FIGURE 7.

NHERF-2 silencing attenuates relative Ca²⁺ recovery in HT-29 cells following exposure to ACh. Cells were transfected with NHERF-2 silencing constructs as described and [Ca²⁺]_i assessed using a Fura-2-based fluorometric assay. For all experiments, agonist injection was at 20 s. A, representative Fura-2 tracings showing relative PMCA-mediated [Ca²⁺]_i recovery following an ACh-evoked Ca²⁺ transient (bath solutions; zero Ca²⁺, 5 μm CPA, extracellular BAPTA). B, relative PMCA-mediated Ca²⁺ efflux was significantly reduced by 32 ± 8% and 31 ± 6% (n = 4; ^**, p < 0.01) for Shag 274 and Shag 361, respectively. C, no significant change in peak [Ca²⁺]_i was observed. D, representative Fura-2 tracings showing relative [Ca²⁺]_i recovery following an ACh-evoked Ca²⁺ transient in the absence of CPA (bath solution; zero Ca²⁺, extracellular BAPTA). E, relative Ca²⁺ efflux was significantly reduced by 50 ± 3% and 53 ± 3% (n = 4; ^***, p < 0.001) for Shag 274 and Shag 361, respectively. F, no significant change in peak [Ca²⁺]_i was observed. All data are representative of 3-4 independent assays where each group was measured from 6-8 wells and are expressed as mean ± S.E.