Chloride conductance of the Amphiuma red cell membrane

Abstract

Like most other red cells, the giant erythrocytes of Amphiuma means possess a system for rapid exchange of chloride across the membrane. Also, there are indications that the net transport of chloride in these cells is slow. The size of Amphiuma erythrocytes allows direct measurements of membrane potential with microelectrodes. The present work exploits the possibility that such measurements can be used to give a quantitative estimate of the chloride conductance (GCl) of the Amphiuma red cell membrane. The membrane potential was measured as a function of extracellular chloride concentration (5-120mM), using an impermeant anion (Para-amino-hippurate) as a substitute. Furthermore, the effect of different pH values (6.0-7.2) was studied. For each extracellular chloride concentration the membrane potential was determined at a pH at which hydroxyl, hydrogen, and bicarbonate ions were in electrochemical equilibrium. From these membrane potentials and the corresponding chloride concentrations in the medium (at constant intracellular ion concentrations), the GCl of the membrane was calculated to be 3.9 x 10-7 omega-1 cm-2. This value is some six orders of magnitude smaller than that calculated from the rate of tracer exchange under equilibrium conditions. The experimental strategy used gives the values for a "partial transference number" which takes into account only ions which are not in electrochemical equilibrium. Whereas this approach gives a value for GCl, it does not permit calculation of the overall membrane conductance. From the calculated value of GCl it is possible to estimate that the maximal value of the combined conductances of hydroxyl (or proton) and bicarbonate ions is 0.6 x 10-7 omega-1 cm-2. The large discrepancy between the rate of exchange of chloride and its conductance is in agreement with measurements on human and sheep red cells employing the ionophore valinomycin to increase the potassium conductance of the membrane. The results in the present study were, however, obtained without valinomycin and an accompaning assumption of a constant field in the membrane. Therefore, the present measurements give independent support to the above mentioned conclusions.

Membrane voltage, resistance, and channel switching in isolated mouse fibroblasts (L cells): a patch‐electrode analysis.

“…4, 7 and 11), we conclude that GO1 is negligible. This result contrasts directly with the supposition of Lamb & MacKinnon (1971 a, b), that the large isotopic Cl-flux would be current-carrying, and argues that the major C1-transport process in L cells is electrically silent, like that in erythrocytes (Lassen,Pape 286 RESTING POTENTIALS AND K+ PERMEABILITY & Vestergaard-Bogind, 1978) in certain heart-muscle cells (Piwnica-Worms, Jacob, Horres & Lieberman, 1983), and in proximal tubular cells of the kidney (Shindo & Spring, 1981;Guggino, London, Boulpaep & Giebisch, 1983). With resting membrane potentials of -50 to -75 mV and the values of EK and ENa noted above, linear equivalent-circuit theory would yield conductance ratios (GNa/GK) of 0-033-0-24.…”

Volume-regulatory responses of Amphiuma red cells in anisotonic media. The effect of amiloride.

“…15) . Direct microelectrode measurements in Amphiuma red cells have shown that the CI concentration ratio can be used to predict the membrane potential, V., under steady state conditions (Lassen, 1977 ;Stoner and Kregenow, 1980;Cala, 1980). These studies show that there is an^-33-mV change in Vm per 10-fold change in the Cl concentration ratio.…”

“…Cala (1980) found that there were no detectable changes in Vm during volume recovery, which suggests that the volume-regulatory mechanisms are electroneutral . It must be emphasized that if volume-regulatory Na and K movements were through conductive pathways, rCl would not be an accurate indicator of the membrane potential (Lassen, 1977 ;Wieth and Brahm, 1980) . The relative conductances of Cl, Na, and K in Amphiuma red cells are 2:1:1 (Lassen, 1977) .…”

“…It must be emphasized that if volume-regulatory Na and K movements were through conductive pathways, rCl would not be an accurate indicator of the membrane potential (Lassen, 1977 ;Wieth and Brahm, 1980) . The relative conductances of Cl, Na, and K in Amphiuma red cells are 2:1:1 (Lassen, 1977) . Thus, if GNa increased >100-fold, as would be required to account for the Na movements observed in the VRI response, Vm would approach EN., and Cl would not equilibrate rapidly enough through the Cl conductance to reflect V.. On the other hand, if the volume-regulatory mechanisms of Amphiuma red cells are electroneutral, as suggested by the data ofCala (1980), the high transport capacity of the anion exchanger would, in parallel with other Cl pathways, re-equilibrate CI within seconds (see Fig.…”

Patch-Clamp Analysis of Membrane Transport in Erythrocytes

“…This protein was identified in 1972 by Cabantchik and Rothstein (Cabantchik & Rothstein, 1972), even if RBC anion permeability had been studied for long time. It was known for long that RBCs anion permeability could be divided into two components: a large exchange component fundamental for the CO 2 -carrying capacity of the blood (Gunn et al, 1973), and a much smaller electrogenic component that normally determines the RBC resting potential (Hunter, 1977;Lassen et al, 1978). This conductive part of chloride permeability ensures a dissipation of chloride gradient across red cell membrane: the membrane potential is clamped at the equilibrium potential for chloride (-12mV) ensuring that Band 3 never has to fight against a chloride gradient to transport bicarbonate ions across red cell membrane.…”