Electrophysiological Characterization of Novel Effects of the Uptake-2 Blocker Decynium-22 (D-22) on Dopaminergic Neurons in the Substantia Nigra Pars Compacta.

Extracellular levels of dopamine (DA) and other monoamines in the brain depend not only on the classic transporters encoded by SLC6A gene family such as DAT, NET and SERT, but also a more recently identified group of low-affinity/high-capacity 'Uptake-2' transporters, mainly OCT3 and PMAT. The most frequently used pharmacological tool in functional studies of Uptake-2 is decynium-22 (D-22) known to block these transporters. However, the effectiveness of this drug in enhancing extracellular DA remains uncertain. Our aim was to test the hypothesis that D-22 increases extracellular levels of DA released from the somatodendritic region of dopaminergic neurons in the substantia nigra pars compacta (SNc) by reducing the OCT3/PMAT-dependent component of DA uptake. Extracellular DA was assessed indirectly, by evoking D2-IPSCs in SNc neurons following stimulated release of this neurotransmitter in midbrain slices obtained from mice. Recordings were conducted after partial inhibition of DAT with nomifensine, and after application of L-DOPA which increased the releasable DA pool. Contrary to our expectations, D-22 reduced, rather than increased, the amplitude of D2-IPSCs. Other effects included inhibition of GABA-IPSCs and I current, and a reduction in firing frequency of nigral neurons. These results show that in addition to the previously known non-specific inhibitory action on α1 adrenoceptors, D-22 exerts additional off-target effects by inhibiting dopaminergic and GABAergic synaptic transmission in the SNc and the spontaneous (pacemaker) activity of nigral neurons. It remains to be established if these novel effects contribute to a reduction in spontaneous locomotor activity reported in previous studies after systemic drug administration.