O’DONNELL, M.J.; RHEAULT, M.R.; McMaster University; McMaster University: Analysis of organic anion transport by isolated cells and tissues: A novel, non-invasive electrophysiological technique.

Renal secretion of organic anions (OAs) is ubiquitous in its distribution. It has been observed in the kidneys of a number of vertebrate and invertebrate species. Prototypical OAs include p-aminohippurate (PAH), methotrexate, and salicylate. OA transport is often studied by measuring fluxes of radiolabelled OAs such as the model compound PAH. Our novel method exploits the ~1500-fold higher selectivity of the anion exchanger tridodecylmethylammonium chloride (TDDMACl) for salicylate relative to chloride. The ~3 um diameter tip of a self-referencing ion-selective microelectrode(SeRIS ME) is moved by computer controlled stepper motors (www.applicableelectronics.com), between two points 50 – 100 um apart in the unstirred layer near the surface of the tissue. [Salicylate] is proportional to SeRIS ME voltage, and salicylate flux is proportional to the measured concentration difference. SeRIS MEs have been used to provide excellent spatial and temporal resolution of salicylate fluxes across isolated tissues of the fruit fly Drosophila. Salicylate transport is highest in the secretory (main) segment of the Malpighian (renal) tubule. The K_m for salicylate influx is ~ 0.4 mmol/l, lower than that previously determined for PAH (>1 mmol/l). Influxes 40%-60% as high as those in the MT were observed in the ileum and rectum. We have also employed SERIS MEs based on Fluka chloride ionophore I (cocktail A), which is highly sensitive to indomethacin. Influxes of indomethacin in the MT secretory segment were of comparable magnitude to those of salicylate.