Early Actions of Parathyroid Hormone and 1,25-Dihydroxycholecalciferol on Isolated Epithelial Cells from Rat Intestine: II. Analyses of Additivity, Contribution of Calcium, and Modulatory Influence of Indomethacin*

Abstract

Epithelial cells isolated from rat intestine were analyzed for responsiveness to treatment in vitro with parathyroid hormone (PTH) and 1,25-dihydroxycholecalciferol [1,25- (OH)₂D₃], alone and in combination, by the criteria of augmented extracellular release of lysosmal enzyme activities and enhanced uptake of ⁴⁵Ca. A 20-min exposure of cells to 1 pM PTH, 65 pM 1,25-(OH)₂D₃, or both hormones together, enhanced ⁴⁵Ca uptake relative to controls. The combined influence of both hormones appeared to be additive by this criterion, suggesting that the two act through different receptors. In contrast, hormone-augmented liberation of three lysosomal hydrolases and alkaline phosphatase reached a maximal level relative to controls in response to either PTH or 1,25-(OH)₂D₃ that was not exceeded in cells treated with the combined hormones. The latter results may indicate a common target cell-type for both hormones. In additional experiments, intestinal cell preparations were treated with the ionophore A23187 to analyze the potential contribution of calcium uptake to hydrolase release. The ionophore promoted liberation of acid phosphatase and N-acetyl-β-D-glucosaminidase to 112% and 123% of controls, respectively, but not cathepsin B or alkaline phosphatase (101% and 99% of controls, respectively), suggesting the likelihood of a calcium-effector coupling mechanism for full reproduction of acute hormone action on membrane loci. This interpretation is supported by the observation that neither PTH nor 1,25-(OH)₂D₃, added directly in vitro to lysosomes isolated from intestinal cells, led to reduce latency of hydrolases. Moreover, isolated intestinal cells treated with indomethacin followed by either PTH or the steroid hormone responded with enhanced ⁴⁵Ca uptake, but iiot with augmented liberaof the hydrolases, relative to corresponding preparations treated with PTH or 1,25-(OH)₂D₃ in the absence of the drug. These data suggest that in intestinal cells, calcium mediates an additional step in hormone-accentuated lysosome activation that is blocked by indomethacin.