H+ permeation and pH regulation at a mammalian serotonin transporter.
The rat serotonin transporter expressed in Xenopus oocytes 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 approximately 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 pKa 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.
Cao
Y
Y
Division of Biology, California Institute of Technology, Pasadena, California 91125, USA.
Mager
S
S
Lester
H A
HA
D000818:Animals; D002352:Carrier Proteins / Q000187:drug effects / Q000235:genetics / Q000502:physiology*; D003891:Desipramine / Q000494:pharmacology; D005260:Female; D005473:Fluoxetine / Q000494:pharmacology; D005998:Glycine / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D007700:Kinetics; D008536:Meglumine / Q000494:pharmacology; D008562:Membrane Glycoproteins / Q000187:drug effects / Q000235:genetics / Q000502:physiology*; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D026901:Membrane Transport Proteins; D009419:Nerve Tissue Proteins; D009865:Oocytes / Q000502:physiology; D018408:Patch-Clamp Techniques; D011522:Protons; D018075:RNA, Complementary; D051381:Rats; D011994:Recombinant Proteins / Q000187:drug effects / Q000378:metabolism; D012701:Serotonin / Q000502:physiology*; D050486:Serotonin Plasma Membrane Transport Proteins; D012964:Sodium / Q000494:pharmacology; D013997:Time Factors; D014158:Transcription, Genetic; D014982:Xenopus laevis
D002352:Carrier Proteins; D008562:Membrane Glycoproteins; D026901:Membrane Transport Proteins; D009419:Nerve Tissue Proteins; D011522:Protons; D018075:RNA, Complementary; D011994:Recombinant Proteins; D050486:Serotonin Plasma Membrane Transport Proteins; C493232:Slc6a4 protein, rat; D005473:Fluoxetine; D012701:Serotonin; D008536:Meglumine; D012964:Sodium; D005998:Glycine; D003891:Desipramine