Kinetic and pharmacological properties of human brain Na+/H+ exchanger isoform 5 stably expressed in Chinese hamster ovary cells

The recently cloned Na+/H+ exchanger isoform 5 (NHE5) is expressed predomin antly in brain, yet Little is known about its functional properties, To fac ilitate its characterization, a full-length cDNA encoding human NHE5 was st ably transfected into NHE-deficient Chinese hamster ovary AP-1 cells. Pharm acological analyses revealed that H-i(+)-activated Na-22(+) influx mediated by NHE5 was inhibited by several classes of drugs (amiloride compounds, 3- methylsulfonyl-4-piperidinobenzoyl guanidine methanesulfonate, cimetidine, and harmaline) at half-maximal concentrations that were intermediate to tho se determined for the high affinity NHE1 and the low affinity NHE3 isoforms , but closer to the latter. Kinetic analyses showed that the extracellular Na+ dependence of NHE5 activity followed a simple hyperbolic relationship w ith an apparent affinity constant (K-Na) of 18.6 +/- 1.6 mM. By contrast to other NHE isoforms, NHE5 also exhibited a first-order dependence on the in tracellular H+ concentration, achieving half-maximal activation at pH 6.43 +/- 0.08. Extracellular monovalent cations, such as H+ and Li+, but not K+, acted as effective competitive inhibitors of Na-22(+) influx by NHE5. In a ddition, the transport activity of NHE5 was highly dependent on cellular AT P levels. Overall, these functional features distinguish NHE5 from other fa mily members and closely resemble those of an amiloride-resistant NHE isofo rm identified in hippocampal neurons.