Kappa opioid receptor (KOR) agonists produce a variety of beneficial effects such as nonaddictive analgesia and diuresis, but their translation into the clinic has been hindered by central adverse effects (dysphoria). In clinical studies, difelikefalin, a KOR agonist recently FDA approved for treatment of pruritus in hemodialysis patients, was shown to be devoid of producing adverse CNS effects suggesting that this compound is peripherally restricted. This is of interest from a renal excretory perspective, since it would be predicted that difelikefalin would not produce diuresis, which is a classical centrally mediated response produced by KOR agonists. However, in preliminary studies in rats we have shown that IV administration of difelikefalin markedly increased urine output and decreased urinary sodium and potassium excretion similar to that produced by other KOR agonists. Therefore, we hypothesized that difelikefalin activates a central KOR pathway to cause diuresis. To test this hypothesis, Sprague-Dawley rats were implanted with bladder, femoral artery, and femoral vein catheters and administered an isotonic saline infusion. To delineate the central and/or peripheral site of action of difelikefalin, renal excretory function was measured in rats pretreated centrally or peripherally with norBNI (KOR antagonist) or vehicle via an intracerebroventricular (ICV) cannula or IV catheter. Following stabilization, conscious rats were administered an IV bolus of difelikefalin (10 µg/kg) and urine output, mean arterial pressure (MAP), and heart rate (HR) were recorded for 90 minutes. The results demonstrated that ICV norBNI pretreatment significantly decreased the diuretic response to IV difelikefalin but had no effect on urinary sodium or potassium concentration (Table 1). In contrast, IV norBNI pretreatment prevented the diuresis to IV difelikefalin and increased urinary sodium/potassium concentration (Table 1). Notably, ICV and IV norBNI pretreatment abolished the difelikefalin-induced decrease in MAP that was also seen in the vehicle pretreated groups. Together, these findings suggest that difelikefalin produces its diuretic and blood pressure effects through a central KOR pathway, while the sodium/potassium retaining effects of difelikefalin are mediated through peripheral KORs.