Non‐passive chloride distribution in mammalian heart muscle: micro‐electrode measurement of the intracellular chloride activity.

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Abstract
Liquid ion-exchanger Cl--sensitive micro-electrodes were used to make continuous measurements of the intracellular Cl activity, .**GRAPHIC**. of quiscent sheep cardiac Purkinje fibers in vitro. .**GRAPHIC**. was higher than that expected from a passive distribution, which would have been .apprx. 5 mM. It was 3-4 times higher in the presence of an extracellular bicarbonate/CO2 buffer-system and was very stable; ECl [chloride equilibrium potential] was .apprx. 35 mV positive to Em [resting membrane potential]. It was over twice as high in the nominal absence of bicarbonate/CO2 (when the buffer-system was HEPES[4-(2-hydroxyethyl)-1-piperazine-N''-2-ethanesulfonic acid]/O2) but was not always so stable; EC1 was about .apprx. 20 mV positive to Em. Experiments designed to assess the maximum possible error likely to occur in the measurement of .**GRAPHIC**. showed that this could not be large and that the estimates of ECl were accurate to within 8 mV. The ability of Cl to move down concentration and potential gradients was established by demonstrating a loss of .**GRAPHIC**. in Cl-free solutions and a gain when Em was depolarized positive to ECl in high-K solutions. In both cases the changes were complete within 100-160 min. The decline of .**GRAPHIC**. in Cl-free solutions (glucuronate-substituted) was not significantly affected by changes of [Ca]o [extracellular concentration] from 0 to 12 mM or by the depolarizations of Em of up to 60 mV that sometimes occurred in low or zero [Ca]o. Only 2-3 mM .**GRAPHIC**. [extracellular Cl activity] mM was sufficient to impede substantially the ready loss of .**GRAPHIC**. in HEPES-buffered solutions. In high-K solutions (45 mM), Cl appeared to be passively distributed since, at equilibrium, Em and ECl differed by less than 2 mV. In HEPES-buffered Tyrode ECl of quiescent papillary muscle of the guinea pig was an average 39 mV positive to Em. Liquid ion-exchanger Cl--sensitive micro-electrodes are suitable for studying the Cl regulation of sheep Purkinje fibers and probably of other cardiac tissues. The measurements of resting .**GRAPHIC**. are accurate when using HEPES or bicarbonate-buffered Tyrode. The results are discussed in relation to estimates of the apparent membrane Cl permeability under various conditions and the possible existence of an inwardly directed Cl pump.