Role of cAMP-phosphodiesterase isozymes in pathogenesis of murine nephrogenic diabetes insipidus

Urea transport, mediated by the urea transporter A1 (UT-A1) and/or UT-A3, is important for the production of concentrated urine. Vasopressin rapidly increases urea transport in rat terminal inner medullary collecting ducts (IMCD). A previous study showed that one mechanism for rapid regulation of urea transport is a vasopressin-induced increase in UT-A1 phosphorylation. This study tests whether vasopressin or directly activating adenylyl cyclase with forskolin also increases UT-A1 accumulation in the plasma membrane of rat IMCD. Inner medullas were harvested from rats 45 min after injection with vasopressin or vehicle. UT-A1 abundance in the plasma membrane was significantly increased in the membrane fraction after differential centrifugation and in the biotinylated protein population. Vasopressin and forskolin each increased the amount of biotinylated UT-A1 in rat IMCD suspensions that were treated ex vivo. The observed changes in the plasma membrane are specific, as the amount of biotinylated UT-A1 but not the calcium-sensing receptor was increased by forskolin. Next, whether forskolin or the V 2 -selective agonist dDAVP would increase apical membrane expression of UT-A1 in MDCK cells that were stably transfected with UT-A1 (UT-A1-MDCK cells) was tested. Forskolin and dDAVP significantly increased UT-A1 abundance in the apical membrane in UT-A1-MDCK cells. It is concluded that vasopressin and forskolin increase UT-A1 accumulation in the plasma membrane in rat IMCD and in the apical plasma membrane of UT-A1-MDCK cells. These findings suggest that vasopressin regulates urea transport by increasing UT-A1 accumulation in the plasma membrane and/or UT-A1 phosphorylation.

Section: Discussionmentioning

confidence: 88%

Vasopressin Increases Plasma Membrane Accumulation of Urea Transporter UT-A1 in Rat Inner Medullary Collecting Ducts

Klein¹,

Fröhlich²,

Blount³

et al. 2006

“…Mice with genetically increased activity of PDE3 and PDE4 show a fourfold higher PDE activity in microdissected IMCD and cortical CD and lack an increase in cAMP in response to V 2 R activation, which is associated with lower AQP2 mRNA and protein expression and lower urine osmolality. 23,24 Pharmacologic studies have argued against a role of PDE3 in urine osmolality. 25 However, treatment of mutant AQP2 heterozygous knock-in mice or mice with hereditary NDI with the PDE4 inhibitor, rolipram, increases urine osmolality and restores AVP-dependent cAMP accumulation in IMCD, indicating a role for PDE4 in the regulation of cAMP and urinary concentration.…”

Section: Discussionmentioning

confidence: 99%

Adenylate Cyclase 6 Determines cAMP Formation and Aquaporin-2 Phosphorylation and Trafficking in Inner Medulla

Rieg

Tang²,

Murray³

et al. 2010

Arginine vasopressin (AVP) enhances water reabsorption in the renal collecting duct by vasopressin V 2 receptor (V 2 R)-mediated activation of adenylyl cyclase (AC), cAMP-promoted phosphorylation of aquaporin-2 (AQP2), and increased abundance of AQP2 on the apical membrane. Multiple isoforms of adenylate cyclase exist, and the roles of individual AC isoforms in water homeostasis are not well understood. Here, we found that levels of AC6 mRNA, the most highly expressed AC isoform in the inner medulla, inversely correlate with fluid intake. Moreover, mice lacking AC6 had lower levels of inner medullary cAMP, reduced abundance of phosphorylated AQP2 (at both serine-256 and serine-269), and lower urine osmolality than wild-type mice. Water deprivation or administration of the V 2 R agonist dDAVP did not increase urine osmolality of AC6-deficient mice to the levels of wild-type mice. Furthermore, AC6-deficient mice lacked dDAVP-promoted inner medullary cAMP formation and phosphorylation of serine-269 and had attenuated increases in both phosphorylation of serine-256 and apical membrane AQP2 trafficking. In summary, AC6 expression determines inner medullary cAMP formation and AQP2 phosphorylation and trafficking, the absence of which causes nephrogenic diabetes insipidus. The anti-diuretic hormone arginine-vasopressin (AVP) is the primary regulator of water reabsorption in the renal collecting duct (CD) and is critically involved in the regulation of water balance and maintenance of plasma osmolality. 1 AVP acts on the CD through the G s protein-coupled vasopressin V 2 receptor (V 2 R) to stimulate adenylyl cyclase (AC) and thus the synthesis of cAMP. 2 cAMP activates protein kinase A (PKA), which phosphorylates the water channel aquaporin-2 (AQP2) in its COOH-terminal tail on serine residue 256 (S256), thereby resulting in apical plasma membrane accumulation of AQP2. [3][4][5][6] In addition, cAMP-independent activation of AQP2 has been reported, which may involve V 2 R action of phosphoinositide-specific phospholipase C. 7 Other non-PKA-targeted AQP2 phosphorylation sites include S261, S264, and S269. 8 Phosphorylation of AQP2 at S264 and S269 requires prior phosphorylation of S256. 9 Immunohistochemistry showed that pS269-AQP2 localizes exclusively in the apical plasma

“…13,14 Similarly, although AQP2 protein is preserved in the vasopressin-deficient Brattleboro rat (with spontaneous mutation of the vasopressin gene leading to "central" DI) or mice with overactive cAMP phosphodiesterase in principal cells, both have polyuria from birth without tubular/renal malformation or kidney failure. 15,16 Interestingly, several other mouse models in which polyuria is associated with perturbations in renal structure are reported to have greatly reduced AQP2 expression, whereas polyuric models with no structural abnormalities have normal AQP2 levels. 14,[17][18][19][20][21][22] Thus, we hypothesized that the tubular abnormalities seen in the neonatal AQP2 knockout animals and/or AQP2 transgenics are due to an intrinsic defect associated with the absence or dysfunction of AQP2 rather than being a secondary effect of polyuria.…”

mentioning

confidence: 99%

Aquaporin 2 Promotes Cell Migration and Epithelial Morphogenesis

Chen

Rice

et al. 2012

The aquaporin 2 (AQP2) water channel, expressed in kidney collecting ducts, contributes critically to water homeostasis in mammals. Animals lacking or having significantly reduced levels of AQP2, however, have not only urinary concentrating abnormalities but also renal tubular defects that lead to neonatal mortality from renal failure. Here, we show that AQP2 is not only a water channel but also an integrin-binding membrane protein that promotes cell migration and epithelial morphogenesis. AQP2 expression modulates the trafficking and internalization of integrin b1, facilitating its turnover at focal adhesions. In vitro, disturbing the interaction between AQP2 and integrin b1 by mutating the RGD motif led to reduced endocytosis, retention of integrin b1 at the cell surface, and defective cell migration and tubulogenesis. Similarly, in vivo, AQP2-null mice exhibited significant retention of integrin b1 at the basolateral membrane and had tubular abnormalities. In summary, these data suggest that the water channel AQP2 interacts with integrins to promote renal epithelial cell migration, contributing to the structural and functional integrity of the mammalian kidney. Aquaporin 2 (AQP2) is a water channel that mediates water absorption through the apical membrane of the principal cells of the kidney collecting ducts, and contributes critically to water homeostasis in mammals. Under physiologic conditions, the trafficking of AQP2 is regulated mainly by vasopressin via a well established signaling pathway that results in the elevation of cAMP, activation of protein kinase A (PKA) and phosphorylation of AQP2. [1][2][3] In addition to regulated trafficking, AQP2 also constitutively recycles between intracellular vesicles and the cell membrane. 4,5 Critical protein/protein interactions that orchestrate the regulated and constitutive trafficking of AQP2 have been extensively investigated and include actin and microtubules, SNAREs, Rab proteins, heat shock protein 70, clathrin, and others. 6-9 However, emerging data from AQP2 knockout and transgenic animals suggested an unusual aspect of AQP2 biology. As expected, induction of AQP2 deficiency in mice results in a urinary concentrating defect known as diabetes insipidus (DI). However, these animals also presented with neonatal mortality from renal failure with renal tubular abnormalities. [10][11][12] This striking phenotype was thought to result from their profound polyuria. However, mice