Arachidonic Acid Stimulates Intracellular Calcium Mobilization and Regulates Protein Synthesis,ATP Levels,and Mucin Secretion in Submandibular Gland Cells

Abstract

Earlier observations that arachidonic acid inhibited the synthesis of membrane inositol phospholipids in rat submandibular acinar cells prompted the present study on whether the fatty acid may also regulate other key physiological processes in the model. Arachidonate, at concentrations above 10 pmol/L, inhibited up to 97% protein synthesis in acinar cells. The acid also lowered cellular ATP levels to 25% of control values by a ouabaininsensitive mechanism. In endoplasmic reticulum-calcium studies in permeabilized cells, arachidonic acid stimulated the mobilization of up to 73% loaded ER-⁴⁵Ca²⁺ to the cytosol, a much greater response than those caused by other calcium translocators, thapsigargin or inositol 1,4,5-trisphosphate. Additionally, arachidonate provoked the release of over 80% of total cell ⁴⁵Ca²⁺ to the extracellular space in intact cells and stimulated mucin secretion in the submandibular model. The inhibitory effect of arachidonic acid on protein synthesis was duplicated by carbachol, thapsigargin, and BAPTA/AM, three agents that cause net efflux of ER-Ca²⁺ by different mechanisms. Furthermore, comparable with the arachidonate effect on ATP, carbachol and thapsigargin also significantly reduced cellular levels of the nucleotide. It is concluded that arachidonic acid acts as a regulator of central synthetic/secretory processes in mucous acinar cells of rat submandibular gland and suggested that at least some of its effects may be secondary to its calcium-mobilizing action.

Keywords

arachidonic acid submandibular gland calcium mobilization protein synthesis ATP

Get full access to this article

View all access options for this article.

References

Brostrom CO , Bocckino SB , Brostrom MA , Galuska EM (1986). Regulation of protein synthesis in isolated hepatocytes by calcium- mobilizing hormones. Molec Pharm 29:104-111.

Chaudhry A. , Laychock SG , Rubin RP (1987). The effects of fatty acids on phosphoinositide synthesis and myo-inositol accumulation in exocrine pancreas. J Biol Chem 262 :17426-17431.

Chaudhry A. , Conway BR , Laychock SG , Rubin RP (1989). Analysis of the regulation of phosphatidylinositol-4,5-bisphosphate synthesis by arachidonic acid in exocrine pancreas. Arch Biochem Biophys 272:488-495.

Chow SC , Jondal M. (1990). Polyunsaturated free fatty acids stimulate an increase in cytosolic Ca2+ by mobilizing the inositol 1,4,5-trisphosphate-sensitive Ca2+ pool in T cells through a mechanism independent of phosphoinositide turnover . J Biol Chem 265:902-907.

Chung HC , Fleming N. (1992). Arachidonic acid regulates the phosphoinositide signal transduction pathway in submandibular acinar cells. J Dent Res 71:1462-1467.

Chung HC , Fleming N. (1995). Regulation of phosphatidylinositol kinases by arachidonic acid in rat submandibular gland cells. Eur J Physiol 429:789-796.

Fleming N. , Bilan PT , Sliwinski-Lis E. (1986). Effects of a phorbol ester and diacylglycerols on secretion of mucin and arginine esterase by rat submandibular gland cells . Eur J Physiol 406:6-11.

Fleming N. , Bilan PT , Sliwinski-Lis E. , Carvalho V. (1987). Muscarinic, α1-adrenergic and peptidergic agonists stimulate phosphoinositide hydrolysis and regulate mucin secretion in rat submandibular gland cells. Eur J Physiol 409:416-421.

Fleming N. , Sliwinski-Lis E. , Burke DN (1989). G regulatory proteins and muscarinic receptor signal transduction in mucous acini of rat submandibular gland. Life Sci 44 :1027-1035.

10.

Fleming N. , Mellow L. , Bhullar D. (1992). Regulation of the cAMP signal transduction pathway by protein kinase C in rat submandibular cells. Eur J Physiol 421:82-89.

11.

Gericke M. , Droogmans G. , Nilius B. (1993). Thapsigargin discharges intracellular calcium stores and induces transmembrane currents in human endothelial cells. Eur J Physiol 422:552-557.

12.

Hall RL , Miller RJ , Peatfield AC , Richardson PS , Williams L. , Lampert I. (1980). A colorimetric assay for mucous glycoproteins using Alcian blue. Biochem Soc Trans 8:72.

13.

Kolesnick RN , Musacchio I. , Thaw C. , Gershengorn MC (1984). Arachidonic acid mobilizes calcium and stimulates prolactin secretion from GH3 cells. Am J Physiol 246:E458-E462.

14.

Piomelli D. , Wang JKT , Sirha TS , Nairn AC , Czernik AJ , Greengard P. (1989). Inhibition of Ca2+/calmodulindependent protein kinase II by arachidonic acid and its metabolites. Proc Natl Acad Sci USA 86:8550-8554.

15.

Plomp PJAM , van Roermund CWT , Groen AK , Meijer AJ , Tager JM (1985). Mechanism of the stimulation of respiration by fatty acids in rat liver. FEBS Lett 193:243-246. Randriamampita C. , Trautmann A. (1990). Arachidonic acid activates Ca2+ extrusion in macrophages . J Biol Chem 265:18059-18062.

16.

Rotman EI , Brostrom MA , Brostrom CO (1992). Inhibition of protein synthesis in intact mammalian cells by arachidonic acid. Biochem J 282:487-494.

17.

Sambrook J. , Fritsch EF , Maniatis T. (1989). Molecular cloning. A laboratory manual. 2nd ed. Cold Spring Harbor: Cold Spring Harbor Laboratory Press, Chapter 7.

18.

Strehler BL (1974). Adenosine-5'-triphosphate and creatine phosphate. Determination with luciferase. In: Methods of enzymatic analysis. Vol. 4. Bergmeyer HU , editor. New York: Academic Press, pp. 2112-2126.

19.

Tanaka Y. , Tashjian AH Jr (1993). Functional identification and quantitation of three intracellular calcium pools in GH4C1 cells: evidence that the caffeine-responsive pool is coupled to a thapsigargin-resistant, ATP-dependent process. Biochemistry 32:12062-12073.

20.

Wolf BA , Turk J. , Sherman WR , McDaniel ML (1986). Intracellular Ca2+ mobilization by arachidonic acid. Comparison with myo-inositol 1,4,5-trisphosphate in isolated pancreatic islets. J Biol Chem 261:3501-3511.