The Lignan (-)-Hinokinin Displays Modulatory Effects on Human Monoamine and GABA Transporter Activities Julie Marie V. Timple, †,§ Lizandra Guidi Magalha ̃ es, ‡ Karen Cristina Souza Rezende, ‡ Ana Carolina Pereira, ‡ Wilson Roberto Cunha, ‡ Ma ́ rcio Luis Andrade e Silva, ‡ Ole Valente Mortensen, † and Andre ́ ia C. K. Fontana* ,†,§ † Department of Pharmacology and Physiology, Drexel University College of Medicine, Philadelphia, Pennsylvania 19102, United States ‡ Nú cleo de Pesquisas em Ciê ncias Exatas e Tecnoló gicas, Universidade de Franca, CEP 14404-600, Franca, SP, Brazil * S Supporting Information ABSTRACT: The neurotransmitter transporters of the SLC6 family play critical roles in the regulation of neurotransmission and are the primary targets of therapeutic agents used to treat clinical disorders involving compromised neurotransmitter signaling. The dopamine and norepinephrine transporters have been implicated in clinical disorders such as attention deficit hyperactivity disorder (ADHD) and substance abuse. The GABA transporters (GATs) serve as a target for anxiolytic, antidepressant, and antiepileptic therapies. In this work, the interaction with neurotransmitter transporters was characterized for a derivative of the lignan (-)-cubebin (1), namely, (-)-hinokinin (2). Using in vitro pharmacological assays, 2 selectively inhibited the human dopamine and norepinephrine transporters, in a noncompetitive manner possibly mediated by binding to a novel site within the transporters, and displayed low affinity for the serotonin transporter. Compound 2 also specifically inhibited the GAT-1 GABA transporter subtype. Compound 2 is not a substrate of the carriers as it had no effect on the efflux of either of the neurotransmitters investigated. This compound is inactive toward glutamate and glycine transporters. These results suggest that 2 may serve as a tool to develop new therapeutic drugs for ADHD and anxiety that target the DAT, NET, and GAT-1 transporters. N eurotransmitter transporters play critical roles in the regulation of neurotransmission that modulates physio- logical responses involving homeostasis, stress response, behavior, motivation, and reward. 1,2 Under normal physio- logical processes, neurotransmitters are stored in vesicles in the presynaptic neuron and released into the synaptic cleft to activate or inhibit postsynaptic receptors. The downstream signaling of these neurotransmitters is terminated by reuptake of the substrate through its respective transporter. 3 Alteration in the regulation of extracellular neurotransmitter levels can lead to various neurological and mental health conditions, and subsequently, neurotransmitter transporters have become a target of interest for therapies altering neurotransmitter clearance. 4,5 Dopamine is involved in neurological processes of reward- driven learning and plays an important role in the normal functions of attention, mood, memory, and sleep. 6 Norepi- nephrine plays a role in learning, memory, attention, and pain perception. 7 The classical biogenic monoamine transporters, the human dopamine transporter (hDAT) and norepinephrine transporters (hNET), are critical for the regulation of dopamine and norepinephrine signaling, as they are responsible for the clearance of the respective neurotransmitters from the synaptic cleft, thereby terminating dopaminergic and norepinergic signaling. 8 Currently, about 5 million children ranging from ages 3 to 17 are diagnosed with attention deficit hyperactivity disorder (ADHD), a neurobehavioral disorder that has the DAT and NET as therapeutic targets for treatment. 9 Disruptions in the extracellular availability of dopamine and norepinephrine have been implicated in ADHD. 10 This deficiency can be overcome, through inhibition of DAT and NET, leading to a subsequent increase in substrate availability and reduction of symptoms related to ADHD. 11-13 The current therapies for this psychiatric condition consist primarily of the DAT and NET substrate amphetamine (Adderall) and the DAT and NET inhibitor and benzylpiperidine derivative methylphenidate (Ritalin). 14,15 Amphetamines act as carrier substrates by competing with the endogenous substrates and can also elicit efflux of dopamine or norepinephrine into the synaptic cleft from the presynaptic terminal. 5,16-18 Additionally, atomoxetine (Strattera) is a selective NET inhibitor that is used in the treatment of ADHD. 19 NET has also been implicated in various Received: June 6, 2013 Published: October 10, 2013 Article pubs.acs.org/jnp © 2013 American Chemical Society and American Society of Pharmacognosy 1889 dx.doi.org/10.1021/np400452n | J. Nat. Prod. 2013, 76, 1889-1895