The macula densa is a unique group of cells that are located within the thick ascending limb at the point where this segment of the loop of Henle comes in contact with its own glomerulus. Macula densa cells detect changes in tubular fluid sodium chloride concentration ([NaCl]) and then transmit information to the underlying mesangial cell/afferent arteriolar complex. This process is called tubuloglomerular feedback and serves to couple tubular flow and ion composition with the regulation of glomerular filtration and blood flow. To date, the nature of the signaling process between macula densa cells and the mesangial cell/afferent arteriolar complex has remained elusive. However, recent studies have now suggested that macula densa cells signal through the release of ATP. NaCl entry into macula densa cells occurs via a Na:2Cl:K cotransporter, thereby leading to a number of cellular responses including elevations in cytosolic [NaCl], basolateral membrane depolarization, changes in cell volume, and increases in intracellular pH and calcium. In contrast to most other cells, intracellular [Na] is removed from macula densa cells via an apically located colonic form of the H:K ATPase. Since the basolateral area of macula densa cells contains a large abundance of mitochondria and very little Na:K ATPase activity, it is possible that this area serves as a reservoir or sink for ATP. In recent patch clamp studies, a large maxi-anion channel has been found at the basolateral membrane of macula densa cells. It has a conductance of ~380 pS and is blocked by Gd3+. Using inside-out patches, this maxi-anion channel was shown to be permeable to ATP. At the present time, the molecular identity of this channel remains unknown. In additional studies, a biosensor assay was developed using either PC12 cells or cultured mesangial cells loaded with fura-2 and placed at the basolateral membrane of macula densa cells. ATP release across the basolateral membrane was measured to be in the range of 5 to 10 ?M. This is well within the range of ATP concentrations that will activate purinergic receptor mediated changes in cytosolic calcium concentration in mesangial cells. Importantly, ATP release was consistent with macula densa cell signaling, including the fact that ATP release was abolished by loop diuretics that inhibit the apical Na:2Cl:K cotransporter. Also, prior salt deprivation, which has been shown to up-regulate macula densa cell signaling likewise caused enhanced release of ATP release. Other work has utilized a mouse cell culture line that has some of the properties of macula densa cells (from J. Schnermann, NIH). Cells were grown on permeable supports and basolateral release of ATP was detected with a luminometer using the luciferase/luciferin-reaction. ATP was released by these cells and could also be inhibited by furosemide or Gd3+. One candidate for the maxi-anion channel in macula densa cells is the voltage dependent anion channel (VDAC, or porin), which serves as a mithocondrial ATP channel. VDAC and the MD maxi-anion channel have similar electrophysiological characteristics, and recent work has identified a plasma membrane isoform of VDAC (plVDAC). In the macula densa cell line mRNA was isolated and used in RT-PCR to identify mithocondrial VDAC’s (VDAC1, 2, 5). Importantly, the mRNA for plVDAC was also present in this macula densa like cell line. These studies indicate that macula densa cells transmit information through the release of ATP via a maxi-anion channel and purinergic signaling.