ACTIVATION BY CALCIUM ALONE OF CHLORIDE SECRETION IN T(84) EPITHELIAL-CELLS

1 The goal of this study was to determine if an increase in cytoplasmi c calcium concentration ([Ca2+]i), in the absence of additional second messengers derived from membrane phospholipid turnover, is a sufficie nt signal to induce chloride secretion across monolayers of the human colonic epithelial line, T84. 2 Thapsigargin was used to increase [Ca2 +]i by inhibiting the endomembrane Ca2+-ATPase. [Ca2+]i was monitored in monolayers by fura-2 fluorescence spectroscopy, chloride secretion by measuring changes in short circuit current (I(sc)) in modified Ussi ng chambers, and inositol phosphates were measured by radio-h.p.l.c. o f extracts of cells prelabelled with [H-3]-inositol. 3 Thapsigargin in creased [Ca2+]i and I(sc) in parallel, without increasing any inositol phosphates. The effect of thapsigargin on I(sc) was abolished by the intracellular calcium chelator, bis-(o-aminophenoxy)-ethane-N,N,N',N'- tetraacetic acid (BAPTA). 4 Increasing [Ca2+]i with thapsigargin did n ot prevent a subsequent calcium response to carbachol or histamine if extracellular calcium was available. In the absence of extracellular c alcium, only one such release of calcium to hormonal stimulation occur red when cells were pretreated with thapsigargin, and a second respons e to either carbachol or histamine was essentially abolished. 5 Additi on of carbachol or histamine to thapsigargin-treated cells mounted in Ussing chambers caused a transient further increase in I(sc) followed by termination of the response, even though [Ca2+]i continued to rise. 6 We conclude that an elevation in [Ca2+]i is a sufficient signal to induce chloride secretion in T84 cells. Rather than being required to stimulate secretory responses, additional second messengers induced by hormonal secretagogues (such as inositol phosphates) may in fact serv e to limit the secretory response.