These studies examine the properties of an apical potassium (K+) channel in macula densa cells, a specialized group of cells involved in tubuloglomerular feedback signal transmission. To this end, individual glomeruli with thick ascending limbs (TAL) and macula densa cells were dissected from rabbit kidney and the TAL covering macula densa cells was removed. Using patch clamp techniques, we found a high density (up to 54 channels per patch) of K+ channels in the apical membrane of macula densa cells. An inward conductance of 41.1 ± 4.8 pS was obtained in cell-attached patches (patch pipette, 140 mM K+). In inside-out patches (patch pipette, 140 mM; bath, 5 mM K+), inward currents of 1.1 ± 0.1 pA (n = 11) were observed at 0 mV and single channel current reversed at a pipette potential of -84 mV giving a permeability ratio (PK/PNa) of over 100. In cell-attached patches, mean channel open probability (N.Po, where N is number of channels in the patch and Po is single channel open probability) was unaffected by bumetanide, but was reduced from 11.3 ± 2.7 to 1.6 ± 1.3 (n = 5, p < 0.02) by removal of bath sodium (Na+). Simultaneous removal of bath Na+ and calcium (Ca2+) prevented the Na+-induced decrease in N.P0 indicating that the effect of Na+ removal on N.P0 was probably mediated by stimulation of Ca2+ entry. This interpretation was supported by studies where ionomycin, which directly increases intracellular Ca2+, produced a fall in N.Po from 17.8 ± 4.0 to 5.9 ± 4.1 (n = 7, p < 0.02). In inside-out patches, the apical K+ channel was not sensitive to ATP but was directly blocked by 2 mM Ca2+ and by lowering bath pH from 7.4 to 6.8. These studies constitute the first single channel observations on macula densa cells and establish some of the characteristics and regulators of this apical K+ channel. This channel is likely to be involved in macula densa transepithelial Cl− transport and perhaps in the tubuloglomerular feedback signaling process. © 1994, Rockefeller University Press., All rights reserved.