The effect of various intracellular Na concentrations (CiNa) and membrane potentials on the Na pump current (Ip) was studied in isolated, cultured sheep cardiac Purkinje cells ('cardioballs'). Ip was identified as cardiac steroid sensitive current. The dependence of Ip on CiNa was investigated at a membrane potential of -40 mV by means of whole-cell recording from cardioballs internally perfused with media containing various Na concentrations. Internal perfusion with a Na free solution abolished Ip. The amplitude of Ip as a function of CiNa displayed saturation kinetics. Half maximal activation of Ip occurred at a CiNa of about 9 mM. The maximal Ip density was estimated to be 1.1 microA/cm2. The potential dependence of Ip was studied by conventional whole-cell recording under various ionic conditions. Generally Ip displayed little voltage dependence at membrane potentials positive to -20 mV. Ip declined at more negative potentials. The pump cycle probably includes only one voltage sensitive step. The potential dependence of Ip was more pronounced at lower CiNa or lower concentrations of the external pump activator Cs+. The findings are in line with the idea that increasingly steeper ionic gradients against which the cations are pumped strengthen the voltage dependence of Ip in the potential range studied. Other factors probably affecting the pump current-voltage (Ip-V) relation are discussed. The results suggest that Ip varies during electrical activity.

A mathematical model of ascending Henle limb (AHL) epithelium has been fashioned using kinetic representations of Na+-K+-2Cl- cotransporter (NKCC2), KCC4, and type 3 Na+/H+ exchanger (NHE3), with transporter densities selected to yield the reabsorptive Na+ flux expected for rat tubules in vivo. Of necessity, this model predicts fluxes that are higher than those measured in vitro. The kinetics of the NKCC and KCC are such that Na+ reabsorption by the model tubule is responsive to variation in luminal NaCl concentration over the range of 30 to 130 mM, with only minor changes in cell volume. Peritubular KCC accounts for about half the reabsorptive Cl- flux, with the remainder via peritubular Cl- channels. Transcellular Na+ flux is turned off by increasing peritubular KCl, which produces increased cytosolic Cl- and thus inhibits NKCC2 transport. In the presence of physiological concentrations of ammonia, there is a large acid challenge to the cell, due primarily to NH4+ entry via NKCC2, with diffusive NH3 exit to both lumen and peritubular solutions. When NHE3 density is adjusted to compensate this acid challenge, the model predicts luminal membrane proton secretion that is greater than the HCO3(-)-reabsorptive fluxes measured in vitro. The model also predicts luminal membrane ammonia cycling, with uptake via NKCC2 or K+ channel, and secretion either as NH4+ by NHE3 or as diffusive NH3 flux in parallel with a secreted proton. If such luminal ammonia cycling occurs in vivo, it could act in concert with luminal K+ cycling to facilitate AHL Na+ reabsorption via NKCC2. With physiological ammonia, peritubular KCl also blunts NHE3 activity by inhibiting NH4+ uptake on the Na-K-ATPase, and alkalinizing the cell.

A mathematical model of the brush border of the proximal tubule (T. A. Krahn, P. S. Aronson, and A. M. Weinstein. Bull. Math. Biol, 56: 459-490, 1994) has been extended by the inclusion of CO2 and H2CO3 as diffusible species and by the inclusion of finite rate constants for the hydration of CO2. This permits the simulation of carbonic anhydrase (CA) activity and its inhibition. We confirm the result of our previous study, which is that, in the presence of CA, the unstirred layer has only a modest effect on the observed formic acid permeability. CA inhibition results in disequilibrium pH gradients, and the effect of these gradients on formic acid permeability depends on the presence of other membrane transport proteins. We also examined the impact of CA activity on the flux of total CO2 through the brush border. Under physiological conditions, CA inhibition depressed NaHCO3 reabsorption through the brush border by interfering with the HCO3(-)-facilitated diffusion of CO2. However, the determination of brush-border CO2 permeability, using an imposed CO2 gradient, was relatively uninfluenced by CA activity. Finally, we inserted a kinetic representation of the Na+/H+ exchanger into the brush-border model. Even when luminal and cytosolic diffusion coefficients were increased 1,000-fold, there was no effect on brush-border Na+ flux. This suggests that variations in the unstirred layer cannot be responsible for the flow dependence of Na+ reabsorption.

Locally produced PD solutions, used with locally available tubing, were used to treat AKI with PD. The mortality and peritonitis rates were similar whether locally produced or commercial supplies were used.