Involvement of intracellular and extracellular Ca 2+ in tetramethylpyrazine‐induced colonic anion secretion

In the present study we investigated the role of Ca 2+ in tetramethylpyrazine (TMP)‐induced anion secretion in the human colonic epithelial cell line, Caco‐2, using the short‐circuit current ( I SC ) technique in conjunction with intracellular Ca 2+ measurements. The results showed that TMP‐induced I SC response was significantly reduced by 58.8% and 38.3% after inhibiting Ca 2+ ATPase of endoplasmic reticulum (ER) with thapsigargin and mobilizing ER stored Ca 2+ release with ATP, respectively. Conversely, thapsigargin‐ and ATP‐evoked I SC responses were also significantly reduced by pretreatment with TMP by 43.2% and 38.5%, respectively. Conversely, removal of extracellular Ca 2+ , apical but not basolateral, or the presence of the Ca 2+ chelator (EGTA) significantly increased TMP‐induced I SC by 47.1% and 37.8%, respectively. Similar to TMP, thapsigargin‐induced current increase was also enhanced by chelating extracellular Ca 2+ or in Ca 2+ free solution; however, removal of extracellular Ca 2+ did not significantly affect 3‐isobutyl‐1‐methylxanthine (IBMX)‐ and forskolin‐induced transepithelial current. Measurement of the intracellular concentration of free Ca 2+ ([Ca 2+ ] i ) with fura‐2/AM showed that TMP could induce an increase in [Ca 2+ ] i but pretreatment with TMP significantly reduced thapsigargin‐evoked, but not ATP‐induced, [Ca 2+ ] i increase. These results suggest that the effect of TMP on colonic anion secretion is partly mediated by TMP‐increased [Ca 2+ ] i by acting on a target similar to thapsigargin. The observed inhibitory effect of extracellular Ca 2+ on Ca 2+ ‐dependent anion secretion represents a novel mechanism by which Ca 2+ ‐dependent regulation of epithelial electrolyte transport may be fine‐tuned by extracellular Ca 2+ in the apical domain.