Differences in Intracellular Ca 2+ Stores of Submandibular Cells of Adult and Newborn Rats

Abstract

The intracellular Ca²⁺ stores of submandibular acinar cells of adult and newborn rats were compared by measuring changes in cytosolic Ca²⁺ ([Ca²⁺]_i) with the Ca²⁺-sensitive fluorescent probe, fura-2, in intact cells and ⁴⁵Ca²⁺ release in permeabilized cells. In cells of adult rats, acetylcholine (ACh) and thapsigargin (TG) elicited, respectively, a 105% and 125% net increase in [Ca²⁺]_i in Ca²⁺-free medium. In cells of newborn animals, the ACh-induced [Ca²⁺]_i increase (218%) was larger; but the TG-induced increase was significantly smaller (77%), suggesting that the TG-sensitive Ca²⁺ pump of the store is less developed, abundant, or responsive in these cells. After ACh or TG, ionomycin elicited a further Ca²⁺ release, Ionomycin by itself completely discharged the ACh- and TG-sensitive store in cells of mature glands but not in cells of immature glands, suggesting a further difference in the distribution of the agonist and ionomycin sensitivities in the two types of cells. After exposure to ACh + ionomycin or TG + ionomycin, collapsing the intracellular pH gradient with monensin caused a third and large Ca²⁺ release (262% and 289%, respectively) in cells of adult rats, but a significantly smaller release (96%, P < 0.001, and 57%, P < 0.001, respectively) in cells of newborn rats, suggesting that a third, IP₃-insensitive acidic store is significantly smaller in cells of immature glands. In permeabilized and ⁴⁵Ca²⁺-loaded cells, inositol-1,4,5-trisphosphate (IP₃, 5 μM) induced an identical ⁴⁵Ca²⁺ release (34% and 33%, respectively) in both types of cells, but TG induced a significant smaller ⁴⁵Ca²⁺ release in cells of immature glands (15%) than in cells of mature glands (33%). Monensin discharged <10% of loaded ⁴⁵Ca²⁺ in both types of cells, indicating that the radiotracer was not loaded into the monensin-sensitive store. These results suggest that in cells of immature glands: (1) the TG-sensitive Ca²⁺ pump of the IP₃-sensitive store is not well developed; (ii) the IP₃-sensitive store is not completely discharged by ionomycin; and (iii) an acidic store likely to be associated with secretory granules is also underdeveloped and of a smaller size than in cells of mature glands.