4). This argues for levamisole-mediated inhibition of reuptake of continuously released substrate rather than for a true releasing action. We previously observed similar spurious

releasing effects Galunisertib with the selective serotonin reuptake inhibitor paroxetine on HEK293-cells expressing SERT (Scholze et al., 2000). To our knowledge, the experiments show for the first time that levamisole directly inhibits the human NET and to a lesser extent SERT and DAT. This inhibition is mediated by a low-affinity interaction with the same site, to which cocaine is bound and thus the SI site. Administration of levamisole to race horses resulted in positive doping tests, because their urine contained aminorex (Barker, 2009). The metabolism of levamisole to the amphetamine-like compound aminorex was later confirmed to also occur in dogs and humans (Bertol et al., 2011 and Hess et al., 2013). Hence, for the sake of comparison, we quantified the inhibition by aminorex of substrate uptake by NET, SERT or

DAT (Fig. 5A). Interestingly, aminorex also preferentially blocked substrate uptake by NET (IC50: 0.33 ± 1.07 μM) and DAT (IC50: 0.85 ± 1.20 μM), while SERT was inhibited only at 20-fold higher concentrations (IC50: 18.39 ± 1.12 μM). Accordingly, the pattern of inhibition (NET > DAT >>> SERT) was reminiscent of the parent compound levamisole, but the inhibitory potency of aminorex was comparable to that of cocaine. To investigate if cocaine has an allosteric modulatory effect on aminorex, we performed uptake-inhibition experiments selleck at increasing concentrations of aminorex in presence of fixed cocaine concentrations

(Fig. 6). The resulting Dixon plots indicated that aminorex and cocaine bound in a mutually exclusive manner. In other words, there was not any appreciable allosteric modulatory effect in SERT, NET or DAT. Aminorex is classified as an amphetamine-like substance, because it is chemically related to amphetamine and it suppresses feeding behavior in a manner similar to amphetamines. However, the neurochemical changes induced by aminorex differ from those of other appetite suppressants (Roszkowski and Kelley, 1963 and Zheng et al., 1997). We therefore Linifanib (ABT-869) investigated its effects on substrate efflux by carrying out superfusion experiments in the presence and absence of monensin (10 μM). Interestingly, aminorex induced significant substrate release only in HEK293-SERT cells whereas efflux was completely absent in HEK293-DAT cells. HEK293-NET cells displayed only a slight response (Fig. 5B-D). Importantly, monensin enhanced efflux as predicted for an amphetamine-like releaser (Scholze et al., 2000). Taken together our experimental data showed that aminorex modulates the neurotransmitter transporters in different ways.