Effects of methamphetamine and lobeline on vesicular monoamine and dopamine transporter-mediated dopamine release in a cotransfected model system

Clare Wilhelm, Robert A. Johnson, Paul G. Lysko, Amy J. Eshleman, Aaron Janowsky

Research output: Contribution to journalArticle

23 Citations (Scopus)

Abstract

Dopamine (DA) retention and drug-induced release kinetics were characterized in human embryonic kidney (HEK)-293 cells stably coexpressing the human DA transporter (hDAT) and human vesicular monoamine transporter (hVMAT2). Cofunction of hDAT and hVMAT2 caused greater retention of [3H]DA at 20 min (37°C), or 45 min (22°C) compared with cells that were treated with dihydrotetrabenazene (DHTB) to block the hV-MAT2. In hDAT- and hVMAT2-coexpressing cells treated with DHTB during [3H]DA loading, methamphetamine (METH)-induced efflux was only 20% of preloaded [ 3H]DA, compared with 50 to 60% efflux in the absence of DHTB. Interestingly, the presence of DHTB (during release only) increased the potency and efficacy of METH at inducing [3H]DA release (without DHTB: EC50 = 33.8 μM, maximal release 51%; release with DHTB: EC 50 = 3.2 μM, maximal release 61%), suggesting that the effects of METH and DHTB on vesicular storage are additive. High concentrations of lobeline induced a statistically significant release of [3H]DA from HEK-hDAT-hVMAT2 cells, but only in the absence of DHTB, suggesting an hVMAT2-mediated effect. Likewise, lobeline did not induce a significant release of [3H]DA from HEK-hDAT cells. The substrates DA and p-tyramine induced robust release of preloaded [3H]DA from cotransfected cells. Cocaine was somewhat effective at blocking substrate-induced [3H]DA efflux. These results suggest that coexpression of the hDAT and hVMAT2 can be used as a model system to distinguish functional pools of DA and to quantify differences in drug effects on DA disposition. In addition, cotransfected cells can be used to determine mechanisms of simultaneous drug interactions at multiple sites.

Original languageEnglish (US)
Pages (from-to)1142-1151
Number of pages10
JournalJournal of Pharmacology and Experimental Therapeutics
Volume310
Issue number3
DOIs
StatePublished - Sep 2004

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Lobeline
Vesicular Monoamine Transport Proteins
Dopamine Plasma Membrane Transport Proteins
Methamphetamine
Dopamine
Kidney
Dopamine Agents
Tyramine
Drug Interactions
Cocaine

ASJC Scopus subject areas

  • Pharmacology

Cite this

Effects of methamphetamine and lobeline on vesicular monoamine and dopamine transporter-mediated dopamine release in a cotransfected model system. / Wilhelm, Clare; Johnson, Robert A.; Lysko, Paul G.; Eshleman, Amy J.; Janowsky, Aaron.

In: Journal of Pharmacology and Experimental Therapeutics, Vol. 310, No. 3, 09.2004, p. 1142-1151.

Research output: Contribution to journalArticle

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title = "Effects of methamphetamine and lobeline on vesicular monoamine and dopamine transporter-mediated dopamine release in a cotransfected model system",
abstract = "Dopamine (DA) retention and drug-induced release kinetics were characterized in human embryonic kidney (HEK)-293 cells stably coexpressing the human DA transporter (hDAT) and human vesicular monoamine transporter (hVMAT2). Cofunction of hDAT and hVMAT2 caused greater retention of [3H]DA at 20 min (37°C), or 45 min (22°C) compared with cells that were treated with dihydrotetrabenazene (DHTB) to block the hV-MAT2. In hDAT- and hVMAT2-coexpressing cells treated with DHTB during [3H]DA loading, methamphetamine (METH)-induced efflux was only 20{\%} of preloaded [ 3H]DA, compared with 50 to 60{\%} efflux in the absence of DHTB. Interestingly, the presence of DHTB (during release only) increased the potency and efficacy of METH at inducing [3H]DA release (without DHTB: EC50 = 33.8 μM, maximal release 51{\%}; release with DHTB: EC 50 = 3.2 μM, maximal release 61{\%}), suggesting that the effects of METH and DHTB on vesicular storage are additive. High concentrations of lobeline induced a statistically significant release of [3H]DA from HEK-hDAT-hVMAT2 cells, but only in the absence of DHTB, suggesting an hVMAT2-mediated effect. Likewise, lobeline did not induce a significant release of [3H]DA from HEK-hDAT cells. The substrates DA and p-tyramine induced robust release of preloaded [3H]DA from cotransfected cells. Cocaine was somewhat effective at blocking substrate-induced [3H]DA efflux. These results suggest that coexpression of the hDAT and hVMAT2 can be used as a model system to distinguish functional pools of DA and to quantify differences in drug effects on DA disposition. In addition, cotransfected cells can be used to determine mechanisms of simultaneous drug interactions at multiple sites.",
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