Extracellular nucleotides regulate epithelial transport via luminal and basolateral P2 receptors. Renal epithelia abundantly express P2 receptors which mediate significant inhibi tion of solute absorption. Recently, we found several P2 receptors in the medullary thick ascending limb (mTAL) including luminal and basolateral P2Y₂ receptors. In addition, we found evidence for a basolateral P2X receptor. It is currently not known if extracellular nucleotides influence transport in this segment. Objective: Find how extracellular ATP influences transport in mTAL. Methods: In this study we use isolated, perfused mTAL from mice to electrically measure Na⁺ absorption. By microelectrodes we determined the transepithelial voltage (V_te) and the transepithelial resistance (R_te) and via these the transepithelial Na⁺ absorption (equivalent short circuit current, I_sc). Results: Non-stimulated mTALs show large transepithelial transport and were characterized by: V_te: +9.030.44 mV, (lumen-positive), R_te: 81.1 W cm², I_sc: 140421.4 mA/cm² (n=16). As expected, luminal furosemide (100 M) completely blocked this transport. Basolateral ATP (100 M, for 10 minutes) acutely (within 1 minute) reduced the absorptive I_sc. After 3 minutes a maximal reduction was measured and amounted to 19.62.8% (n=8). In most experiments transport inhibition was sustained during the application of ATP. Basolateral UTP, a P2Y₂/P2Y₄ receptor agonist was without effect as was adenosine. Conclusion: These data define that basolateral ATP exerts a significant inhibition of Na⁺ absorption in mouse mTAL. Our data point to a P2X receptor-mediated mechanism. Intriguingly, these data add yet another example of P2 receptor mediated inhibition of tubular transport in intact renal epithelium.