One of the major functions of the prostate gland is the production and accumulation of high levels of citrate due to the tissue-specific Zn2+-dependent activity of a rate-limiting oxidizing enzyme, m-aconitase (Costello et al. 1997). Citrate is released into the lumen by a membrane transport mechanism which at present is not well understood.
We adopted the normal human prostate cell line PNT2-C2 to characterise citrate transport electrophysiologically. Two different experimental protocols were used: citrate (sodium salt) was introduced into the cells either via the intracellular patch pipette or the cells were perfused with extracellular citrate. The membrane currents induced under voltage-clamp conditions were recorded. Data were analysed as means ± S.E.M.
At a holding potential of -45 mV, intracellular citrate (0.5 or 10 mM) induced an outward current (Icit) which slowly reached values of 124 ± 17 and 180 ± 25 pA, respectively (n > 30). Equimolar intracellular NaCl had no effect. Other Krebs cycle intermediates like malate, succinate and isocitrate were 50-70 % less effective. Extracellular citrate (10 mM) elicited much smaller, inward currents of -42 ± 8.1 pA. Application of intracellular citrate had no effect on normal breast epithelial MCF-10A cells. Increasing the extracellular K+ concentration from 2.5 to 118 mM caused a significant, dose-dependent decrease in Icit. On the other hand, changes in extracellular Na+, Cl–, Ca2+ or Mg2+ concentrations had no influence. The following pharmacological agents potentiated (+) or inhibited (-) Icit: phloretin, +14% diethyl pyrocarbonate, -18% LiCl, -14% and 4-aminopyridine, -13 %. SCH28080, ouabain and dinitrophenol did not produce any change (n > 10 for each). The effects of the ionic changes and the pharmacological agents were essentially the same on the current elicited by extracellular citrate, consistent with both currents being due to the activity of the same transport mechanism. We propose that normal prostatic epithelial cells express a novel citrate transporter which is electrogenic and co-transports trivalent citrate anion with K+ outward.
This research was supported by The Wellcome Trust.