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Amphetamine actions at the serotonin transporter rely on the availability of phosphatidylinositol-4,5-bisphosphate

Author:
Buchmayer, Florian, Schicker, Klaus, Steinkellner, Thomas, Geier, Petra, Stübiger, Gerald, Hamilton, Peter J., Jurik, Andreas, Stockner, Thomas, Yang, Jae-Won, Montgomery, Therese, Holy, Marion, Hofmaier, Tina, Kudlacek, Oliver, Matthies, Heinrich J. G., Ecker, Gerhard F., Bochkov, Valery, Galli, Aurelio, Boehm, Stefan, Sitte, Harald H.
Source:
Proceedings of the National Academy of Sciences of the United States of America 2013 v.110 no.28 pp. 11642-11647
ISSN:
0027-8424
Subject:
amphetamine, binding sites, extracellular space, humans, ion channels, mutants, mutation, nerve tissue, norepinephrine, phospholipase C, rats, serotonin, transporters
Abstract:
Nerve functions require phosphatidylinositol-4,5-bisphosphate (PIP ₂) that binds to ion channels, thereby controlling their gating. Channel properties are also attributed to serotonin transporters (SERTs); however, SERT regulation by PIP ₂ has not been reported. SERTs control neurotransmission by removing serotonin from the extracellular space. An increase in extracellular serotonin results from transporter-mediated efflux triggered by amphetamine-like psychostimulants. Herein, we altered the abundance of PIP ₂ by activating phospholipase-C (PLC), using a scavenging peptide, and inhibiting PIP ₂-synthesis. We tested the effects of the verified scarcity of PIP ₂ on amphetamine-triggered SERT functions in human cells. We observed an interaction between SERT and PIP ₂ in pull-down assays. On decreased PIP ₂ availability, amphetamine-evoked currents were markedly reduced compared with controls, as was amphetamine-induced efflux. Signaling downstream of PLC was excluded as a cause for these effects. A reduction of substrate efflux due to PLC activation was also found with recombinant noradrenaline transporters and in rat hippocampal slices. Transmitter uptake was not affected by PIP ₂ reduction. Moreover, SERT was revealed to have a positively charged binding site for PIP ₂. Mutation of the latter resulted in a loss of amphetamine-induced SERT-mediated efflux and currents, as well as a lack of PIP ₂-dependent effects. Substrate uptake and surface expression were comparable between mutant and WT SERTs. These findings demonstrate that PIP ₂ binding to monoamine transporters is a prerequisite for amphetamine actions without being a requirement for neurotransmitter uptake. These results open the way to target amphetamine-induced SERT-dependent actions independently of normal SERT function and thus to treat psychostimulant addiction.
Agid:
1737270