Studies of aggregate-carrying vesicles and actin cytoskeleton during hydrosmotic response to ADH in amphibian urinary bladder

Abstract

Antidiuretic hormone (ADH) increases the water permeability of the apical membrane of the granular epithelial cells in amphibian urinary bladder and mammalian collecting duct by a cAMP-mediated signaling pathway. Cytoskeletal elements are considered to be involved in the action of the hormone. Upon ADH stimulation, cytoplasmic aggregate-carrying vesicles termed aggrephores fuse with the apical membrane and deliver particle aggregates which are related to water channels.;The first part of this thesis study concerns the dynamics of aggrephore fusing, how the aggrephores behave during continuous ADH stimulation, and whether the aggrephores are part of the acid-endosome system. It has been demonstrated that aggrephore fusions occur as early as 2 minutes after ADH exposure. Retrieved aggrephores labeled with horseradish peroxidase (HRP) are observed at 5 minutes of ADH stimulation. Cessation of ADH stimulation, followed by a second stimulation, greatly reduced the number of labeled aggrephores. Based on these data, combined with the finding that aggrephores are not acidic organelles, it is proposed that in response to ADH, aggrephores recycle between cytoplasm and apical membranes without entering into an acidic compartment.;In the second part of the thesis study I have explored the response of the actin cytoskeleton of urinary bladder epithelial cells to ADH. It has been demonstrated that ADH induces rapid F-actin depolymerization via a cAMP-mediated signal pathway, and stabilization of F-actin with NBD-phallacidin selectively inhibits water flow. It is proposed that depolymerization of F-actin may facilitate aggrephore fusion.