Activation energies (Ea) for water movement across vasopressin-(ADH) sensitive epithelia have been reported to be about 10 kcal/mol (1, 12). The present study shows that measurements of Ea for osmotic water flow across toad bladders are unreliable, because a temperature change induces marked alterations in membrane permeability to water within a 2.5-min interval. Thus bladders equilibrated with ADH either at room temperature or at 33 degrees C and then suddenly subjected to a lower temperature were found to exhibit a marked increase in membrane permeability to water. This observation suggests that there is a rapid turn-over of water permeability sites and that sudden exposure to cold inhibits the removal more than the induction of sites by ADH. To stabilize ADH-induced water channels for Ea measurements, bladders were exposed to ADH at room temperature, fixed with glutaraldehyde, and subjected to osmotic gradients at different temperatures. The Ea values for osmotic water flow across these ADH-permeabilized, glutaraldehyde-fixed bladders were 5.1 (4-12 degrees C), 4.3 (12-21 degrees C), 3.6 (21-36 degrees C), and 3.6 kcal/mol (30-38 degrees C). Ea values for shear viscosity of water in these temperature ranges were calculated to be 4.7, 4.2, 4.1, and 3.6 kcal/mol, respectively. The close correlation between Ea values for bulk water viscosity and osmotic water flow across the bladder wall suggests that an equivalent number of hydrogen bonds must be broken to achieve an increase in water flow through ADH-induced channels and an increase in fluidity of water in bulk solution.