CORTICOSTEROID-MODULATED EPITHELIAL NA+ & HC03- TRANSPORT

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This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The project goal is to define mechanisms of corticosteroid-regulated ion transport across vas deferens epithelia. Different luminal environments are required for sperm maturation, storage and activation in the male duct. We established protocols to study ion transport in freshly isolated human and porcine ducts, in primary ductile epithelial cell cultures, and in immortal porcine vas deferens epithelial cells. Each of these systems exhibits all or many of the regulatory properties observed in the intact tissue. I will use this set of experimental systems to achieve the following specific aims. Aim 1: To define corticosteroid signaling pathways that regulate Na+ transport across vas deferens epithelium. Vas deferens epithelial cells exhibit amiloride-sensitive ion transport only when exposed to glucocorticoids. In tissues such as kidney, lung, and semicircular canal duct, corticosteroids are linked to changes in epithelial Na+ transport by either mineralocorticoid or glucocorticoid receptors and by various 2nd messengers. Whether these receptors and messengers affect activity of the epithelial Na+ channel in the male reproductive duct is unknown. Pharmacological modulators and molecular techniques will be used to test for discrete changes in these cytosolic regulators and to determine the sequential or parallel order in which messengers are modulated by corticosteroids. Aim 2: To define mechanisms by which glucocorticoids regulate epithelial HCO3- transport. Glucocorticoid receptor agonists and antagonists modulate cAMP-stimulated anion secretion. Corticosteroid receptors could be linked directly to HCO3- transport by a second messenger pathway or indirectly by effects of cation channels on membrane voltage, which is a driving force that supports both basolateral anion entry and apical anion exit. We will use pharmacological modulators and molecular techniques to test for discrete effects on the expression or activity of HCO3- transporters. The relationship of corticosteroids to HCO3- secretion is unknown, but in the context of the vas deferens, likely plays a pivotal role in male fertility. Results from these studies will identify targets for pharmacological interventions to modulate luminal pH with the most direct application to male fertility. We will establish a mechanistic model(s) to account for acute modulation of epithelial Na+ and HCO3- transport that can be extended to other complex cellular systems in the body.