h-89 and 5--adenylyl-(beta-gamma-methylene)diphosphonate

In our previous observations, adenosine triphosphate (ATP) was found to evoke immediate elevations in intracellular free calcium concentration ([Ca2+]i) in HT4 neuroblastoma cells of mice. We tried to see if a brief pretreatment of glucocorticoids could inhibit the Ca2+ response and reveal the underlying signaling mechanism.. Measurement of [Ca2+]i was carried out using the dual-wavelength fluorescence method with Fura-2 as the indicator.. Pre-incubation of HT4 cells for 5 min with corticosterone (B) or bovine serum albumin conjugated corticosterone (B-BSA) inhibited the peak [Ca2+]i increments in a concentration-dependent manner. Cortisol and dexamethasone had a similar action, while deoxycorticosterone and cholesterol were ineffective. Both extracellular Ca2+ influx and internal Ca2+ release contributed to ATP-induced [Ca2+]i elevation. The brief treatment with only B attenuated Ca2+ influx. Furthermore, the [Ca2+]i elevation induced by the P2X receptor agonist adenosine 5'-(beta, gamma-methylene) triphosphate (beta, gamma-meATP) was also suppressed. The rapid inhibitory effect of B can be reproduced by forskolin 1 mmol/L and blocked by H89 20 mmol/L. Neither nuclear glucocorticoid receptor antagonist mifepristone nor protein kinase C inhibitors influenced the rapid action of B.. Our results suggest that glucocorticoids modulate P2X receptor-medicated Ca2+ influx through a membrane-initiated, non-genomic and PKA-dependent pathway in HT4 cells.

Topics: Adenosine Triphosphate; Animals; Calcium; Cell Line, Tumor; Corticosterone; Cyclic AMP-Dependent Protein Kinases; Dexamethasone; Dose-Response Relationship, Drug; Hydrocortisone; Isoquinolines; Mice; Neuroblastoma; Purinergic P2 Receptor Agonists; Receptors, Purinergic P2X; Serum Albumin; Signal Transduction; Sulfonamides