H⁺ Permeation and pH Regulation at a Mammalian Serotonin Transporter

The rat serotonin transporter expressed in Xenopusoocytes displays an inward current in the absence of 5-HT when external pH is lowered to 6.5 or below. The new current differs from the leakage current described previously in two ways. (1) It is ∼10-fold larger at pH 5 than the leakage current at pH 7.5 and reaches 1000 H⁺/sec per transporter at extremes of voltage and pH with no signs of saturation. (2) It is selective for H⁺ by reversal potential measurements. Similar H⁺-induced currents are also observed in several other ion-coupled transporters, including the GABA transporter, the dopamine transporter, and the Na⁺/glucose transporter. The high conductance and high selectivity of the H⁺-induced current suggest that protons may be conducted via a hydrogen-bonded chain (a “proton-wire mechanism”) formed at least partially by side chains within the transporter. In addition, pH affects other conducting states of rat serotonin transporter. Acidic pH potentiates the 5-HT-induced, transport-associated current and inhibits the hyperpolarization-activated transient current. The dose–response relationships for these two effects suggest that two H⁺ binding sites, with pK_a values close to 5.1 and close to 6.3, govern the potentiation of the 5-HT-induced current and the inhibition of the transient current, respectively. These results are important for developing structure-function models that explain permeation properties of neurotransmitter transporters