Mobilization of intracellular calcium by substance p in a human astrocytoma cell line (U‐373 MG)

Variations in intracellular free calcium concentration (delta[Ca2+]i) were measured in intact and isolated human astrocytoma cells (U373 MG) loaded with fura-2 acetoxymethylester. Microperfusion of 50 nM substance P (SP), applied for 1 s, increased [Ca2+]i by 351 nM from a stable basal level of [Ca2+]i of 26 nM. The peak delta[Ca2+]i induced by SP was dose dependent with a threshold of 10(-3) nM, an ED50 of 1.3 nM and a maximal effect for concentrations of SP greater than 100 nM. The NK1 receptor agonist, [Sar9Met(O2)11]SP, mimicked the effect of SP, while the NK2 and NK3 selective receptor agonists, [N1(10)]NKA(4-10) and senktide, respectively, had no effect. The delta[Ca2+]i induced by SP was unaffected by 100 microM cadmium or by removal of extracellular calcium ions. Caffeine up to 30 mM had no effect on [Ca2+]i. In contrast, thapsigargin increased resting [Ca2+]i by 92 nM and reduced the delta[Ca2+]i induced by SP. A pertussis treatment (500 ng/ml-24 h) did not modify the delta[Ca2+]i induced by SP. We conclude that SP, acting on a NK1 receptor, mobilizes cytosolic calcium from an intracellular calcium pool which can be partially depleted by thapsigargin.