Sodium is not required for chloride efflux via chloride/bicarbonate exchanger from rat thymic lymphocytes

Abstract

Sodium-dependent Cl-/ H C O 3 - exchanger acts as a chloride (Cl-) efflux in lymphocytes. Its functional characterization had been described when Cl- efflux was measured upon substituting extracellular sodium (Na+) by N-methyl-D-glucamine (NMDG). For Na+ and Cl- substitution, we have used D-mannitol or NMDG. Thymocytes of male Wistar rats aged 7-9 weeks were used and intracellular Cl- was measured by spectrofluorimetry using MQAE dye in bicarbonate buffers. Chloride efflux was measured in a Cl--free buffer (Cl - substituted with isethionate acid) and in Na+ and Cl--free buffer with D-mannitol or with NMDG. The data have shown that Cl- efflux is mediated in the absence of Na+ in a solution containing D-mannitol and is inhibited by H2DIDS. Mathematical modelling has shown that Cl- efflux mathematical model parameters (relative membrane permeability, relative rate of exchanger transition, and exchanger efficacy) were the same in control and in the medium in which Na+ had been substituted by D-mannitol. The net Cl - efflux was completely blocked in the NMDG buffer. The same blockage of Cl- efflux was caused by H2DIDS. The study results allow concluding that Na+ is not required for Cl- efflux via Cl-/ H C O 3 - exchanger. NMDG in buffers cannot be used for substituting Na+ because NMDG inhibits the exchanger. © 2014 Donatas Stakišaitis et al.