Recent evidence suggests that vasopressin exerts a regulatory influence on transport processes in the rabbit cortical collecting duct via both the baso-lateral and luminal membranes. The present study was undertaken to examine whether luminal vasopressin receptors, coupled to changes in intracellular calcium, were also present in microperfused rat medullary thick ascending limb (mTAL), a key element of the urine concentrating mechanism. Addition of 1 nM vasopressin to the luminal microperfusate elicited a small but significant and sustained rise in intracellular calcium, from basal values of 100.1 ± 20.1 to 169.6 ± 24.1 nM after 250 s. The effect observed following luminal addition of vasopressin was dose-dependent, with a larger increment of 190.2 ± 32.2nM evoked by addition of 1 µM vasopressin. Addition of 1 µM oxytocin to the lumen did not cause a significant increase in intracellular calcium concentration, consistent with the response to vasopressin being mediated by specific luminal vasopressin receptors. In the absence of calcium in the bath and lumen together or in the bath alone, a residual response to 1 µM luminal vasopressin was still evident, suggestive of a small component of release of calcium from intracellular stores. Selective calcium removal from the luminal microperfusate alone left the response intact. These data are congruous with a model of vasopressin-induced entry of calcium which occurs via the basolateral membrane following ligand binding to the apical membrane. These findings, coupled with earlier observations in the collecting duct, suggest that a fundamental re-assessment of where and how vasopressin, and perhaps other hormones, acts in the kidney may be required. Hormonal regulation of distal tubular function may not therefore be determined only by blood-borne delivery of hormone, but may also involve tubular fluid delivery to apical receptors in distal nephron