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1.
Dopamine Synthesis in Synaptosomes: Relation of Autoreceptor Functioning to pH, Membrane Depolarization, and Intrasynaptosomal Dopamine Content 总被引:2,自引:2,他引:0
Abstract: Factors affecting dopamine (DA) synthesis in rat striatal synaptosomes were examined by measuring the conversion of [3 H]tyrosine (Tyr) to [3 H]DA. Any [3 H]DA that was synthesized was extracted into a toluene-based scintillation cocktail and quantitated by liquid scintillation spectrometry. The extraction was facilitated using di-(2-ethylhexyl) phosphoric acid (DEHP), a liquid cation exchanger. DA, apomorphine, and other DA agonists were much less potent inhibitors of DA synthesis in striatal synaptosomes at pH 6.2 than at pH 7.2. 3-(3-Hydroxyphenyl)- N - n -propylpiperidine (3-PPP), a putative DA autoreceptor agonist, was inactive at pH 6.2. However, at pH 7.2, 3-PPP did inhibit DA synthesis. This inhibition was reversed by sulpiride, a DA receptor antagonist, but not by benztropine, a DA uptake blocker, suggesting that 3-PPP inhibits DA synthesis by stimulating the DA autoreceptor. DA release from synaptosomes was much greater at pH 6.2 than at pH 7.2, most probably because the synaptosomal membrane appears to be depolarized at pH 6.2, as measured by the accumulation of [3 H]tetraphenylphosphonium ions. Since tyrosine hydroxylase is inhibited by DA, this finding suggested that low assay buffer pH (i.e., pH 6.2) might interfere with the ability of 3-PPP and other DA agonists to inhibit DA synthesis, by promoting DA release. Likewise, reserpine and tetrabenazine, compounds which disrupt vesicular DA storage, were much less effective inhibitors of DA synthesis at pH 6.2 (high basal DA release). Moreover, d -amphetamine and high buffer potassium concentrations, treatments which promote DA release, also interfered with the ability of 3-PPP to inhibit DA synthesis. Thus, modulation of the release of DA in equilibrium with tyrosine hydroxylase may be a mechanism by which the DA autoreceptor regulates DA synthesis. 相似文献
2.
Regulation of catecholamine synthesis in rat brain synaptosomes 总被引:9,自引:9,他引:0
Abstract— Catecholamine synthesis in synaptosomal preparations of rat striatum, cortex and brain stem was investigated. The striatum had much higher activity than either the cortex or brain stem. Equilibration of labelled tyrosine between tissue and incubation medium was completed within 2 min. The apparent Km of tyrosine hydroxylase (EC 1.14.3a) and of the overall catecholamine synthetic pathway were both approximately 5 ± 10?6m for tyrosine. The following amines were found to inhibit striatal dopamine synthesis: dopamine, 25% inhibition at 5 ± 10?7m ; noradrenaline, 25% inhibition at 5 ± 10?6m ;and serotonin, 30% inhibition at 10?5m . The catecholamine-induced inhibition of synthesis was antagonized by pre-incubation with cocaine. Increasing the potassium concentration from 5 to 55 mm caused a release of amines into the medium which was accompanied by a 40% increase in dopamine synthesis, when synthesis was measured during the first 5 min of exposure to elevated potassium. These results indicate that synaptosomal catecholamine synthesis is inhibited by increases in intra-synaptosomal amine levels, and that short-term exposure to depolarizing concentrations of potassium can increase synthesis. 相似文献
3.
Abstract— A modified tritium release assay for the measurement of synaptosomal tyrosine hydroxyl-ation. with a sensitivity suitable for use on areas of the rat brain with a low density of catecholamine terminals. is described. The apparent Km , for tyrosine hydroxylase in the hippocampus was 9.3 μM. in the hypothalamus 6.1 μM and in the striatum 9.9 μM Preparations from all three regions showed a pH optimum of 6.0–6.2, and the activities were reduced to a small % of control by synaptosomal disruption. 3-iodotyrosine. noradrenaline and reserpine. Membrane depolarization at a pH of 6.1 did not elevate tyrosine hydroxylation rates in any of the regions studied, although striatal tyrosine hy-droxylation rates were elevated at a pH of 7.2 by 55 mM-K+ . The addition of dibutyryl cyclic AMP (0.5 mM) to the medium produced a 20-30% elevation of the rates of hydroxylation in all three regions studied: addition of tetrahydrobiopterin (0.2 mM) elevated hydroxylation rates in the hypothalamus and striatum. These results indicate that many characteristics of tyrosine hydroxylase from the three regions are similar. In each case the enzyme is apparently sensitive to end-product inhibition and to cyclic AMP activation. 相似文献
4.
The effect of veratridine-mediated depolarization on rat brain synaptosomal respiration in the presence and absence of calcium was investigated. Studies on respiration were performed employing three different pretreatments of the synaptosomes which attempted to deplete endogenous substrates. First, synaptosomes were preincubated for 10 min in the absence of any substrates in medium either containing or devoid of calcium. Second, synaptosomes were preincubated for either 15 or 60-min periods in the presence and absence of calcium, and the incubation medium was changed by centrifugation and resuspension of synaptosomes in their respective media. Irrespective of the prior treatment, maximal stimulation of respiration (400-600%) during veratridine (100 microM) elicited depolarization was observed only when calcium was present in the incubation media. In incubations performed in the absence of calcium, veratridine addition either modestly stimulated (10- and 15-min preincubated synaptosomes) or did not affect (60-min preincubated synaptosomes) the rate of respiration. However, when calcium was added back to these incubations the rate of respiration in the presence of veratridine was stimulated by five- to six-fold. Similarly, the rates of 14CO2 production from [1-14C]- and [2-14C]pyruvate were increased by veratridine only when synaptosomes were incubated in calcium-replete medium. These data indicate that calcium plays an obligatory role in depolarization-elicited stimulation of synaptosomal oxidative processes. 相似文献
5.
Influence of Calcium on Dopamine Synthesis and Tyrosine Hydroxylase Activity in Rat Striatum 总被引:5,自引:5,他引:0
Abstract: We have investigated three aspects of the relationship between calcium and tyrosine hydroxylase activity in rat striatum. In the first series of experiments, we examined the hypothesis that the rise in dopamine synthesis during increased impulse flow results from a calcium-induced activation of tyrosine hydroxylase. Calcium (12.5–200 μ M ) had no effect when added to crude enzyme or enzyme partially purified by gel filtration. Moreover, incubation of synaptosomes with excess calcium (up to 3.5 m M ) had little or no effect on dopamine synthesis. Incubation with the depolarizing alkaloid veratridine (75 μ M ) did increase dopamine synthesis, but did not alter the activity of tyrosine hydroxylase subsequently prepared from the synaptosomes, despite the presumed rise in intracellular calcium. In the second series we examined the hypothesis that increased dopamine synthesis after axotomy results from activation of tyrosine hydroxylase owing to a decrease in intracellular calcium. Addition of the calcium chelator EGTA (100 μ M ) to crude or partially purified enzyme was without effect, whereas incubation of synaptosomes with EGTA (500 μM ) decreased cell-free enzyme activity. In the third experimental series we examined the relationship between calcium and activation of tyrosine hydroxylase by dibutyryl cyclic AMP. EGTA failed to alter the increase in the activity of tyrosine hydroxylase prepared from synaptosomes incubated with dibutyryl cyclic AMP. However, it blocked the increase in synaptosomal dopamine synthesis and dopamine content normally produced by the cyclic AMP analogue. Thus, tyrosine hydroxylase does not appear to be activated by either increases or decreases in calcium availability. However, calcium may be important for the maintenance of basal tyrosine hydroxylase activity, and may play an indirect role in the expression of tyrosine hydroxylase activation produced by other means. 相似文献
6.
Mechanisms controlling choline transport and acetylcholine synthesis in motor nerve terminals during electrical stimulation
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Electrical stimulation of the chick ciliary nerve leads to a frequency-dependent increase in the Na+-dependent high affinity uptake of [3H]choline (SDHACU) and its conversion to acetylcholine (ACh) in the nerve terminals innervating the iris muscle. The forces that drive this choline (Ch) uptake across the presynaptic membrane were evaluated. Depolarization with increased [K+] out or veratridine decreases Ch accumulation. In addition to the electrical driving force, energy is provided by the Na+ gradient. Inhibition of the Na,K-ATPase decreased the Ch taken up. Thus, changes in the rate of Ch transport are dependent on the electrochemical gradients for both Ch and Na+. Ch uptake and ACh synthesis were increased after a conditioning preincubation with high [K+] out or veratridine. As is the case for electrical stimulation, this acceleration of Ch uptake and ACh synthesis was strongly dependent on the presence of Ca++ in the incubation medium. Na+ influx through a TTX-sensitive channel also contributed to this acceleration. Inasmuch as membrane depolarization reduces the initial velocity of Ch uptake and ACh synthesis, their increases during electrical stimulation therefore cannot be the direct effect of the depolarization phase of the action potential. Instead they are the result of the ionic fluxes accompanying the presynaptic spike. It is concluded that stimulation of Ch uptake and ACh synthesis by nerve activity depends first, on the ACh release elicited by Ca++ influx after depolarization and second, on the activation of the Na,K-ATPase due to Na+ entry. Furthermore, it is suggested that the release of ACh after stimulation drives translocation of cytoplasmic ACh into a protected compartment (probably vesicular). This recompartmentation of intraterminal ACh stimulates ACh synthesis by mass action, allowing further accumulation of Ch. 相似文献
7.
The effect of [Ca2+] and [H+] on the functional recovery of rat brain synaptosomes from anoxic insult in vitro.
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The energy status (as measured by the ATP/ADP ratio), oxidative metabolism (14CO2 output) and neurotransmitter synthesis ( [14C]acetylcholine production) by rat brain synaptosomes utilizing [U-14C]glucose has been studied. The ability of anoxia in vitro to permanently alter these parameters was investigated with reference to external [Ca2+] and [H+]. It has previously been shown that anoxic damage to synaptosomal preparations is only apparent when their metabolism is stimulated by veratridine [Harvey, Booth & Clark (1982) Biochem. J. 206, 433-439]. It is concluded that low [Ca2+] ameliorates, and high [H+] exacerbates, the damage sustained by veratridine-stimulated anoxic synaptosomes. The combined effects of low pH, anoxia and veratridine stimulation on synaptosomal metabolism most closely approximated to the irreversible damage to brain metabolism observed during acute hypoxia in vivo [Booth, Harvey & Clark (1983) J. Neurochem. 40, 106-110]. Suitably treated synaptosomal preparations may therefore be usefully employed as models to study impaired neurotransmitter synthesis in vivo. 相似文献
8.
In Vitro Synthesis of Dopamine and Noradrenaline in the Isolated Rat Pineal Gland: Day-Night Variations and Effects of Electrical Stimulation 总被引:1,自引:0,他引:1
Kurt Racké Heike Krupa Hannsjörg Schröder Lutz Vollrath 《Journal of neurochemistry》1989,53(2):354-361
Isolated rat pineal glands were incubated in vitro in a medium containing [14C]dopamine or [14C]tyrosine, and the tissue contents of 14C-labelled and total dopamine and noradrenaline were determined by HPLC followed by electrochemical detection and scintillation spectrometry. During incubation with [14C]dopamine, the labelled amine accumulated in pineal glands and was partially converted into [14C]noradrenaline. Nomifensine, a neuronal amine uptake blocker, largely inhibited the accumulation of [14C]dopamine and the formation of [14C]noradrenaline. These experiments demonstrated dopamine beta-hydroxylase activity in the sympathetic nerves of the pineal gland. During incubation with [14C]tyrosine, formation of [14C]dopamine and [14C]noradrenaline was observed in the pineal tissue, indicating that noradrenaline can also be synthesized from dopamine, endogenously formed in the gland. Electrical stimulation of the stalk region of the pineal gland during incubation with [14C]dopamine enhanced the accumulation of [14C]dopamine and synthesis of [14C]noradrenaline. Electrical stimulation also enhanced the formation of [14C]dopamine during incubation with [14C]tyrosine. Compared to that at midday, the tissue content of endogenous noradrenaline at midnight was enhanced by 50% and that of dopamine by 450%. The in vitro accumulation of [14C]dopamine, as well as the synthesis of [14C]dopamine and [14C]noradrenaline, was also increased at midnight. In conclusion, sympathetic nerves in the rat pineal gland contain tyrosine hydroxylase and dopamine beta-hydroxylase, the two enzymes required for the synthesis of noradrenaline.(ABSTRACT TRUNCATED AT 250 WORDS) 相似文献
9.
Paulino Gómez-Puertas Alberto Martínez-Serrano Pablo Blanco Jorgina Satrústegui Elena Bogónez 《Journal of neurochemistry》1991,56(6):2039-2047
Temporal changes in the phosphorylation level of synaptosomal phosphoproteins following depolarization of synaptosomes were investigated under conditions restricting calcium influx. High-K+ depolarization in media of low [Na+]o (32 mM during preincubation and depolarization) at pH 6.5 resulted in a pronounced fall in the cytosolic free calcium concentration transient, and in a reduction in the initial K(+)-stimulated 45Ca2+ uptake and endogenous acetylcholine release relative to the values obtained with control synaptosomes (preincubated and depolarized in Na(+)-based media). This reduction was paralleled by a decrease in the rate of dephosphorylation of the synaptosomal protein P96. A slower dephosphorylation of P96 also was observed on exposure to 20 microM veratridine at 0.5 mM external calcium. Our results indicate that, similar to synapsin I phosphorylation, P96 dephosphorylation shows a graded response to the amount of calcium entering the presynaptic terminal. Depolarization of synaptosomes under conditions restricting the influx of calcium revealed a transient dephosphorylation (reversed within 10 s) of the phosphoprotein P65. The possible significance of this finding to the process of neurotransmitter release is discussed. 相似文献
10.
Tyrosine Hydroxylase Regulation: Apparent Kinetic Alterations Following Incubation of Brain Slices in a Sodium-free Medium 总被引:5,自引:5,他引:0
In an attempt to determine if alterations in intraneuronal Ca2+ may regulate tyrosine hydroxylase activity, brain slices were subjected to experimental manipulations known to increase the intraneuronal concentration of free Ca2+ ions. Incubation of either striatal or olfactory tubercle slices in a Na+-free medium for 15 min at 37 degrees resulted in a marked increase in the activity of tyrosine hydroxylase present in the 20,000 g supernatant fraction of homogenates prepared from the slices. Tyrosine hydroxylase isolated from slices previously incubated in a Na+-free, choline-enriched medium or in a Na+-free, sucrose-enriched medium exhibited maximal activities when assayed at pH 6.0 and 7.0, respectively. However, the percentage stimulation of enzyme activity induced by incubation of the slices in a Na+-free medium was maximal when the enzyme assays were performed at pH 7.0. The observed increase in enzyme activity seems to be mediated by a decrease in the apparent Km of the enzyme for pteridine cofactor, regardless of whether the kinetic enzyme analyses were conducted at pH 6.0 or 7.0, and by an increase in the Ki of the enzyme for end-product inhibitor dopamine. The apparent kinetic changes in the enzyme do not seem to result from alterations in the endogenous dopamine content of the slices, and they are independent of any increase in dopamine release that might have occurred as a response to the augmented intraneuronal Ca2+ concentration. Furthermore, the activation of tyrosine hydroxylase produced by incubating slices in a Na+-free medium is observed even in slices depleted of dopamine by pretreatment of rats with reserpine 90 min before preparation of brain slices. The activation of tyrosine hydroxylase observed under these experimental conditions does not seem to be mediated by cAMP or by a cAMP-dependent phosphorylation process. It is suggested that the changes in tyrosine hydroxylase reported are mediated primarily by a rise in the free Ca2+ concentration within the nerve tissue. These observations are consistent with the hypothesis that the kinetic activation of tyrosine hydroxylase produced after depolarization of central dopaminergic neurons may occur through a Ca2+-dependent even other than transmitter release. 相似文献
11.
In rat striatal synaptosomes incubated with [14C]tyrosine, the evolution of 14CO2, taken as a measure of dopamine synthesis, was inhibited by exogenous dopamine and by the dopaminergic receptor agonist ADTN. The inhibition was not counteracted by dopaminergic receptor antagonists (haloperidol, sulpiride, pimozide or domperidone). Instead, it was prevented by dopamine uptake blockers, suggesting that dopamine and ADTN (a substrate of the dopamine carrier) acted once inside the nerve endings and not through activation of autoreceptors on their external membrane. The dopamine uptake inhibitors nomifensine, benztropine and cocaine increased 14CO2 evolution from incubated striatal synaptosomes. Depolarization with KCl also increased dopamine synthesis and this action was potentiated when the reuptake of the released catecholamine was prevented by carrier blockers. The rate of dopamine synthesis was lowered when synaptosomal dopamine was raised upon incubation with monoamine oxidase inhibitors or with l-DOPA. The inhibition was counteracted by dopamine reuptake blockers. The data suggest that dopamine synthesis in striatal nerve endings is under the inhibitory control of the transmitter recaptured following release. 相似文献
12.
Synaptosomes from rat forebrain were analyzed for the presence of phosphotyrosine-containing proteins by immunoblotting with antiphosphotyrosine antibodies. Using this technique, 10-11 phosphotyrosine-containing proteins were detected. Depolarization of synaptosomes by transfer to a high (41 mM) K+ medium resulted in increases in the phosphotyrosine content of several synaptosomal proteins, the most pronounced increase being associated with a membrane protein of M(r) 117,000 (ptp117). Additional proteins exhibiting depolarization-dependent increases in phosphotyrosine content had molecular weights of 39,000, 104,000, 135,000, and 160,000. The depolarization-dependent increase in the phosphotyrosine content of ptp117 was apparent within 30 s of the onset of depolarization, reached a maximum between 3 and 5 min, and then decreased to near control values by 30 min. The increase in tyrosine phosphorylation of ptp117 was dependent on the concentration of K+ in the depolarizing medium and was maximal with [K+] in excess of 50 mM. It was also calcium dependent and did not occur in the absence of extracellular calcium. The addition of veratridine to the incubation medium also resulted in an increase in the tyrosine phosphorylation of ptp117. The results suggest that the phosphorylation of synaptic proteins on tyrosine residues may be involved in the regulation or modulation of synaptic activity. 相似文献
13.
Veratridine in low concentrations (20 μ) and at high pH (pH 9) acts as a synergist for carbamylcholine-induced depolarizations in the electroplax of electric eel. This potentiation is not sensitive to tetrodotoxin, but is significantly reduced by d-tubocurarine. Veratridine alone does not depolarize this preparation at the concentration used (20 μ). The increased carbamylcholine depolarization arising in the presence of veratridine does not simply sum with the carbamylcholine depolarization; the fractional contribution of veratridine to the total depolarization decreases as the carbamylcholine concentration is increased, and at 50 μM carbamylcholine no significant difference is apparent between groups with and without veratridine. Depolarization with increased external K+, unlike carbamylcholine depolarization, is not potentiated by veratridine. 相似文献
14.
Maria Ereciska Malgorzata M. Zaleska David Nelson Itzhak Nissim Marc Yudkoff 《Journal of neurochemistry》1990,54(6):2057-2069
The synaptosomal metabolism of glutamine was studied under in vitro conditions that simulate depolarization in vivo. With [2-15N]glutamine as precursor, the [glutamine]i was diminished in the presence of veratridine or 50 mM KCl, but the total amounts of [15N]glutamate and [15N]aspartate formed were either equal to those of control incubations (veratridine) or higher (50 mM [KCl]). This suggests that depolarization decreases glutamine uptake and independently augments glutaminase activity. Omission of sodium from the medium was associated with low internal levels of glutamine which indicates that influx occurs as a charged Na(+)-amino acid complex. It is postulated that a reduction in membrane potential and a collapse of the Na+ gradient decrease the driving forces for glutamine accumulation and thus inhibit its uptake and enhance its release under depolarizing conditions. Inorganic phosphate stimulated glutaminase activity, particularly in the presence of calcium. At 2 mM or lower [phosphate] in the medium, calcium inhibited glutamine utilization and the production of glutamate, aspartate, and ammonia from glutamine. At a high (10 mM) medium [phosphate], calcium stimulated glutamine catabolism. It is suggested that a veratridine-induced increase in intrasynaptosomal inorganic phosphate is responsible for the enhancement of flux through glutaminase; calcium affects glutaminase indirectly by modulating the level of free intramitochondrial [phosphate]. Because phosphate also lowers the Km of glutaminase for glutamine, augmentation of the amino acid breakdown may occur even when depolarization lowers [glutamine]i. Reducing the intrasynaptosomal glutamate to 26 nmol/mg of protein had little effect on glutamine catabolism, but raising the pH to 7.9 markedly increased formation of glutamate and aspartate. It is concluded that phosphate and H+ are the major physiologic regulators of glutaminase activity. 相似文献
15.
The mechanisms of tyrosine hydroxylase (TH) activation by depolarization or exposure of dopaminergic terminals to cyclic AMP have been compared using rat striatal slices. Tissues were incubated with veratridine or 60 mM K+ (depolarizing conditions), on the one hand, and forskolin or dibutyryl cyclic AMP, on the other. K+-(or veratridine-)induced depolarization triggered an activation of TH (+75%) that persisted in soluble extracts of incubated tissues. This effect disappeared when drugs (EGTA, N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide, Gallopamil) preventing Ca2+- and calmodulin-dependent processes were included in the incubating medium. In contrast, prior in vivo reserpine treatment or in vitro addition of benztropine did not affect the depolarization-induced activation of TH. In vitro studies of soluble TH extracted from depolarized tissues indicated that activation was associated with a marked increase in the enzyme Vmax but with no change in its apparent affinity for the pteridin cofactor 6-methyl-5,6,7,8-tetrahydropterin (6-MPH4) or tyrosine. Furthermore, the activated enzyme from depolarized tissues exhibited the same optimal pH (5.8) as native TH extracted from control striatal slices. In contrast, TH activation resulting from tissue incubation in the presence of forskolin or dibutyryl cyclic AMP was associated with a selective increase in the apparent affinity for 6-MPH4 and a shift in the optimal pH from 5.8 to 7.0-7.2. Clear distinction between the two activating processes was further confirmed by the facts that heparin- and cyclic AMP-dependent phosphorylation stimulated TH activity from K+-exposed (and control) tissues but not that from striatal slices incubated with forskolin (or dibutyryl cyclic AMP). In contrast, the latter enzyme but not that from depolarized tissues could be activated by Ca2+-dependent phosphorylation. These data strongly support the concept that Ca2+- but not cyclic AMP-dependent phosphorylation is responsible for TH activation in depolarized dopaminergic terminals. 相似文献
16.
Ca2+ transport by intact synaptosomes: the voltage-dependent Ca2+ channel and a re-evaluation of the role of sodium/calcium exchange 总被引:6,自引:0,他引:6
The verapamil-sensitive Ca2+ channel in the synaptosomal plasma membrane is investigated. Verapamil is without effect on Ca2+ uptake or steady-state content in synaptosomes with a polarized plasma membrane, but completely inhibits the additional Ca2+ uptake following plasma-membrane depolarization by high [K+], by veratridine plus ouabain or by high concentrations of the permeant cation tetraphenylphosphonium. Verapamil-insensitive Ca2+ influx and steady-state content are identical in polarized and depolarized synaptosomes, even though the Na+ electrochemical potential is greatly decreased in the latter, indicating that Na+/Ca2+ exchange is not a significant mechanism for Ca2+ efflux under these conditions. A transient Na+-dependent Ca2+ efflux can only be observed on addition of Na+ to Na+-depleted depolarized synaptosomes. While 0.2 mM verapamil decreases the ate of 86Rb+ efflux and 22Na+ entry during depolarization induced by veratridine plus ouabain, the final steady-state Na+ accumulation is not inhibited. Ca2+ efflux from synaptosomes following mitochondrial depolarization does not occur by a verapamil-sensitive pathway. 相似文献
17.
The potential interaction between kappa-opiate receptors and dopamine activity was examined in this study by monitoring the effect of U-50,488H on the release of endogenous dopamine from rat striatal slices in both the absence and presence of 10 microM nomifensine, a potent dopamine uptake inhibitor. Basal dopamine release was increased 10-fold in the presence of nomifensine, and the normally steady base line was observed to increase gradually under these conditions. U-50,488H, a potent kappa-agonist, enhanced the spontaneous release of dopamine, but only at relatively high concentrations (40.0 microM) and only in the absence of nomifensine. Likewise, nomifensine and U-50,488H (40.0 microM) each significantly inhibited the synaptosomal uptake of [3H]dopamine. As with basal release, nomifensine markedly enhanced the potassium-evoked release of dopamine, and this evoked release was significantly attenuated by U-50,488H (0.4 and 40.0 microM) in both the absence and presence of nomifensine. This opiate-mediated inhibition of evoked dopamine release was antagonized in a time-dependent manner by the putative kappa-antagonist, WIN 44,441-3, suggesting that striatal kappa-receptor activation modulates dopamine release. 相似文献
18.
These studies were carried out to characterize the activation of rat striatal tyroxine hydroxylase produced by depolarization of the medial forebrain bundle and to evaluate the possible role of cyclic AMP as a mediator of this activation. The enzymatic properties of tyrosine hydroxylase following in vivo depolarization were compared to those produced by treatment of striatal synaptosomes with dibutyryl cyclic AMP (dbcAMP). Similar effects were observed with regard to enzyme distribution, altered sensitivity to dopamine-induced inhibition, and activity as a function of tyrosine concentration. However, differences between the two treatments were also apparent. First, treatment with dbcAMP shifted the pH optimum from 6.2 to 7.0. In contrast, electrical stimulation decreased the rate of decline in activity as the pH was increased above the optimum, but did not shift the pH optimum. Second, plots of tyrosine hydroxylase activity versus cofactor concentration revealed two enzyme forms for both control and electrically stimulated preparations. However, dbcAMP treatment converted the enzyme to a single high affinity form. These results can be explained by one of the following: (1) cyclic AMP is the sole mediator of enzyme activation, but does not produce a maximally activated enzyme following in vivo depolarization (2) cyclic AMP is only one of several mediators involved or (3) cyclic AMP is not involved in depolarization-induced activation, with activation occurring via the mediation of other intracellular messengers, such as calcium. 相似文献
19.
Relationships between the neuronal sodium/potassium pump and energy metabolism. Effects of K+, Na+, and adenosine triphosphate in isolated brain synaptosomes 总被引:6,自引:0,他引:6
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The relationships between Na/K pump activity and adenosine triphosphate (ATP) production were determined in isolated rat brain synaptosomes. The activity of the enzyme was modulated by altering [K+]e, [Na+]i, and [ATP]i while synaptosomal oxygen uptake and lactate production were measured simultaneously. KCl increased respiration and glycolysis with an apparent Km of about 1 mM which suggests that, at the [K+]e normally present in brain, 3.3-4 mM, the pump is near saturation with this cation. Depolarization with 6-40 mM KCl had negligible effect on ouabain-sensitive O2 uptake indicating that at the voltages involved the activity of the Na/K ATPase is largely independent of membrane potential. Increases in [Na+]i by addition of veratridine markedly enhanced glycoside-inhibitable respiration and lactate production. Calculations of the rates of ATP synthesis necessary to support the operation of the pump showed that greater than 90% of the energy was derived from oxidative phosphorylation. Consistent with this: (a) the ouabain-sensitive Rb/O2 ratio was close to 12 (i.e., Rb/ATP ratio of 2); (b) inhibition of mitochondrial ATP synthesis by Amytal resulted in a decrease in the glycoside-dependent rate of 86Rb uptake. Analyses of the mechanisms responsible for activation of the energy-producing pathways during enhanced Na and K movements indicate that glycolysis is predominantly stimulated by increase in activity of phosphofructokinase mediated via a rise in the concentrations of adenosine monophosphate [AMP] and inorganic phosphate [Pi] and a fall in the concentration of phosphocreatine [PCr]; the main moving force for the elevation in mitochondrial ATP generation is the decline in [ATP]/[ADP] [Pi] (or equivalent) and consequent readjustments in the ratio of the intramitochondrial pyridine nucleotides [( NAD]m/[NADH]m). Direct stimulation of pyruvate dehydrogenase by calcium appears to be of secondary importance. It is concluded that synaptosomal Na/K pump is fueled primarily by oxidative phosphorylation and that a fall in [ATP]/[ADP][Pi] is the chief factor responsible for increased energy production. 相似文献
20.
Evidence that [3 H]Dopamine Is Taken Up and Released from Nondopaminergic Nerve Terminals in the Rat Substantia Nigra In Vitro 总被引:2,自引:0,他引:2
Potassium chloride (25 mM) and (+)-amphetamine (100 microM) both stimulated the release of radioactivity from slices of substantia nigra preincubated with [3H]3,4-dihydroxyphenylethylamine [( 3H]dopamine). Potassium chloride (25 mM) released radioactivity from slices of both zona compacta and zona reticulata. Prior 6-hydroxydopamine (6-OHDA) lesions of one nigrostriatal pathway did not reduce the spontaneous release of radioactivity, or the potassium chloride- or amphetamine-induced release of radioactivity from slices of nigra ipsilateral to the lesion after preincubation with [3H]dopamine. The accumulation of radioactivity following incubation of nigral slices from 6-OHDA-lesioned animals with [3H]dopamine was increased when compared to uptake into slices from intact tissue. In synaptosomal preparations of striatum, nomifensine but not desipramine or fluoxetine inhibited [3H]dopamine uptake. In contrast, nomifensine, desipramine, and fluoxetine all inhibited [3H]dopamine uptake in nigral synaptosomal preparations. Following 6-OHDA lesions of one nigrostriatal pathway the uptake of [3H]dopamine into nigral synaptosomal preparations was unchanged but uptake into striatal preparations was substantially decreased. In contrast, bilateral electrolesions of the dorsal and medial raphe nuclei reduced [3H]dopamine uptake into nigral preparations but not into striatal synaptosomes. The uptake of [3H]5-hydroxytryptamine ([3H]5-HT) into synaptosomal preparations of substantia nigra was abolished by fluoxetine and reduced by desipramine, but was unaffected by nomifensine. In contrast, fluoxetine, desipramine, and nomifensine all inhibited [3H]5-HT uptake into striatal synaptosomal preparations. Following 6-OHDA lesions of one nigro-striatal pathway the uptake of [3H]5-HT into nigral synaptosomal preparations was unchanged but uptake into striatal preparations was reduced.(ABSTRACT TRUNCATED AT 250 WORDS) 相似文献