Calcium-dependent inhibition of synaptosomal serotonin transport by the α₂-adrenoceptor agonist 5-bromo-N-[4,5- dihydro-1H-imidazol-2-yl]-6-quinoxalinamine (UK14304)

Termination of serotonergic transmission is the function of the plasma membrane 5-hydroxytryptamine (serotonin, 5-HT) transporter (SERT), which is also a high-affinity target in vivo for antidepressants, amphetamines, and cocaine. Studies show that SERT is regulated by protein kinase- and phosphatase-linked pathways. In contrast, receptor-linked modulation of SERT is only minimally defined. Because noradrenergic stimulation is reported to influence 5-HT release, we explored possible presynaptic adrenoceptor-mediated regulation of SERT. In mouse forebrain synaptosomes, α₂-adrenoceptor agonists, particularly 5-bromo-N-[4,5-dihydro-1H-imidazol-2-yl]-6-quinoxalinamine (UK14304), triggered a concentration- and time-dependent decrease in 5-HT transport. In contrast, 5-HT up-take was unaffected by pharmacological α₁-adrenoceptor activation. Kinetically, UK14304 significantly decreased the apparent substrate affinity, K_m without altering transport capacity, V_max. At concentrations of UK14304 supporting maximal inhibition of SERT in synaptosomes, no effect on SERT in transfected cells was observed, suggesting that UK14304 acts indirectly to reduce SERT activity. The effect of UK14304 on 5-HT uptake was not shared by other Na⁺ and Cl^--dependent transporters. UK14304-mediated inhibition of SERT function was yohimbine-sensitive, as was inhibition triggered by norepinephrine, and was abolished in the absence of added Ca²⁺. Moreover, UK14304 effects were attenuated by voltage-sensitive Ca²⁺ channel antagonists, consistent with a role for Ca²⁺ in UK14304 effects. In agreement with altered 5-HT transport activity in vitro, in vivo chronoamperometry studies revealed that UK14304 significantly prolonged 5-HT clearance. Our findings suggest that UK14304 modulates-SERT function in vitro and in vivo via signaling pathways, possibly supported by an influx of Ca²⁺ through voltage-sensitive Ca²⁺ channels.