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M E Reith

Publications and source records attributed to M E Reith.

At least 73 records · Page 4Linked to original sources
PubMed1994

GBR 12909 and 12935 block dopamine uptake into brain synaptic vesicles as well as nerve endings.

GBR 12909 and 12935, commonly used as potent neuronal dopamine uptake blockers, also inhibit dopamine uptake into brain synaptic vesicles. The concentrations required for the latter activity (34-45 nM) are one order of magnitude higher than those required for inhibiting neuronal uptake of dopamine (1-6 nM). In contrast, the two activities differ by three orders of magnitude for cocaine (137 microM versus 0.35 microM). We propose that the vesicular effect of GBR-type dopamine uptake blockers should be taken into account when interpreting in vivo experiments.

Animals↗
PubMed1994

[3H]WIN 35,428 binding to the dopamine uptake carrier. I. Effect of tonicity and buffer composition.

In the present study, the dopamine transporter on rat striatal membranes was labeled with [3H]WIN 35,428 (2 beta-carbomethoxy-3 beta-(4-fluorophenyl)-tropane), and its binding state or form was manipulated by changing tonicity and buffer composition. Binding to P2 membranes was enhanced by the presence of sucrose in the assay. This effect was not due solely to factors relating to tonicity because creation of isotonicity by dextrose or N-methyl-D-glucamine was less effective, and an increase in binding by sucrose was also observed in assays that were already isotonic by a mixture of sodium phosphate and NaCl. Under the latter conditions, fructose and mannose were equally effective as sucrose. Other important factors were the presence of sodium phosphate in the homogenizing buffer and the presence of sucrose during resuspension of the membranes. When P2 membranes were prepared from homogenates in 0.32 M sucrose, the effect of sucrose in the polytronning step or in the binding assay was restricted to a decrease in the Kd of the main binding component.

Animals↗
PubMed1994

[3H]WIN 35,428 binding to the dopamine uptake carrier. II. Effect of membrane fractionation procedure and freezing.

In the present study the dopamine transporter on rat striatal membranes was labeled with [3H]WIN 35,428 (2 beta-carbomethoxy-3 beta-(4-fluorophenyl)-tropane) and its binding state or form was compared between fresh and frozen tissue fractionated by various protocols. Freezing striatal tissue resulted in a decrease in binding to crude or P2 membranes, but did not consistently generate upward concave Scatchard plots. Among various conditions, sodium phosphate buffers containing sucrose, employed for homogenization and for measurement of binding to freshly prepared P2 membranes, were most likely to give multiple binding component resolutions. These results allow us to reconcile the existing literature on [3H]WIN 35,428 binding and to identify (1) the 'universal' binding component common to all studies thus far, (2) a high-affinity component measured with sufficiently low ligand concentration, and (3) a low-affinity component observed with sufficiently high ligand concentration.

Animals↗
PubMed1994

Effects of locally applied cocaine, lidocaine, and various uptake blockers on monoamine transmission in the ventral tegmental area of freely moving rats: a microdialysis study on monoamine interrelationships.

Microdialysis was used to compare the effect of local perfusion of cocaine with that of functionally similar compounds on extracellular norepinephrine, dopamine, and serotonin (measured simultaneously) in the ventral tegmental area of freely moving rats. Tetrodotoxin (1 microM) potently inhibited both basal and cocaine-induced dialysate monoamine outputs. The local anesthetic lidocaine produced little or no effect on the monoamine output, whereas all uptake blockers tested (at 0.1-1,000 microM) increased the monoamine output in a dose-dependent manner. The selective norepinephrine-uptake blockers desipramine and nisoxetine did not show any selectivity for norepinephrine, whereas the selective serotonin-uptake blockers fluoxetine and citalopram, as well as the selective dopamine-uptake blocker GBR 12935, preferentially (but not exclusively) increased their target amine. Cocaine at low concentrations (1-10 microM) increased the three amines similarly, but at higher concentrations (100-1,000 microM) caused a relatively higher dopamine output. A positive relationship between blocker-induced dialysate norepinephrine and dopamine outputs suggests significant interactions between monoamine systems. The present results indicate that cocaine's action in the ventral tegmental area involves not only a dopamine-, but also a norepinephrine- and a serotonin-related component, and that cocaine-induced monoamine increase is independent of its local anesthetic property.

Animals↗
PubMed1994

Isolation of a gametophyte-specific cDNA encoding a lipoxygenase from the red alga Porphyra purpurea.

A cDNA clone encoding a lipoxygenase was obtained from a subtracted cDNA library specific for the gametophyte of Porphyra purpurea. The amino acid sequence of the P. purpurea lipoxygenase is most similar to the sequences of animals and plants in the iron-binding, catalytic region located in the C-terminal half of the enzymes. Northern hybridization confirmed that the gene represented by this cDNA is expressed only in the gametophyte.

Amino Acid Sequence↗
PubMed1994

Structure-activity relationships for cocaine congeners in inhibiting dopamine uptake into rat brain synaptic vesicles and bovine chromaffin granule ghosts.

Structure-activity relationships of cocaine congeners were determined in inhibiting reserpine-sensitive, Mg++/ATP-dependent uptake of [3H]dopamine into rat brain synaptic vesicles under initial velocity conditions. The C2 carbomethoxy group could be deleted without loss of activity, whereas movement of this group from axial to equatorial orientation increased the potency. There was no need for the ester linkage between the tropane and phenyl rings, nor was there need for the ethylene bridge between C1 and C5 that makes cocaine a tropane instead of a piperidine structure. The structure-activity relationships were different from those for inhibiting neuronal amine transport or blocking voltage-dependent sodium channels. There was no correlation between block of uptake and degree of lipophilicity. The equally lipophilic compounds cocaine and pseudococaine, and WIN 35,065-3 and WIN 35,140, differed in uptake-blocking potency by an order of magnitude (137 vs. 22 microM and 65 vs. 4 microM, respectively). In bovine chromaffin granules, used as a less complex model system for the vesicular uptake system, the rank order of d-amphetamine, cocaine and pseudococaine in perturbing the proton gradient and in changing the membrane potential was different from that in inhibiting uptake. The inhibition of uptake is discussed in terms of the compounds acting as weak bases, transporter substrates or transporter blockers.

Animals↗
PubMed1994

Autoregulation and monoamine interactions in the ventral tegmental area in the absence and presence of cocaine: a microdialysis study in freely moving rats.

Extracellular levels of norepinephrine (NE), dopamine (DA) and serotonin (5-HT) were measured simultaneously by microdialysis in the ventral tegmental area (VTA) of conscious rats, and locomotor activity was monitored. Extracellular NE, DA and 5-HT was increased by both local infusion (30 microM) and i.p. injection (20 and 40 mg/kg) of cocaine with 5-HT responding most rapidly. DA was the only amine that showed a significantly higher increase to the systemic cocaine dose of 40 mg/kg than 20 mg/kg, and a higher response to local cocaine (30 microM) than systemic cocaine (20 mg/kg). In the following experiments, the monoamine autoreceptor antagonists idazoxan (alpha-2, 100 microM), sulpiride (D2 DA/D3 DA, 10 microM) and methiothepin (5-HT1/5-HT2, 20 microM) were focally applied into the VTA before cocaine application. Idazoxan or methiothepin increased only local cocaine-induced NE or 5-HT output, whereas sulpiride increased both local and systemic cocaine-induced DA output, consonant with the importance of somatodendritic monoamine autoreceptors in addition to accumbens-VTA feedback pathways. Both idazoxan and methiothepin increased the basal output of all three amines without modifying cocaine (30 microM or 20 mg/kg)-induced output of DA/5-HT or NE/DA, whereas sulpiride promoted cocaine-induced NE output without modifying basal NE/5-HT output and cocaine-induced 5-HT output, implying complex interactions between monoamines. Idazoxan or methiothepin depressed, whereas sulpiride stimulated cocaine (20 mg/kg)-induced motor activity. The analysis of behavioral/neurochemical relationships revealed a negative correlation between dialysate NE output and motor activity in the cocaine alone and idazoxan/cocaine groups, and a positive correlation between dialysate DA output and motor activity in the sulpiride/cocaine and methiothepin/cocaine groups.

Animals↗
PubMed1993

Cationic and anionic requirements for the binding of 2 beta-carbomethoxy-3 beta-(4-fluorophenyl)[3H]tropane to the dopamine uptake carrier.

The present study reports the ion dependency of 2 beta-carbomethoxy-3 beta-(4-fluorophenyl)[3H]tropane ([3H]CFT) binding to the dopamine transporter in the rat striatum. The results indicate that [3H]CFT binding to synaptosomal P2 membranes requires low concentrations of Na+ (peak binding between 20 and 50 mM Na+), is stimulated by phosphate anion or I-, but is unaffected or only slightly affected by F-, Cl-, Br-, NO3-, or SO(4)2-. Concentrations of Na+ of > 50 mM become inhibitory except in the presence of I-, which shifts peak binding levels toward higher Na+ concentrations and also elevates the peak binding level. K+ strongly decreased [3H]CFT binding with a shallow inhibition curve, and Na+ could not overcome this effect. Saturation analysis of [3H]CFT binding revealed a single binding site changing its affinity for CFT depending on the concentration of sodium phosphate buffer (6, 10, 30, 50, 130, or 200 mM; 1 mM plus 49 mM NaCl versus 10 mM plus 40 mM NaCl; or 1 mM plus 129 mM NaI versus 10 mM plus 120 mM NaI). No differences were observed in the density of CFT binding sites between any of the conditions examined.

Animals↗
PubMed1993

[3H]dopamine and [3H]serotonin release in vitro induced by electrical stimulation in A9 and A10 dopamine regions of rat brain: characterization and responsiveness to cocaine.

The present study investigated [3H]dopamine (DA) and [3H]5-hydroxytryptamine (5-HT) release evoked by electrical stimulation in superfused ventral mesencephalon (VM) slices containing A9 and A10 DA neurons. Electrically induced [3H]DA release from VM was, at least, of two origins: one was from DA somatodendrites, regulated by DA autoreceptors, and increased by DA uptake blockers; another was from 5-HT terminals, modulated by 5-HT autoreceptors, and could be minimized by the copresence of fluoxetine during the labeling of the slices. Release of both origins was Ca(++)-dependent and tetrodotoxin-sensitive. Cocaine (10 and 100 microM) modestly increased electrically induced [3H]DA release from DA somatodendrites and concentration-dependently inhibited that from 5-HT terminals. Electrically induced [3H]5-HT release from VM was exclusively from 5-HT terminals, dependent on Ca++, partly blocked by tetrodotoxin, increased by 5-HT uptake blockers and regulated by 5-HT autoreceptors. The autoregulation of [3H]5-HT release partly counteracted the stimulatory effect of cocaine (10 microM) on [3H]5-HT release. Inhibition of 5-HT uptake by cocaine reduced the effectiveness of 5-methoxytryptamine to suppress electrically induced [3H]5-HT release. No evidence was found to support the notion that postsynaptic 5-HT receptors modulate somatodendritic DA release. 5-HT facilitated spontaneous [3H]DA release from VM via a 5-HT/DA transporter-dependent exchange process and inhibited electrically induced [3H]DA release from 5-HT terminals via a 5-HT autoreceptor-mediated mechanism. The dual effects of cocaine on electrically induced [3H]DA release from VM may be attributed to the complex 5-HT/DA and 5-HT autoreceptor/cocaine interactions as well as the relative densities of 5-HT/DA transporters in this region, indicating a possible involvement of the 5-HT system in cocaine's behavioral effects.

Animals↗
PubMed1993

Dopamine and serotonin release-regulating autoreceptor sensitivity in A9/A10 cell body and terminal areas after withdrawal of rats from continuous infusion of cocaine.

The effects of dopamine (DA) and 5-hydroxytryptamine (5-HT) autoreceptor agents on electrically induced [3H]DA and [3H]5-HT release from superfused slices of striatum, nucleus accumbens and ventral mesencephalon (VM) containing A9 and A10 neurons were investigated in rats made tolerant to the stimulatory effect of cocaine on locomotor behavior by a 14-day continuous infusion of cocaine (29 mg/kg/day) by s.c. implanted osmotic minipumps followed by a 7-day drug-free period. In VM, electrically induced [3H]DA was increased, the ability of pergolide to inhibit this release was abolished, but the ability of sulpiride to facilitate the release was potentiated, implicating a higher concentration of synaptic DA modifying the responsiveness of somatodendritic D2 autoreceptors to D2 agents. Both electrically induced [3H]5-HT release from VM and the stimulatory effect of in vitro cocaine on this release were enhanced whereas the effects of both 5-methoxytryptamine and methiothepin were attenuated, indicating that subsensitivity of 5-HT autoreceptors developed in DA cell body regions. In striatum and nucleus accumbens, no significant changes were observed in [3H]DA and [3H]5-HT release, except for a modest reduction in the effects of both pergolide and sulpiride on electrically induced [3H]DA release from striatum. These results emphasize the importance of pretreatment-induced changes in DA cell body regions, rather than terminal areas, under the present conditions. The observed increase in DA autoinhibitory tone and subsensitivity of 5-HT release-regulating autoreceptors in the VM may contribute to the locomotor tolerance upon cocaine challenge after continuous cocaine.

Animals↗
PubMed1992

Evidence for mutually exclusive binding of cocaine, BTCP, GBR 12935, and dopamine to the dopamine transporter.

The present study addressed the possibility that there are distinct but allosterically interacting populations of binding sites for dopamine/cocaine and BTCP/GBR (N-[1-(2-benzo[b]thiophenyl)cyclohexyl]piperidine/1-(2-diphenylmethox y) - ethyl]-4-(3-phenylpropyl)piperazine) (selective dopamine uptake blockers) on the dopamine transporter in the rat striatum. Dopamine uptake sites were labeled in vitro with the cocaine analog [3H]CFT (2 beta-carbomethoxy-3 beta-(4-fluorophenyl)-tropane), and the inhibition of binding by CFT or cocaine was measured. A graphic method was adopted for studying shifts in inhibitory potency resulting from the addition of a second compound. Under the conditions used, the co-presence of dopamine, GBR 12935, or BTCP decreased the inhibitory potency of CFT or cocaine to the extent predicted by a model in which all compounds bind to the same site or the binding of all compounds is mutually exclusive. No evidence for negative allosteric interactions between CFT and BTCP was found in experiments comparing inhibition of [3H]CFT binding by BTCP at a low and high concentration of [3H]CFT.

Animals↗
PubMed1992

Determination of radioligand specific activity using competition binding assays.

Radioligand binding assays are routinely utilized in laboratories throughout the world to study receptors and their related binding sites, carrier proteins, and enzymes. To accurately estimate equilibrium binding parameters, such as the equilibrium dissociation constant (Kd) and maximal number of binding sites (Bmax), the investigator must know the correct value of the specific activity of the radioligand. If the specific activity is overestimated the Kd and Bmax values will be underestimated, while underestimation of the specific activity results in an overestimation of the Kd and Bmax. The present communication describes a simple and rapid method for determining the specific activity of a radioligand using homologous competition binding assays. Performing the competition assays at two or more different concentrations of the radioligand allows the specific activity to be determined from the IC50 values without the need of analytical methods to quantify minute amounts of the radioligand. In addition to providing the specific activity, use of this method estimates the Kd for the radioligand. This method was utilized to determine the specific activity and Kd for two blockers of the dopamine uptake carrier, [3H]GBR-12935 and [3H]-CFT, which share a common binding site in the striatum.

Animals↗
PubMed1992

Dopamine releasing effect of phenylbiguanide in rat striatal slices.

The present study explored the mechanisms underlying the dopamine releasing effect of phenylbiguanide, a compound commonly used as a 5-HT3 receptor agonist. Phenylbiguanide, and also serotonin and 2-methyl-serotonin, enhanced the outflow of radioactivity from superfused rat striatal slices preloaded with [3H]dopamine. The presence of the dopamine uptake blocker nomifensin prevented the increase in outflow. The effect of phenylbiguanide was not antagonized by 5-HT3 receptor antagonists, did not require the presence of Ca2+ in the superfusion buffer, and also occurred in reserpinized preparations with depleted dopamine stores. Phenylbiguanide caused a greater shift in the distribution of superfusate radioactivity from DOPAC to dopamine than did nomifensin. All these results are in agreement with an exchange mechanism by which phenylbiguanide promotes the efflux of dopamine by operation of the uptake carrier in the reversed direction. In consonance, phenylbiguanide, and also serotonin and 2-methyl-serotonin, inhibited the binding of [3H]CFT to dopamine uptake sites, although the rank order for promoting outflow, serotonin greater than phenylbiguanide greater than 2-methyl-serotonin, differed from that for inhibiting [3H]CFT binding to dopamine uptake sites, 2-methylserotonin approximately serotonin greater than phenylbiguanide. The present results raised the possibility that phenylbiguanide has an additional activity in releasing vesicular dopamine into the cytoplasmic pool.

Animals↗
PubMed1992

Radiolabeling of dopamine uptake sites in mouse striatum: comparison of binding sites for cocaine, mazindol, and GBR 12935.

This study addressed the possibility of a unique binding interaction between cocaine and the dopamine transporter as compared with other blockers of dopamine uptake. Cocaine binding sites in a fresh P2 fraction of mouse striatum were labeled with [3H]CFT, a phenyltropane analog of cocaine also known as WIN 35,428, and compared with sites labeled with [3H]mazindol or [3H]GBR 12935. Under the conditions used, homogeneous binding was observed that was inhibited monophasically by cocaine, CFT, and mazindol; the same potencies were observed with the three radioligands. Saturation analysis in the presence and in the absence of unlabeled inhibitor (CFT, mazindol, cocaine) indicated a change in the Kd but not the Bmax, consonant with a competitive mechanisms. Tris-HCl reduced the affinity of each radioligand and unlabeled inhibitor without changing the Bmax. N-Ethylmaleimide reduced the binding of all radioligands equally and cocaine offered protection. The dissociation rate of [3H]CFT and [3H]mazindol binding was not affected by the presence of mazindol and CFT, respectively. The Bmax of [3H]CFT and [3H]mazindol binding was the same; the relatively higher value for [3H]GBR 12935 binding in analyses involving varying tritiated GBR 12935 only, was due primarily to an underestimation of the specific activity of [3H]GBR 12935. All results are in agreement with a one-site model in which cocaine, CFT, mazindol, and GBR 12935 share a common binding site in mouse striatum.

Animals↗
PubMed1992

Cocaine binding sites in mouse striatum, dopamine autoreceptors, and cocaine-induced locomotion.

BALB/cByJ mice received cocaine (25 mg/kg IP) once a day for 3 days, resulting in a greater locomotor response to cocaine on day 3 than on day 1. On day 4, a dose (0.03 mg/kg SC) of apomorphine, targeted at dopamine autoreceptors, caused the same degree of locomotor depression in cocaine- as in saline-pretreated mice. In addition, no change was found in either the affinity or density of cocaine binding sites in their striatum as measured by the binding of [3H]CFT. C57BL/6ByJ, mice displayed a greater locomotor response to cocaine than BALB/cByJ mice, but had the same number of striatal [3H]CFT binding sites with the same affinity. Factors other than striatal cocaine binding sites, or dopamine autoreceptors as measured by apomorphine-induced depression of locomotion, should be considered for the explanation of the enhancement of the locomotor response upon daily cocaine administration in BALB/cByJ mice, or for the different locomotor response to cocaine of this strain compared with the C57BL/6ByJ strain of mice.

Aging↗
PubMed1991

Carrier-mediated efflux of [3H]dopamine and [3H]1-methyl-4-phenylpyridine: effect of ascorbic acid.

The carrier-mediated efflux of [3H]1-methyl-4-phenylpyridine (MPP+) and [3H]dopamine was examined in mouse striatal synaptosomal P2 fractions. Although the two compounds are transported by the same carrier, the translocation of the carrier-ligand complex is more rapid with MPP+ than with dopamine. With dopamine-stimulated efflux of preloaded [3H]dopamine, externally present dopamine at a concentration of 1.3 microM reduced the intrasynaptosomal concentration of [3H]dopamine by 50% (the ECR value) with 8 min of incubation. The ECR value of dopamine in promoting the efflux of [3H]MPP+, however, was only 0.15 microM. Similarly, ascorbic acid was weaker in enhancing the efflux of [3H]dopamine (ECR greater than 2000 microM) than that of [3H]MPP+ (ECR = 567 microM). This effect of ascorbic acid on the efflux of [3H]MPP+ was attenuated by mazindol, a blocker of dopamine uptake. It is proposed that ascorbic acid has a neuromodulatory role involving changes at the level of carrier-membrane translocation and/or orientation.

1-Methyl-4-phenylpyridinium↗
PubMed1991

Sertraline and cocaine-induced locomotion in mice. I. Acute studies.

The present study assessed the behavioral and pharmacokinetic interaction between the serotonin uptake blocker sertraline and cocaine in C57BL/6ByJ mice. Pretreatment with sertraline (1-32 mg/kg IP) did not affect the total amount of spontaneous locomotor activity during 50 min following administration of cocaine (15-40 mg/kg IP). At doses of sertraline (16 and 32 mg/kg) much higher than those found to inhibit ex vivo neuronal uptake of serotonin by 50% (1-2 mg/kg), the peak of cocaine-induced locomotor activity was shifted towards a later time. A similar effect was seen after pretreatment with serotonin uptake blockers other than sertraline, and also after desipramine. Sertraline (16 and 32 mg/kg), given 60 min prior to cocaine, did not affect levels of cocaine in brain and plasma, and cocaine administration did not alter the brain level of sertraline. Although female mice were more responsive to cocaine than male mice, they were not different in their response to sertraline.

1-Naphthylamine↗