Measurement of intracellular chloride in guinea-pig vas deferens by ion analysis, 36chloride efflux and micro-electrodes

Abstract

1. Cl-sensitive micro-electrodes were used to measure the intracellular Cl activity (a(Cl) (i)) in smooth muscle cells of the guinea-pig vas deferens. The values obtained were compared with those of intracellular Cl (Cl(i)) found by both ion analysis and (36)Cl efflux.2. Various combinations of filling solution for recording membrane potential (E(m)), and type of micro-electrode were tested. The most successful, which allowed continuous recording of a(Cl) (i) for several hours, was a double-barrelled electrode using the reference liquid ion exchanger (RLIE; Thomas & Cohen, 1981). However, a(Cl) (i) measured both by simultaneous impalements of separate cells with Cl-sensitive and conventional micro-electrodes, and by double-barrelled micro-electrodes, was about 42 mM in normal Krebs solution. This is five times higher than the value from a passive distribution. E(Cl) was about -24 mV, more than 40 mV positive to E(m).3. On complete removal of extracellular Cl (Cl(o)), a(Cl) (i) fell to an apparent level of about 3 mM. If this represents interference from other anions, the maximum error in E(Cl) measured in normal Krebs solution is 2.5 mV. Replacement of Cl(o) caused a rapid increase in a(Cl) (i). This must be caused by an active transport of Cl(-) ions into the cell against their electrochemical gradient.4. The stabilized values of a(Cl) (i) measured at different levels of Cl(o) agree surprisingly well with a(Cl) (i) estimated from ion analysis and (36)Cl efflux, assuming that the intracellular activity coefficient was the same as measured in the normal Krebs solution. The relationship of a(Cl) (i) to Cl(o) was hyperbolic.5. It is concluded that Cl-sensitive micro-electrodes accurately measure a(Cl) (i) in smooth muscle cells. The remarkable agreement between the direct and indirect methods of measuring Cl(i) suggests that Cl(-) ions are not bound to a significant extent and that the compartment seen by the micro-electrodes is probably representative of the whole cell.