Discrimination of Monoamine Uptake by Membranes of Adrenal Chromaffin Granules

Prada, M. Da; Obrist, R.; Pletscher, A.

doi:10.1111/j.1476-5381.1975.tb07357.x

Cited by 42 publications

(16 citation statements)

References 23 publications

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“…Johnson et al (14) have also recently shown isoproterenol to be taken up by chromaffin granules in an ATP-activated, reserpine-sensitive manner, just like natural substrates. However, the detailed kinetic properties of isoproterenol have not been extensively evaluated (4,(14)(15)(16).In the present study we report that Me2E is indeed a classical competitive inhibitor of (R)-norepinephrine transport. However, we also found that the inhibition constant was extremely high and that [3H]Me2E was not transported.…”

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confidence: 67%

“…Chromaffin granules were prepared by differential centrifugation in 0.3 M sucrose as described in earlier studies from this laboratory (17,18). The purified granules were maintained on ice at a concentration equivalent to an optical density of 5.0 at 540 nm for no more than 15 min prior to the initiation of the experiment.…”

Section: Methodsmentioning

confidence: 99%

Section: Fig 1 Structures Of Me2e (A) and Isoproterenol (B) [3h]mementioning

confidence: 99%

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Chemical evidence that catecholamines are transported across the chromaffin granule membrane as noncationic species.

Ramu¹,

Levine²,

Pollard³

1983

Proc. Natl. Acad. Sci. U.S.A.

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Catecholamines are transported into chromaffin granules by a Mg2+/ATP-driven process under conditions in which the substrate exists primarily as a positively charged or neutral species. In order to distinguish between these two states, we studied the transport properties of a permanently charged quaternary analogue of epinephrine, (R,S)-dimethylepinephrine. We found that this compound was a classical competitive inhibitor of (R)-[3H]norepinephrine uptake, with a K; of 3.8 mM for the racemic form, or 1.9 mM for the R form. However, the [3H]dimethylepinephrine was not transported at all into granules. Our control for steric hindrance as an explanation for deficient translocation was analysis of the transport properties of isoproterenol, a secondary catecholamine with an isopropyl group around the amine residue. (R)-Isoproterenol was an effective competitive inhibitor of (R)-[3H]norepinephrine transport, with a K; of 91 ,uM. In contrast to dimethylepinephrine, (RS)-[3H]isoproterenol was clearly translocated across the granule membrane, with a Km of 123 aiM, or 61.5 IAM for the R isomer. Thus, the positive charge on dimethylepinephrine and not the size of the amine moiety appeared to be responsible for the lack of translocation. We interpret these data to indicate that, although the positively charged species can interact with the transport site, an uncharged species is the one actually transported.Catecholamines are transported into chromaffin granules by a Mg2+/ATP-driven process, which has become increasingly attractive as a general and simple system to analyze chemiosmotic energy coupling to transport (1-5). Many models have been proposed to explain the process, but distinguishing among them has been made difficult by lack of knowledge of whether the neutral, cationic, or anionic catecholamine species is the true substrate (4,(6)(7)(8)(9). Recently, on the basis of observations that both the binding constants for dopamine and serotonin and the calculated cationic fractions were constant between pH 6.8 and 7.6, Knoth et al. (10) concluded that the cationic species was translocated. However, using a similar experimental approach, Scherman and Henry (11) came to the opposite conclusion. They observed that the Km for norepinephrine declined between pH 6.5 and 8.5, and they interpreted this as due to an increase in the concentration of the uncharged species, the proper substrate. Yet, on the basis of entirely different considerations, Johnson and Scarpa (7) initially claimed that the species transported was most likely neutral, and more recently summarized by saying that a conclusion was not yet possible (8).We therefore decided to reinvestigate this question directly by examining the transport properties of a quaternary analogue of epinephrine. The compound we used was (R,S)-dimethylepinephrine (Me2E). * Me2E and its tritium-labeled derivative have the structure shown in Fig. 1 FIG. 1. Structures of Me2E (A) and isoproterenol (B). [3H]Me2E waslabeled at the p carbon (*). The wavy line on isoproterenol in ...

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confidence: 67%

Section: Methodsmentioning

confidence: 99%

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Chemical evidence that catecholamines are transported across the chromaffin granule membrane as noncationic species.

Ramu¹,

Levine²,

Pollard³

1983

Proc. Natl. Acad. Sci. U.S.A.

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“…Vesicles have been found to store most, if not all, of the amines recognized as neurotransmitters; the affinity of the vesicle for a foreign amine is usually similar to the affinity for the native amine (for example, Da Prada & Pletscher, 1969;Phillips, 1974;Da Prada, Obrist & Pletscher, 1975). Our experiments constitute the first in vivo study in which uptake 'FAST A KONAL TRANSPORT OF FOREIGN TRANSMITTERS 269 across the plasma membrane has been circumvented.…”

Section: Selectivity Of Uptake Into Vesictesmentioning

confidence: 89%

Fast axonal transport of foreign transmitters in an identified serotonergic neurone of Aplysia californica.

Goldberg¹,

Schwartz²

1980

The Journal of Physiology

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4. Injection of large amounts of dopamine and octopamine reduced the export into the axon of [3H]serotonin injected into the same cell. Large amounts of choline did not reduce export of [3H]serotonin. We conclude that the biogenic amines compete with serotonin for the vesicular storage site and that they move by fast transport because they are sequestered by the serotonergic storage vesicle.5. The specificity of uptake into the storage vesicle as assayed with this in vivo system is similar to the specificity of uptake into aminergic vesicles as previously studied in vitro.

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“…The isolated membranes of adrenal chromaffin granules were used as a model system since it has not yet been possible to obtain pure amine storage organelles from the brain. Chromaffin granules are available in a highly purified form, and their membranes have been shown to take up biogenic amines by a specific, probably carriermediated adenosine-5'-triphosphate (ATP)-dependent transport mechanism (Taugner & Hasselbach, 1966;Taugner, 1971;Kirshner, 1974;Da Prada, Obrist & Pletscher, 1975 (Helle, 1971;. Previous …”

Section: Introductionmentioning

confidence: 99%

Effect of Neuroleptics and Other Drugs on Monoamine Uptake by Membranes of Adrenal Chromaffin Granules

Pletscher

1977

British J Pharmacology

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The effects have been investigated of various reserpine‐like, neuroleptic, antidepressant and other compounds on the adenosine‐5′‐triphosphate (ATP)‐dependent uptake of noradrenaline (NA) (reserpine‐sensitive) and tryptamine (reserpine‐resistant) by membranes of isolated chromaffin granules of bovine adrenal medulla. Reserpine and Ro 4–1284 (2‐hydroxy‐2‐ethyl‐3‐isobutyl‐9,10‐dimethoxy‐hexahydro‐11bH‐benzo(a)quinolizine) as well as neuroleptics (e.g. chlorpromazine and haloperidol) inhibited the NA uptake, but the reserpine‐like drugs were more potent. In contrast, Ro 4–1284 showed a considerably weaker effect than the neuroleptics in interfering with tryptamine uptake. Chlorpromazine had about the same potency in inhibiting NA and tryptamine uptake, whereas the action of haloperidol was more pronounced on the uptake of NA than of tryptamine. The relative potencies of neuroleptic drugs in inhibiting NA uptake by granule membranes in vitro corresponded only partly to their relative potencies in enhancing dopamine turnover in vivo. The inhibition of NA uptake by chlorpromazine and Ro 4–1284 appeared to be of the noncompetitive type. Chlorpromazine did not influence the decrease in ATP induced by granule membranes in the incubation medium. Other basic, but not acidic compounds also inhibited NA uptake by granule membranes; their potency was of the order of that of chlorpromazine (antidepressants) or weaker (e.g. benzodiazepines). In conclusion, the mechanism of action of neuroleptics probably differs from that of reserpine‐like drugs in the inhibition of monoamine uptake by membranes of catecholamine storage organelles. While interference with the granular storage of dopamine at the granule membrane level may contribute to the in vivo action of neuroleptics (e.g. in enhancing dopamine turnover), additional effects of these drugs must be involved in vivo, e.g. blockade of pre‐ and postsynaptic dopamine receptors.

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Discrimination of Monoamine Uptake by Membranes of Adrenal Chromaffin Granules

Cited by 42 publications

References 23 publications

Chemical evidence that catecholamines are transported across the chromaffin granule membrane as noncationic species.

Chemical evidence that catecholamines are transported across the chromaffin granule membrane as noncationic species.

Fast axonal transport of foreign transmitters in an identified serotonergic neurone of Aplysia californica.

Effect of Neuroleptics and Other Drugs on Monoamine Uptake by Membranes of Adrenal Chromaffin Granules

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