The human organic cation transporter 2 (hOCT2) has been mainly detected in the basolateral membrane of proximal tubules of human kidneys and is thought to be the major transporter for excretion of various organic cations. Beside protein kinase A- and phosphatidylinositol-3-kinase-dependent regulatory pathways, the transport rate of hOCT2 is strongly influenced by inhibition of the Ca²⁺/calmodulin-complex (Ca²⁺/CaM) with calmidazolium (Cetinkaya et al., Am J Physiol Renal Physiol 284: F293-F302, 2003). Aim of this study was to investigate the mechanisms of hOCT2-regulation by Ca²⁺/CaM. The function of hOCT2 stably expressed in HEK-cells was examined microfluorimetrically with the cation 4-[4-(dimethylamino)-styryl]-N-methylpyridinium (ASP) as substrate. The expression of membrane-associated hOCT2 was quantified by FACScan measurements using a specific antibody against the extracellular loop of hOCT2. The K_m-values for ASP-uptake were not significantly influenced by calmidazolium incubation (29 mM without and 26 mM with calmidazolium), while V_max decreased significantly (160 photons/sec to 99 photons/sec). Calmidazolium led to a significant decrease of membrane-associated fluorescence (-22±6%, n=9). These results imply that inhibition of Ca²⁺/CaM causes a change of V_max probably via hOCT2-trafficking.