Characterization of whole cell chloride conductances in a mouse inner medullary collecting duct cell line mIMCD-3

The chloride conductance of inner medullary collecting duct cells (mIMCD-3 cell line) has been investigated using the whole cell configuration of the patch clamp technique. Seventy-seven percent of cells were chloride selective when measured with a NaCl-rich bathing solution and a TEACl-rich pipette solution. Seventy-five percent of chloride-selective cells (90/144) had whole cell currents which exhibited an outwardly-rectifying (OR) current-voltage (I/V) relationship, while the remaining cells exhibited a linear (L) I/V relationship. The properties of the OR and L chloride currents were distinct. OR currents (mean current densities at +/- 60 mV of 66 +/- 5 pA/pF and 44 +/- 3 pA/pF), were time- and voltage-independent with an anion selectivity (from calculated permeability ratios) of SCN- (2.3), NO3- (1.8), ClO4- (1.7), Br- (1.7), I- (1.6), Cl- (1.0), HCO3- (0.5), gluconate- (0.2). Bath additions of NPPB, flufenamate, glibenclamide (all 100 microM) and DIDS (500 microM) produced varying degrees of block of OR currents with NPPB being the most potent (IC50 of approximately 50 microM) while DIDS was the least effective. Linear chloride currents had similar current densities to the OR chloride currents and were also time- and voltage-independent. The anion selectivity sequence was SCN- (2.5), NO3- (1.9), Br- (1.4), I- (1.1), Cl- (1.0), ClO4- (0.5), HCO3- (0.5), gluconate- (0.3). In contrast to the OR conductance, glibenclamide was the most potent and DIDS the least potent blocker of L currents. An IC50 of > 100 microM was observed for NPPB block. Neither OR of L chloride currents were affected by acutely or chronically increased intracellular cAMP and were not affected when intracellular Ca2+ levels were increased or decreased. The molecular identity and physiological role of OR and linear currents in mIMCD-3 cells are discussed.