Effect of Heat Shock, (Ca(2+))-i, and cAMP on Inositol Trisphosphate in Human Epidermoid A-431 Cells
Abstract
The basal levels of inositol monophosphate, inositol bisphosphate, and inositol trisphosphate (InsP3) in A-431 cells incubated in Na+ Hanks' solution were, respectively, 1.23 +/-0.18%, 0.17+/-0.03%, and 0.69+/-0.07% of the total radioactivity in the cell. When cells were heated, InsP3 increased in a temperature-dependent manner related to the duration of heating. The active form of InsP3, inositol 1,4,5-trisphosphate, increased 237 +/- 17% after heating (45 C. 20 min) then returned to baseline within 15 min in InsP3 was not observed in the absence of extracellular Ca2+ or with amiloride treatment. Treatment with the nonhydrolyzable GTP analog Gpp(NH)p stimulated that component of the InsP3 increase due to G proteins. U-73122, an inhibitor of PL-C-mediated processes blocked the increase in InsP3 resulting from heat exposure. Both pertussis toxin (30 ng/mL, 24 h), an inhibitor of G inhibitory protein, and cholera toxin (I microgram/mL, 1 h), a stimulator of G-stimulatory protein, increased InsP3 in unheated cells, and heating failed to induce a further increase, suggesting heat activates G proteins. Likewise, 8-Br-cAMP, IBMX, Ro 20-1724, or forskolin increased InsP3 in unheated cells, and heat did not cause an additional increase. The InsP3 increase induced by 8-Br-cAMP was inhibited by removal of extracellular Ca2+ or treatment with verapamil, suggesting that an influx of extracellular Ca2+ stimulates InsP3 production. These data are the first to suggest that the heat-induced increase in InsP3 results from both an increase in intracellular Ca2+ concentration and activation of G proteins, while the cAMP- induced increase is due to a rise in intracellular Ca2+ alone.
Document Details
- Document Type
- Technical Report
- Publication Date
- Jan 01, 1993
- Accession Number
- ADA269092
Entities
People
- David E. Mcclain
- Juliann G Kiang
Organizations
- Walter Reed Army Institute of Research