Many human diseases are caused by loss- or gain-of-function mutations in distinct G protein-coupled receptors (GPCRs). The V2 vasopressin receptor (V2R) is a class I GPCR that is almost exclusively expressed in the renal collecting duct system. Most patients (>90%) with congenital nephrogenic diabetes insipidus (NDI) harbor inactivating mutations in the V2R gene which is located on the long arm of the X chromosome. This form of NDI, which is commonly referred to as X-linked NDI (XNDI), therefore almost exclusively manifests itself in males. After its release into the blood stream from the posterior pituitary gland, arginine vasopressin (AVP) binds to renal V2Rs, triggering the activation of the stimulatory G protein, Gs. The resulting elevation in intracellular cAMP levels eventually promotes the insertion of aquaporin-2 (AQP2) water channels into the luminal membrane of the principal cells of the renal collecting duct system. This mechanism allows for the passive movement of water from the tubule lumen into the kidney interstitium and eventually into the blood stream. In XNDI patients, the loss of V2R function interferes with water reabsorption in the renal collecting duct system, resulting in the production of large volumes of dilute urine (polyuria) and excessive thirst and water intake (polydipsia). Hypernatremia and dehydration, which are among the most severe complications associated with XNDI, may cause chronic renal insufficiency as well as mental retardation, at least in a subgroup of XNDI patients. Large dilatations of the urinary tract and bladder as well as kidney failure secondary to bilateral hydronephrosis represent long-term complications of the disease. Moreover, newborns suffering from XNDI frequently often show poor weight gain and an overall failure to thrive, sometimes complicated by convulsions or death due to hypertonic dehydration, especially if XNDI is not diagnosed early enough. At present, no specific, effective pharmacological therapy exists for the treatment of XNDI, primarily due to the lack of suitable animal models. We previously created V2R knockout (KO) mice lacking functional V2Rs throughout development (Yun et al. J Clin Invest 106, 1361-1371, 2000). However, none of the mutant XNDI pups survived the first postnatal week. During the past year, we were able to establish the first viable mouse model of XNDI. In these newly developed mutant mice, the V2R gene can be deleted in a conditional (4-OH-tamoxifen-dependent) fashion in the kidneys of adult mice. The resulting V2R KO mice showed all key symptoms of XNDI, including the production of large amounts of dilute urine (polyuria) and polydipsia. Following dDAVP administration, the V2R mutant mice were unable to increase their urine osmolality, consistent with the complete absence of renal V2R binding sites observed in radioligand binding assays. Kidneys from V2R KO mice also showed a distension of the renal pelvis, a characteristic morphological deficit that is typically a consequence of massive polyuria. Immunoblotting studies showed that renal AQP2 and AQP3 expression levels were drastically reduced (by 70-80%) in V2R KO mice. AQP2 and AQP3 are two water channels that play a key role in mediating V2R-dependent water reabsorption. We speculated that renal collecting duct cells might express Gs-coupled receptors (besides the V2R) that could serve as targets for the treatment of XNDI. We demonstrated that mouse inner medullary collecting cells express another Gs-coupled receptor, the EP4 prostanoid receptor, at significant levels. To test the potential usefulness of EP4 receptor agonists for the therapy of XNDI, we treated V2R KO mice with the selective EP4 receptor agonist, ONO-AE1-329 (ONO). Remarkably, both acute and chronic treatment of V2R mutant mice with ONO greatly reduced all major manifestations of XNDI, leading to striking reductions in urine output and water intake and pronounced increases in urine osmolality. Moreover, MRI studies showed that prolonged ONO treatment of V2R KO mice could prevent the further progression of renal pelvic distension normally seen with V2R KO mice. We also demonstrated that prolonged treatment of V2R KO mice with ONO increased renal AQP2 levels by 70%, probably due to EP4 receptor-mediated elevations of cAMP levels in kidney collecting duct cells. ONO treatment of kidney collecting duct tubule preparations led to a pronounced increase in cAMP levels and enhanced water permeability, consistent with the concept that the beneficial effects of ONO in the treatment of XNDI mice are due to a direct action on renal EP4 receptors. Taken together, we have generated the first viable mouse model of human XNDI. Our findings suggest that selective EP4 receptor agonists may represent a new class of drugs useful for the treatment of XNDI, due to their direct action on kidney collecting duct cells. The newly generated V2R mutant mice should prove generally useful for testing new drugs for the therapy of XNDI.

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