Serotonin and norepinephrine transporters: possible relationship between oligomeric structure and channel modes of...

In the past several years there has been significant progress made on the biophysics of neurotransmitter transporters, leading to the proposal of new models of substrate and ion permeation across membranes. Questions arising from these studies are as follows: How are substrate uptake and substrate-induced current related? Where and how does substrate-ion coupling occur? What is the functional significance of the coupled and uncoupled currents? Because of a long-standing interest and collaboration, and because of their importance for normal function and disease, the authors have focused on the properties of human norepinephrine and serotonin transporters, using other clones and mutations as specific needs arise. It has been know for decades that hNETs (human norepinephrine transporters) clear NE+ (norepinephrine) following its release in peripheral sympathetic and central noradrenergic synapses. Neuronal activity influences NE+ uptake, so one is also interested in the acute regulation of hNET. To study these problems, hNET-expressing cells have been developed that are suitable for patch clamp, radioligand uptake, biochemistry, and transiently expressed clones for structure-function analysis, and new protocols have been designed combining patch-clamp, microamperometry, Ca2+ imaging, and native catecholamine transporter preparations to study transporters in whole cells and isolated patches. Using these methods, Na-dependent, NE+-induced hNET currents that are blocked by cocaine and antidepressants, channel modes of NE+ conduction, voltage-dependent uptake coupled to NE+-induced ion channel activity, PKC (phosphokinase C) regulation of NE+ uptake, and transporter modulation by [Ca2+]i have all been discovered. There is also provocative new data on other transporters in this family, such as Li/Na mole fraction experiments in the Drosophila serotonin transporters and sided enkephalin block in proline transporters. These studies have led one to postulate the existence of a narrow pore within transporters through which the substrate (NE+ or serotonin, 5HT+) and other ions (principally Na+) pass. It is hypothesized that the pore resides in an oligomeric structure and that separate gene products of hNET or hSERT (human serotonin transporters) come together to form a channel.