The effects of extracellular polyvalent cations on the cytosolic free Ca2+ concentration ([Ca2+]i) of isolated segments of the mouse nephron were investigated using fura 2 microfluorometry. Extracellular Ca2+ concentration ([Ca2+]o), gadolinium (Gd3+), and neomycin (Neo) increased the [Ca2+]i in cortical thick ascending limb (CTAL) tubules with effective doses (ED50) of approximately 3.5 mM for Ca2+, 20 microM for Gd3+, and 40 microM for Neo. This effect was reproduced by Ba2+ but not by Mg2+. High [Ca2+]o inhibited the responses to Gd3+, Neo, and Ba2+. The Gd(3+)- and Neo-evoked [Ca2+]i transients persisted in the absence of external Ca2+ and were abolished by the depletion of internal Ca2+ stores with thapsigargin (TG). The responses to rises in [Ca2+]o were similarly inhibited by TG and slightly reduced by 20 microM La3+ but not by 10 microM nifedipine. Mn2+ also mobilized a TG-sensitive internal Ca2+ store and stimulated its own entry. External Ca2+, Gd3+, and Neo induced small but significant increases in [Ca2+]i in distal convoluted tubule, cortical collecting duct, and outer medullary collecting duct segments, transiently increased [Ca2+]i in some medullary TAL (MTAL) tubules, but had no effect on descending thin limb. We conclude that a Ca(2+)-mobilizing Ca2+/polyvalent cation sensor resembling that of the parathyroid gland cells is predominantly located in the mouse CTAL but also in the MTAL and, to a lesser extent, in more distal segments.