The K–Cl Cotransporter in the Lobster Stretch Receptor Neurone—A Kinetic Analysis

Abstract

Experiments were performed to define quantitatively the substrate (K + and Cl −) dependence of the transport function (production of equally large and oppositely directed K +and Cl − flows/currents) of an earlier (Theander et al., 1999) identified electroneutral K–Cl cotransporter in the slowly adapting stretch receptor neurone of the European lobster. The experiments were based on microelectrode techniques. This allowed us to perform steady-state measurements of the so-called “instantaneous” current–voltage relationships (around a holding voltage of −65 mV after a blockage of the cell's action potential and hyperpolarization-activated currents) and intracellular ion concentrations at various settings of the extracellular K + and Cl − concentrations. From the results, we could then define steady-state values of all of the cell's non-KCl cotransporter K + and Cl − currents. Finally, the negative sums of the inferred non-KCl cotransporter K + and Cl − currents could be taken as equivalents of the K–Cl cotransporter's K + and Cl − currents for the reason that, in steady state, all membrane currents add up to zero. For the cotransporter currents, thus inferred for a range from 2.5/410.5 to 40.0/448.0 mM external K +/Cl −, we found that their absolute values increased in a nonlinear fashion from about 5 nA cell −1 at the lowest, to about 20 nA cell −1 at the highest external K +/Cl − concentrations. Formally, this relationship could be reproduced by a Hill function-based enzyme kinetic expression simulating inward and outward transmembrane electroneutral ion transports. Following insertion of this expression into a comprehensive model of electrical membrane functions and intracellular solute and solvent control in the lobster stretch receptor neurone, the model predictions suggested that the K–Cl cotransporter does play an important role in (a) keeping intracellular Cl − low for a proper function of the cell's inhibitory system, and (b) enabling rapid transmembrane K + shifts that provide for a stabilization of the cell's membrane voltage and membrane excitability in cases of varying extracellular K + concentrations. The model predictions gave, however, no clear evidence that the K–Cl cotransporter is critically involved in the cell's volume regulation in conditions of varying extracellular osmolalities.