Membrane potential, chloride exchange, and chloride conductance in Ehrlich mouse ascites tumour cells.

Abstract

SUMMARY1. The steady-state tracer exchange flux of chloride was measured at 10-150 mM external chloride concentration, substituting either lactate or sucrose for chloride. The chloride flux saturates in both cases with a K1 about 50 and 15 mm, respectively.2. The inhibitory effect of other monovalent anions on the chloride transport was investigated by measuring the 3601-efflux into media where either bromide, nitrate, or thiocyanate had been substituted for part of the chloride. The sequence of increasing affinity for the chloride transport system was found to be: Br-< Cl1 < SCN-= NO3-.3. The chloride steady-state exchange flux in the presence of nitrate can be described by Michaelis-Menten kinetics with nitrate as a competitive inhibitor of the chloride flux.4. The apparent activation energy (EA) was determined to be 67 + 6-2 kJ/mole, and was constant between 7 and 38 0C.5. The membrane potential (Vm) was measured as a function of the concentration of external K+, substituting K+ for Na+. The transference number for K+ (tK) was estimated from the slope of Vm vs. log10 (K+)e, and tcl and tNa were calculated, neglecting current carried by ions other than Cl-, K+, and Na+. The diffusional net flux of K+ was calculated from the steady-state exchange flux of 42K+, assuming the flux ratio equation to be valid. From this value the K+ conductance and the Na+ and Cl-conductances were calculated. The experiments showed that %t, GN., and GK are all about 14 ,US/cm2. 6. The net (conductive) chloride permeability derived from the chloride conductance was 4 x 10-8 cm/sec compared with the apparent permeability of 6 x 10-7 cm/ sec as calculated from the chloride tracer exchange flux. These data suggest that about 95 % of the chloride transport is mediated by an electrically silent exchange diffusion.7. Comparable effects of phloretin (0.25 mM) on the net (conductive) permeability and the apparent permeability to chloride (about 80 % inhibition) may indicate that the chloride exchange and conductance pathways are not completely separate and Experiments were performed at both addresses.

Role of volume-regulated and calcium-activated anion channels in cell volume homeostasis, cancer and drug resistance

“…14 Figure 1A illustrates Cl -efflux under isotonic, hypotonic and hypertonic conditions, and it has been determined that under isotonic conditions less than 5% represents conductive Cl -flux with the rest being electroneutral anion exchange (AE). 15 In contrast, increased Clflux under swollen and shrunken conditions represent Cl -conductance (VRAC) and electroneutral Cl -transport via the Na, K,2Cl cotransporter (NKCC), respectively. 10 Figure 1B shows the volume regulatory response in EATC under hypotonic conditions, where the intrinsic volume sensitive K C channel has been blocked by quinine but a high K C conductance has been ensured by addition of the cation-ionophore gramicidin, hence making Cl -movement rate limiting for volume changes.…”

“…Under hypertonic conditions an increasing part of the Cl -efflux constitutes cation dependent electroneutral Na, K, 2Cl cotransporter (NKCC). Values from 15,158 and Figure adapted from 7 . (B) Transient increase in the Cl -permeability following hypoosmotic cell swelling.…”

Activation of the TASK-2 channel after cell swelling is dependent on tyrosine phosphorylation

“…K ϩ content was determined by atomic absorption flame photometry (model 2380, Perkin Elmer atomic absorptions spectrophotometer) using samples and K ϩ standards diluted 100-fold with 1 mM CsCl. 86 Rb ϩ activity (cpm/g cell dry wt), estimated from the extracellular 86 Rb activity (cpm/ml medium) and the cell dry weight (g/ml medium), was plotted versus time, and the rate constant for 86 Rb ϩ efflux (min Ϫ1 ) was determined by fitting the efflux curve between 1 and 10 min to an exponential function (18). The 86 Rb ϩ efflux (cpm/ml cell water ϫ min Ϫ1 ) was calculated as the product of the rate constant and the cellular concentration (cpm/ml cell water).…”

Amino acid transport and cell volume regulation in Ehrlich ascites tumour cells.

“…Measurements of cellular and extracellular concentrations of amino acids and ions Na+, K+ and Cl-were determined as described by Hoffmann, Simonsen & Sj0holm (1979). For determination of the cellular amino acid pool, 1-5 ml.…”