FIGURE 3.

O-GlcNAc elevation obtained by pre-incubation of cells with PUGNAc and glucosamine blunts IClswell in ICln-transfected cells. IClswell activation was monitored in HEK 293 Phoenix cells transfected for 24 h with ICln as well as the transfection marker EGFP as separate proteins. Single transfected cells were selected and voltage clamped using the whole-cell patch-clamp technique. (A) Original recordings obtained in isotonic (upper panels) and hypotonic (lower panels) extracellular solutions in cells pre-incubated for 1 h with 0.1% DMSO plus 5 mM D-glucose (left panels) or 100 μM PUGNAc plus 5 mM glucosamine (right panels) and stimulated with voltage increments of 20 mV from –120 to +100 mV applied from a holding potential of 0 mV (lower right inset). (B) Current density-to-voltage relationship determined after a 20-min exposure to the extracellular hypotonic solution (left) and current density-to-time relationship (right) of IClswell in cells pre-incubated with 100 μM PUGNAc plus 5 mM glucosamine or the vehicle. *p < 0.05, unpaired Student’s t-test. (n) refers to the number of cells. EGFP, enhanced green fluorescent protein.