Vasopressin V2 Receptor, Tolvaptan, and ERK1/2 Phosphorylation in the Renal Collecting Duct

Activation of ERK1 and ERK2 is essential in regulation of a wide variety of cellular and physiological processes. In native inner medullary collecting ducts, vasopressin (AVP) working through the V2 subtype vasopressin receptor (V2R)-mediated activation of Gas, inhibits ERK1 and ERK2 activity. However, it has been reported that V2R can signal independently of Gas through the activation of ß-arrestin, which activates ERK1 and ERK2. Vaptans, V2R antagonists that function as so-called “inverse agonists”, have the potential of promoting cell proliferation via ß-arrestin-dependent ERK activation. Here we use the mpkCCD cell line which natively expresses V2R to investigate the effects of AVP, the V2-selective analog dDAVP, and tolvaptan on ERK1 and ERK2 phosphorylation and activation. We demonstrated that ERK1 and ERK2 phosphorylation in mpkCCD cells was significantly reduced by either AVP or dDAVP, in contrast to the increases seen in non-collecting duct cells overexpressing V2R. We also found that tolvaptan has a strong effect to increase ERK1 and ERK2 phosphorylation in the presence of dDAVP and that the tolvaptan effect to increase ERK1 and ERK2 phosphorylation is absent in PKA-null mpkCCD cells. Thus, it appears that the tolvaptan effect to increase ERK activation is PKA-dependent and, therefore, not mediated by the ß-arrestin pathway. Overall, the studies show that AVP decreases and that tolvaptan increases ERK1 and ERK2 activation in cells expressing V2R at endogenous levels, and provide no evidence for a role for ß-arrestin in the regulation of ERK1 and ERK2 activity. Overall design: Here we use the mpkCCD cell line to address the following questions regarding the endogenously expressed V2 vasopressin receptor: 1) What are the effects of AVP and dDAVP on ERK1 and ERK2 phosphorylation/activation? 2) What is the effect of tolvaptan on ERK1 and ERK2 phosphorylation/activation in the presence and absence of dDAVP? 3) What is the role of PKA in vasopressin-mediated regulation of ERK1 and ERK2 phosphorylation/activation and on the effect of tolvaptan?