Effect of temperature on apical membrane remodeling in ADH-stimulated toad urinary bladders

Pretreatment and removal of vasopressin (ADH) in toad urinary bladder renal model tissues induces endocytosis at 25°C. The objective of the current study is to determine if apical membrane remodeling, as well as transepithelial water flow, can be affected by lowering the temperature to 15°C. Control toad urinary bladders in the presence of an osmotic gradient at either 25°C or 15°C when visualized by scanning electron microscopy (SEM) show a typical apical membrane surface with no apparent surface differences. ADH-treated tissues following 15-min stimulation at 25°C or 15°C revealed a propagation of apical microvilli on their surface membranes. After 15 min following removal of ADH, bladder tissues at 25°C or 15°C showed surface invaginations involving over 44% and 80% of granular cells, respectively. The rate of water flow in tissues at 15°C remained elevated compared to tissues held at 25°C. This was consistent with the observation that ADH-stimulated tissues following washout at 15°C still had marked apical membrane surface involvement. However, at 30 min and 60 min postwashout, ADH-stimulated tissues at 15°C recovered considerably, with a reduction in the number of shallow apical membrane invaginations involving fewer than 33% and 20% of granular cells respectively. This may indicate that the membrane undergoes continuous remodeling even at cold temperature conditions but with a different half-time. Control bladder tissues subjected to transmission electron microscopy (TEM) reveal a dense cytoplasmic profile with a scattered distribution of secretory granules, rough ER cisternae, mitochondria, and little or no vacuolation. In contrast, ADH-stimulated bladder tissues displayed a vacuolated cytoplasm, expanded rough ER cisternae, and ruffled basolateral membranes. These observations suggest that the apical membrane undergoes considerable reorganization following cessation of hormone action and that lowering the temperature reduces the rate of membrane remodeling and thus may provide a means to monitor the processes of endocytosis and the mechanisms responsible for water channel retrieval.