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Coincident with an increase in the water permeability of toad urinary bladder induced by serosal hypertonicity, a transformation of the ridge-like surface structures of the granular cells into individual microvillous structures occurs. This study was initiated to establish whether the transformation is mediated by the cytoskeletal network and, thus, can be prevented by disruption of microtubule-microfilament function with colchicine or cytochalasin B (CB). Scanning electron microscopy revealed the characteristic branching ridges on granular cells of control bladder incubated with colchicine or CB. In contrast, transformation of ridges to discrete microvilli was observed in experimental bladders exposed to serosal hypertonicity alone or in combination with either colchicine or CB. These results suggest that the mechanism underlying hypertonicity-induced surface changes which are associated with increased water permeability does not involve either microtubules or microfilaments.

Coincident with an increase in water permeability, the ridge-like surface structures of toad bladder granular cells transform to individual microvilli after stimulation with vasopressin (VP) or cyclic adenosine monophosphate (cAMP) by a mechanism that is yet to be defined. To explore the possible role of microtubules and microfilaments in this cell response, colchicine and cytochalasin B were employed to determine whether interference with the function of these components of the cytoskeletal system would prevent the VP- and cAMP-induced conversion of ridges to microvilli. Incubation of toad urinary bladders in 10(-4) M colchicine for 4 hours or 10(-5) M cytochalasin B for 90 minutes before stimulation with 20 mU. per ml. of VP markedly inhibited osmotic water flow. However, neither agent prevented the striking conversion of ridges to surface microvilli induced by VP and cAMP as seen with scanning electron microscopy. In addition, the ridges characteristic of granular cells were maintained in control bladders incubated with colchicine or cytochalasin B, but left unstimulated. Under the conditions of these experiments, these findings suggest that microtubules and microfilaments are not essential for maintenance of normal surface configuration in granular cells of toad urinary bladder, and that they are not involved in the mechanism responsible for VP- and cAMP-induced surface changes that occur in association with increased water permeability of this epithelium.

The mucosal surface of toad urinary bladder was examined with scanning electron microscopy following its exposure to 20 mU/ml of vasopressin (VP), 10(-4) M 8-bromo-cAMP, 1 mM acetylcholine chloride, serosal hypertonicity, or a hypotonic bathing medium. After a 30-min exposure to VP, the arborizing ridge-like surface pattern typical of granular cells was transformed into microvilli, a response that was not dependent on transepithelial osmotic water movement. An identical response occurred following a 30-min exposure of the bladder to 8-bromo-cAMP, again in the absence of an osmotic gradient. Microvillus formation was not observed when cell volume was increased by incubation of tissue in half-normal amphibian Ringer's solution for 30 min, or with exposure to acetylcholine, which caused accentuation of the convexity of the apical surface of the granular cell similar to that observed with VP-induced osmotic water flow. However, 60 min of incubation in a hypertonic serosal medium (mannitol, 240 mM) caused transformation of ridges to microvilli mimicking the picture obtained with VP. These findings establish that transepithelial osmotic water flow with cell swelling is not required for microvillus formation on the apical surface of granular cells following VP stimulation, and that the surface changes are not due to cell swelling alone or to changes in the configuration of the apical plasmalemma. The results also suggest that the response to VP is mediated via the generation of cAMP. Finally, this study demonstrates that serosal hypertonicity also causes transformation of ridges to microvilli by a mechanism that is yet to be defined.