functional activities of NA neurons differ considerably from those of DA neurons: Only in the cerebral cortex and hippocampus are the NA neurons slightly sensitive to hypoxia, whereas the DA neurons are most sensitive in all brain regions.
Effect of Chronic Desipramine Treatment on Dihydroalprenolol, Imipramine, and Desipramine Binding Sites: A Quantitative Autoradiographic Study in the Rat Brain
Authors: Biegon Anat
Published: July 2010

Desmethylimipramine (DMI) administered once daily for 10 days caused a significant decrease in P‐adrenergic receptor binding, as measured by quantitative autoradiography in discrete brain regions. The decrease was observed 72 h after the last injection throughout the cortex and in hippocampus but not in other regions, much richer in p‐receptors, such as the caudate, olfactory tubercle, superior colliculus, dorso‐medial thalamus, substantia nigra, or pineal. The same paradigm did not affect imipramine (IMI) binding in the cortex or in regions with high concentrations of IMI binding sites. DMI binding was not decreased, either. Significant increases in DMI binding were observed in frontal cortex and in the ventral aspect of the bed nucleus of the stria terminalis. We conclude that a reduction in tricyclic binding is not a general phenomenon following chronic treatment with tricyclic antidepressants, and changes in binding, when they do occur, are not correlated with areas of high binding site density.
An In Vivo Study of Dopamine Release and Metabolism in Rat Brain Regions Using Intracerebral Dialysis
Authors: Sharp T. , Zetterström T. , Ungerstedt U.
Published: July 2010

Intracerebral dialysis was used with a specifically designed HPLC with electrochemical detection assay to monitor extracellular levels of endogenous 3,4‐dihydroxyphenylethylamine (dopamine, DA) and its major metabolites, dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA), in brain regions of the halo‐thane‐anesthetized rat. Significant amounts of DA, DOPAC, and HVA were detected in control perfusates collected from striatum and n. accumbens whereas the medial prefrontal cortex showed lower monoamine levels. The ratio of DA in perfusate to DA in whole tissue suggests that in f. cortex, compared to n. accumbens and striatum, there is a greater amount of DA in the extracellular space relative to the intraneuronal DA content. The DOPAC/HVA ratio in control perfusates varied between regions in accordance with whole tissue measurements. This ratio was highest in n. accumbens and lowest in f. cortex. The monoamine oxidase inhibitor pargyline (100 mg/kg i.p.) caused an exponential decline in DOPAC, but not of HVA, in regional perfusates, an effect that was associated with an increase in DA. The data indicated a higher turnover of extracellular DOPAC in n. accumbens than in striatum and the lowest DOPAC turnover in f. cortex. The rate of decline in extracellular DA metabolite levels was slow compared to whole tissue measurements. In the perfusates there was no statistical correlation between basal amounts of DA in the perfusates and DOPAC and HVA levels or DOPAC turnover for any of the areas, indicating that measurement of DA metabolism in the brain under basal conditions does not provide a good index of DA release. In summary, this study shows clear regional differences in basal DA release and metabolite levels, metabolite patterns, and DOPAC turnover rates in rat brain in vivo.
Regulation of Taurine Transport in Rat Skeletal Muscle
Authors: Iwata Heitaroh , Obara Takeo , Kim Bong Ki , Baba Akemichi
Published: July 2010

Taurine concentration of soleus muscle (SL, slow‐twitch) was initially about twofold higher than that of extensor digitorum longus muscle (EDL, fast‐twitch). Taurine concentration in gastrocnemius muscle (GC) was intermediate between that of EDL and SL. Four days after sciatic nerve section, taurine concentration in the EDL but not in the SL was increased by 2.5‐fold. The increase was not due to the muscle atrophy and was observed 28 days after denervation. Tenotomy did not increase the total taurine content of the EDL. The increase in taurine concentration of the denervated EDL was prevented by simultaneous ingestion of guanidinoethane sul‐fonate, a competitive inhibitor of taurine transport. The initial and the maximal rates of [3H]taurine uptake were significantly higher in SL than in EDL. Denervation dramatically accelerated the initial and the maximal rates of the transport in EDL, whereas it significantly reduced those in SL. In contrast, the electrical stimulation of sciatic nerve accelerated the uptake of taurine by EDL and SL of the control but not of the curare‐treated rats. These results suggest that transport of taurine into rat skeletal muscles is regulated differently by neural information and by muscular activity, and that the regulation is dependent on the muscle phenotype.
Brain Adrenoceptor Binding Sites in Mice Susceptible (DBA/2J) and Resistant (C57 Bl/6) to Audiogenic Seizures
Authors: Jazrawi S. P. , Horton R. W.
Published: July 2010

We have examined if the age‐related susceptibility of DBA/2J mice to audiogenic seizures is the result of an abnormality in the number or sensitivity of brain adrenoceptors. The binding of α1‐‐, α2‐‐, and β‐adrenoceptor ligands to membranes prepared from whole brain or regions of brain of DBA/2J mice was measured at various ages, corresponding to the periods before, during, and after the maximal sensitivity to audiogenic seizures. For comparison, we have studied concurrently age‐matched C57 B1/6 mice, a strain resistant to audiogenic seizures at all ages. There was no difference in the binding of α2‐‐or β‐adrenoceptor ligands to whole brain membranes between the two strains of mice at any age. The maximal number of α1‐‐adrenoceptor binding sites was lower in whole brains of DBA/2J mice than of C57 B1/6 mice at all ages studied except 13–15 days of age. The differences were small (maximally 17%) but were statistically significant at 21–23 days of age, the time of maximal sensitivity of DBA/2J mice to audiogenic seizures. No difference between the two strains was found in the number or affinity of α1‐‐or α2‐‐adrenoceptor binding sites at any age in any of the brain regions studied. The age‐related susceptibility of DBA/2J mice to audiogenic seizures is not the result of an abnormality in number or sensitivity of α2‐‐or β‐adrenoceptor binding sites, but a reduced number of α1‐‐adrenoceptor binding sites may be involved.
Reevaluation of the Role of Gangliosides as Receptors for Tetanus Toxin
Authors: Critchley David R. , Habig William H. , Fishman Peter H.
Published: July 2010

Binding of tetanus toxin to rat brain membranes was of lower affinity and capacity when binding was determined in 150 mMNaCl, 50 mMTris‐HCl (pH 7.4) than in 25 mMTris‐acetate (pH 6.0). Binding under both conditions was reduced by treating the membranes with neuraminidase. Pronase treatment, however, reduced toxin binding only in the Tris‐saline buffer (pH 7.4). In addition, the concentration of gangliosides required to inhibit toxin binding was 100‐fold higher in Trissaline compared to Tris‐acetate buffer. The toxin receptors in the membranes were analyzed by ligand blotting techniques. Membrane components were dissolved in sodium dodecyl sulfate, separated by polyacrylamide gel electrophoresis, and transferred to nitrocellulose sheets, which were overlaid with125I‐labeled toxin. Tetanus toxin bound only to material that migrated in the region of the dye front and was extracted with lipid solvents. Gangliosides isolated from the lipid extracts or other sources were separated by TLC on silica gel and the chromatograms were overlaid with labeled tetanus toxin. The toxin bound to areas where the major rat brain gangliosides migrated. When equimolar amounts of different purified gangliosides were applied to the chromatogram, binding of the toxin was in the order GDlb≅ GTlb≅ GQ1b> GD2> GD3≫ GD1a≅ GM1. Thus, the toxin appears to have the highest affinity for gangliosides with a disialyl group linked to the inner galactosyl residue. When binding of tetanus toxin to transfers and chromatograms was determined in the Tris‐saline buffer (pH 7.4), the toxin bound to the same components but the extent of binding was markedly reduced compared with the low‐salt and ‐pH conditions. Our results indicate that the interaction of tetanus toxin with rat brain membranes and gangliosides is greatly reduced under more physiological conditions of salt and pH and raise the possibility that other membrane components such as sialoglycoproteins may be receptors for the toxin under these conditions.
Studies of the Regulation of Basal Adenylate Cyclase Activity by Membrane Polyunsaturated Fatty Acids in Cultured Neuroblastoma
Authors: Murphy M. G.
Published: July 2010

The role of membrane polyunsaturated fatty acids (PUFAs) in the regulation of basal adenylate cyclase activity was examined in intact N1E‐115 neuroblastoma cells. Addition of linoleic acid (50 μM) to the culture medium for 48 h resulted in a significant increase in phospholipid PUFA content and in a two‐ to fivefold increase in basal accumulation of cyclic AMP (cAMP). Both phenomena were reversed on removal of linoleate from the medium. PUFA enrichment stimulated cell proliferation by —20% without altering the relative proportion of cellular protein. The supplemented cells synthesized significantly larger amounts of prostaglandin (PG) E and D than did the controls; however, blockade of PG synthesis by indomethacin or ibuprofen did not alter cAMP formation. Supplemented cells contained higher levels of malondialdehyde (MDA) than did controls, and MDA formation was reduced by coculture with α‐tocopherol; however, its inclusion in the medium did not affect cAMP accumulation. Linoleate‐supplemented cells responded to cyclase‐activating agonists to the same extent as did control cells. Responses to inhibitory agonists (e.g., isoproterenol and carbamylcholine) were altered, but not to a sufficient extent to account for the PUFA‐dependent increases in basal adenylate cyclase activity.
Molecular Forms of Acetylcholinesterase and Butyrylcholinesterase in the Aged Human Central Nervous System
Authors: Atack John R. , Perry Elaine K. , Bonham James R. , Candy John M. , Perry Robert H.
Published: July 2010

The distributions of acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) molecular forms and their solubility characteristics were examined, using density gradient centrifugation, in various regions of the postmortem human CNS. Total AChE activity varied extensively (50‐fold) among the regions investigated, being highest in the telencephalic subcortical structures (caudate nucleus and nucleus of Meynert); intermediate in the substantia nigra, cerebellum, and spinal cord; and least in the fornix and cortical regions (hippocampus and temporal and parietal cortex). Total BChE activity was, in contrast, much more evenly distributed, with only a threefold variation between the regions studied. Although the patterns of molecular forms of each enzyme were broadly similar among the different areas, regional variations in the distribution and abundance of the various forms of AChE were much greater than those of BChE. Thus, although the tetrameric G4 form of AChE constituted the majority of the total AChE activity in all regions examined, the ratio of the G4 form to the monomeric Gl form, the latter of which constituted the majority of the remaining activity, varied markedly, ranging from 21 in the caudate nucleus to 1.7 in the tem poral cortex. In addition to the G4 and Gl forms of AChE, the dimeric G2 form was observed in the nucleus of Meynert and a fast‐sedimenting (16S) species was found in samples of both the parietal cortex and spinal cord. In contrast, the G4 and Gl forms of BChE were the only molecular species observed in the different areas and the G4.G1 ratio varied from 3.3 in the substantia nigra to 0.9 in the temporal cortex. Regarding the solubility characteristics of the individual AChE and BChE molecular forms, the majority of the G4 form of AChE was extractable only in the presence of detergent, indicating a predominantly membrane‐bound localization of this species. The smaller AChE forms (Gl and G2) and both the Gl and G4 forms of BChE were all relatively evenly distributed between soluble and membrane‐bound species. These findings are discussed in relation to neurochemical and neuroanatomical, particularly cholinergic, features of the regions examined.