A calcium–sensing receptor (CaR) has functionally been described in the cortical thick ascending limb of Henle's loop (CTAL) of rat and mouse. This G protein–coupled receptor activates phospholipase C and increases the intracellular Ca²⁺ concentration. We observed that in the mouse CTAL cAMP formation, induced by 10^–8 mol/l AVP, was inhibited by more than 90% when the extracellular Ca²⁺ concentration ([Ca²⁺]_e) was increased from 0.5 to 3 mmol/l. Measurements of transepithelial potential difference (PD_te) in rat and mouse CTAL and medullary thick ascending limb (mTAL) segments and of transepithelial ion net fluxes in the mouse CTAL (isotonic perfusion conditions: 150 mmol/l NaCl in the lumen and bath) showed that an increase in the [Ca²⁺]_e had no effect on basal and arginine vasopressin (AVP, 10^–10 mol/l)–stimulated transepithelial PD_te, NaCl and Mg²⁺ transport. However, Ca²⁺ reabsorption was strongly inhibited by increased [Ca²⁺]_e. Addition of AVP reversed this inhibitory effect of increased [Ca²⁺]_e. Under hypotonic perfusion conditions (lumen 50 mmol/l NaCl; bath 150 mmol/l NaCl), a high [Ca²⁺]_e induced a 50% decrease in Mg²⁺ reabsorption which was restored by AVP. Under these conditions, the effects on Ca²⁺ transport described above were still observed. In conclusion, activation of the CaR in the mouse TAL has no effect on basal and AVP–stimulated transepithelial NaCl reabsorption despite its large inhibitory effect on cAMP synthesis. The CaR, however, could play a role in the regulation of transepithelial Ca²⁺ and Mg²⁺ reabsorption.