Intracellular retention of functional vasopressin V2 receptors (V2R) is a major cause of congenital Nephrogenic Diabetes Inspidus (NDI). Rescue of V2R mutants by pharmacological chaperones may restore their basolateral membrane (BM) localization and function. However, the criteria for efficient rescue of V2R by pharmacological chaperones at clinically feasible concentrations are unknown. The four non-peptide antagonists SR49059, OPC31260, OPC41061 and SR121463B induced maturation and rescued the BM expression of eight V2R mutants, stably expressed in polarized MDCK cells. The extent of maturation and BM localization correlated with the antagonists’ concentration and affinity for the V2R. Displacement of the antagonists by AVP and the subsequent cAMP generation inversely correlated with the antagonists’ affinities for the V2R, but is partially influenced by antagonist-specific effects. The low-affinity SR49059 optimally induced functional rescue at high concentrations, due to its easy displacement by AVP. At clinically feasible concentrations, however, only the high-affinity antagonists OPC31260 and OPC41061 induced functional rescue, as at these concentrations the extent of BM expression became limited. As OPC31260 and OPC41061 are clinically safe, they are promising candidates to relieve NDI. Since receptor activation requires cell-permeable antagonists to be displaced by an agonist, the ability of non-peptide agonists to act as pharmacological chaperones was tested. Six ER-retained mutants responded to treatment with non-peptide agonists, whereas no effect was observed for the peptide agonist AVP. Therefore, non-peptide agonists represent a second class of promising therapeutics to relieve NDI in patients.