UCLA

The chloride gradient plays an important role in regulating cell volume, membrane potential, pH, secretion, and the reversal potential of inhibitory glycine and GABA_A receptors. Measurement of intracellular chloride activity, [Formula: see text], using liquid membrane ion-selective microelectrodes (ISM), however, has been limited by the physiochemical properties of Cl^- ionophores which have caused poor stability, drift, sluggish response times, and interference from other biologically relevant anions. Most importantly, intracellular [Formula: see text] may be up to 4 times more abundant than Cl^- (e.g. skeletal muscle) which places severe constraints on the required selectivity of a Cl^- - sensing ISM. Previously, a sensitive and highly-selective Cl^- sensor was developed in a polymeric membrane electrode using a trinuclear Hg(II) complex containing carborane-based ligands, [9]-mercuracarborand-3, or MC3 for short. Here, we have adapted the use of the MC3 anion carrier in a liquid membrane ion-selective microelectrode and show the MC3-ISM has a linear Nernstian response over a wide range of a_Cl (0.1 mM to 100 mM), is highly selective for Cl^- over other biological anions or inhibitors of Cl^- transport, and has a 10% to 90% settling  time of 3  sec. Importantly, over the physiological range of a_Cl (1 mM to 100 mM) the potentiometric response of the MC3-ISM is insensitive to [Formula: see text] or changes in pH. Finally, we demonstrate the biological application of an MC3-ISM by measuring intracellular a_Cl, and the response to an external Cl-free challenge, for an isolated skeletal muscle fiber.