SUMMARY1 Amiloride inhibits Na transport and short-circuit current (SCC) across the toad bladder. It is 1000 times more active at the mucosal than serosal surface. The lowest effective concentration was 10-7 M.2. The inhibition was non-competitive with the sodium on the mucosal side of the bladder.3. Vasopressin, cyclic adenosine monophosphate (AMP) and aldosterone increased Na transport and SCC across the bladder and these effects were inhibited by amiloride.4. The antagonism of amiloride, for vasopressin was non-competitive.5. Amphotericin B also increases Na transport across the bladder but its action was not changed by amiloride.6. Amiloride was without effects on SCC and diffusion potentials in bladders metabolically inhibited with CN-and iodoacetic acid (IAA).7. Neither plasma albumin, Ca2+ nor adenosine triphosphate (ATP) altered the effects of amiloride.8. The only structural analogue of amiloride found to reduce SCC similarly was guanidine which was 1000 times less active. Pyrazine and a substituted pyrazine analogue were without effect. Neither guanidine nor the substituted pyrazine compound were competitive with amiloride.9. Amiloride had no effect on the osmotic permeability of the toad bladder either in the presence or absence of vasopressin.10. Na transport across the toad colon was also reduced by 10-5 M amiloride at the mucosal surface.11. The possible mechanism of action of amiloride is discussed.

1. An in vitro preparation of the urinary bladder of Bufo marinus is described. 2. Small doses of 'Pituitrin' markedly increase the rate of water transfer across the bladder wall when the solutions inside the bladder are hypotonic.3. Passive movement is small and increases slightly with increases in the osmotic gradient across the bladder wall. It is unaffected by changes in substrate levels or any of the metabolic inhibitors tested except for cyanide which increases it in some cases.4. The vasopressor neurohypophysial fraction is more active than the oxytocic one in increasing water transfer across the bladder wall.5. The increase in water transfer depends on an intact oxygen supply and sufficient glucose or pyruvate.6. Iodoacetate, malonate, cyanide, 2-4-dinitrophenol, and bubbling 5% CO2+95% O2 through Ringer's solution inhibit the water transfer in response to neurohypophysial extract. 7. Diamox is only an effective inhibitor at very high concentrations. 8. The possible mechanism of the water transfer is discussed.The reabsorption of water stored in the urinary bladder of Bufo regularis when injected with neurohypophysial extract has been shown by Ewer [1952]. The possi¬ bility that the water may have been absorbed after régurgitation into the cloaca was eliminated by Sawyer & Schisgall [1956], who concluded that the water was in fact transferred across the bladder wall.Preparations of frog and toad skin in vitro have been used for the study of the movements of water and salt across biological membranes [see Sawyer, 1956]. From Ewer's observations it seemed likely that the isolated urinary bladder of the toad could be used for similar studies. Moreover, the simpler histological structure, in particular the absence of the numerous secretory glands present in the frog and toad skin, might clarify interpretation of results. The experiments reported in this paper demonstrate that the isolated bladder of the toad is a very suitable preparation for the study of neurohypophysial effects on water transport in vitro. Results are also presented from a continuing investigation of factors which influence the effect of neurohypophysial extracts. 13Endoc. 17, 3

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