Effects of anions on cellular volume and transepithelial Na+ transport across toad urinary bladder

Abstract

The effects of complete substitution of gluconate for mucosal and/or serosal medium Cl- on transepithelial Na+ transport have been studied using toad urinary bladder. With mucosal gluconate, transepithelial potential difference (VT) decreased rapidly, transepithelial resistance (RT) increased, and calculated short-circuit current (Isc) decreased. Calculated ENa was unaffected, indicating that the inhibition of Na+ transport was a consequence of a decreased apical membrane Na+ conductance. This conclusion was supported by the finding that a higher amiloride concentration was required to inhibit the residual transport. With serosal gluconate VT decreased, RT increased and Isc fell to a new steady-state value following an initial and variable transient increase in transport. Epithelial cells were shrunken markedly as judged histologically. Calculated ENa fell substantially (from 130 to 68 mV on average). Ba2+ (3 mM) reduced calculated ENa in Cl- Ringer's but not in gluconate Ringer's. With replacement of serosal Cl- by acetate, transepithelial transport was stimulated, the decrease in cellular volume was prevented and ENa did not fall. Replacement of serosal isosmotic Cl- medium by a hypo-osmotic gluconate medium (one-half normal) also prevented cell shrinkage and did not result in inhibition of Na+ transport. Thus the inhibition of Na+ transport can be correlated with changes in cell volume rather than with the change in Cl-per se. Nystatin virtually abolished the resistance of the apical plasma membrane as judged by measurement of tissue capacitance. With K+ gluconate mucosa, Na+ gluconate serosa, calculated basolateral membrane resistance was much greater, estimated basolateral emf was much lower, and the Na+/K+ basolateral permeability ratio was much higher than with acetate media. It is concluded the decrease in cellular volume associated with substitution of serosal gluconate for Cl results in a loss of highly specific Ba2+-sensitive K+ conductance channels from the basolateral plasma membrane. It is possible that the number of Na+ pump sites in this membrane is also decreased.

Electrical transients produced by the toad urinary bladder in response to altered medium osmolality.

“…Although short-circuited bladders bathed in normal gluconate Ringer solution modified their transepithelial currents when challenged with an osmotic step, we have not located a consistent lower limit of osmolality of the serosal bathing medium beyond which short-circuit current would be unaffected by hypo-osmotic steps. However, as reported by others (Lewis et al 1985), a bladder bathed on the serosal side by a gluconate Ringer solution in which sodium gluconate concentration was reduced by 50 %, produced a current approximately equal to that generated by the same bladder when bathed by a normal chloride Ringer solution. Hypo-osmotic steps in this diluted gluconate medium had little further effect on the SCC, indicating that a limit had been reached or passed.…”

“…If the sodium pump has a fixed flux ratio, sodium to potassium, then both the activation or recruitment of sodium pumps and the hyperpolarization of cells following a hypo-osmotic step would produce an increase in activity of cell potassium unless the basolateral membrane potassium conductance adjusted to match the changed uptake. Although specific evidence for changes in basolateral membrane conductance was not obtained in these studies, others (Schultz, 1981;Lewis et al 1985) have found a positive correlation between pump rate and basolateral membrane conductance.…”

“…Also, in a short-circuited bladder, there is a transient increase in current following the partial isosmotic exchange of gluconate for chloride in the serosal medium (L. G. M. Gordon, personal observation). As the basolateral membrane is impermeable to gluconate (Lewis et al 1985), a chloride efflux should produce an initial reduction in short-circuit current under these conditions, not the observed increase. This evidence suggests that the conductance for chloride in this membrane is significantly lower than that for potassium.…”

Fluid absorption by rat lung in situ: pathways for sodium entry in the luminal membrane of alveolar epithelium.

“…Another proposal regarding gluconate effects has been put forward by Lewis, Butt, Bowler, Leader & Macknight (1985). They found that gluconate reduces the Na+ permeability of the luminal membrane in toad bladder epithelium.…”

Single anion-selective channels in basolateral membrane of a mammalian tight epithelium.

“…Membrane chloride conductance is an essential feature of cellplar models for chloride transport across the cornea (1), trachea (2), and thick ascending limb of Henle's loop (3) and is thought to be an important regulatory site for fluid secretion and cell-volume regulation in some cells (4)(5)(6)(7). However, unlike sodium and potassium channels, which have been characterized using fluctuation analysis (8,9), little is known regarding anion permeability in most epithelia, and its constituent ion channels have not been identified (10).…”