Structural responses to voltage-clamping in the toad urinary bladder

Abstract

The structural consequences of clamping the transepithelial potential difference across the toad's urinary bladder have been examined. Reducing the potential to zero (short-circuiting) produced no apparent changes in the morphology of any of the four cell types which comprise the epithelium. Computer assisted, morphometric analysis of quick frozen specimens revealed no measurable difference in granular cell volume between open- and short-circuited preparations. However, when the open-circuit potential was quantitatively reversed (serosa negative with respect to mucosa), some of the preparations showed a marked increase in granular cell volume. To examine this more systematically twelve preparations were voltage-clamped at 50 mV (serosa negative); eight of the twelve revealed prominent granular cell swelling relative to control, short-circuited preparations. Only in this group of eight had the external circuit current fallen substantially during the clamping interval. Mitochondria-rich cells were not affected detectably. Application of the diuretic amiloride prior to clamping at reversed potential prevented granular cell swelling in every case. Goblet cells which were often affected by the −50 mV clamp were not protected by the diuretic. Granular cell swelling thus appeared to be dependent on sodium entry at the mucosal surface. We also observed that, after voltage reversal, the apical “tight” junctions of the bladders were blistered as they are with hypertonic mucosal media. This blistering was associated with an increase in passive ionic permeability and was not prevented by application of amiloride. This finding is consistent with the evidence that the junction is a complex barrier with asymetric, and hence, rectifying properties for intrinsic ionic conductance as well as hydraulic permeability. These findings, together with others from the literature, lead to the conclusion that the granular cells constitute the principal, if not sole, elements for active sodium transport across toad urinary bladder and that they swell when sodium entry exceeds the transport capacity of the pump at the basal-lateral surface.