Abstract
The effect of changes of pHi on Cai were studied using fluorescent dyes in cells of the cultured smooth muscle-like line, BC3H-1. Resting Cai in these cells was 182 ± 12 nM (n = 74) at pHo of 7.4. Upon exposure to NH4Cl, which rapidly alkalinized cells, a transient increase of Cai to 349 ± 55 nM (n = 29) was observed. The peak of the transient occurred within 30 s of exposure to NH4Cl and returned to baseline within 1 minute. Two other procedures which resulted in rapid cellular alkalinization also caused a transient rise in Cai: exposure to and then removal of CO2 (Cai increased from 182 ± 22 to 248 ± 28 nM; n = 8); and exposure to and then removal of Na propionate (Cai increased from 242 ± 32 to 456 ± 71 nM; n = 9). The NH4Cl-induced Cai transient was eliminated by exposure to 0.2 mM TMB8 and to Ca-free solutions, but not by exposure to 0.5 mM LaCl3. Sustained changes of pHi can be induced by varying pHo. When pHo was lowered to 6.9, Cai fell by 49 ± 11 nM but increased by 203 ± 51 nM (n = 6) when pHo was raised to 7.9. These data indicate that rapid alkalinization of BC3H-1 cells results in a rapid transient rise of Cai. This transient is most likely due to the release of Ca from intracellular stores but may also involve an increase of Ca influx. Steady state values of Cai are positively correlated with steady state pHi. These data may have implications for the contractile state of smooth muscle during periods of acid/base disturbances and relate to the role of elevated pHi in cells from hypertensive animals.
Similar content being viewed by others
References
Bers DM, Ellis D: Intracellular calcium and sodium activity in sheep heart Purkinje fibres. Effect of changes of external sodium and intracellular pH. Pflüg Arch 393: 171–178, 1982
Abercrombie RA, Roos A: The intracellular pH of frog skeletal muscle: its regulation in hypertonic solutions. J Physiol 345: 189–204, 1983
Hendey B, Mamrack MD, Putnam RW: Thrombin induces a calcium transient that mediates an activation of the Na+/H+ exchanger in human fibroblasts. J Biol Chem. In press
Villereal ML: Regulation of Na+/H+ exchange: role of Ca2+. In: S Grinstein (ed.) Na/H Exchange. CRC Press, Inc, Boca Raton, Florida, 1988, pp 243–253
Mahnensmith RL, Aronson PS: The plasma membrane sodium-hydrogen exchanger and its role in physiological and pathophysiological processes. Circ Res 56: 773–788, 1985
Nakamaru Y, Schwartz A: The influence of hydrogen ion concentration on calcium binding and release by skeletal muscle sarcoplasmic reticulum. J Gen Physiol 59: 22–32, 1972
Siskind MS, McCoy CE, Chobanian A, Schwartz JH: Regulation of intracellular calcium by cell pH in vascular smooth muscle cells. Am J Physiol 256 (Cell Physiol 25): C234-C240, 1989
Shepherd JT: Circulation to skeletal muscle. In: JT Shepherd and FM Abboud (eds.) Handbook of Physiology, sec 11, vol III. Peripheral Circulation and Organ Blood Flow, Part 1. American Physiological Society, Washington, D.C., 1983, pp 319–370
Danthuluri NR, Deth RC: Effects of intracellular alkalinization on resting and agonist-induced vascular tone. Am J Physiol 256 (Heart Circ Physiol 25): H867-H875, 1989
Corry DB, Yanagawa N, Tuck ML: Inhibition of cation transport pathways alters cytosolic calcium (Cai) in vascular smooth muscle cells (VSMC) (Abstr). Amer Soc Nephrol. Abstracts 21st Annual Meeting: 176A, 1988
Nguyen-Hoang H, Bamosa AO, Spurway NC: Effects of variations of intracellular pH, induced by exposure to weak acids or bases, on vascular smooth muscle contractility (Abstr). Proc Int Union Physiol Sci 17: 398, 1989
Aviv A: The link between cytosolic Ca2+ and the Na+-H+ antiport: a unifying factor for essential hypertension. J Hypertension 6: 685–691, 1988
Muslin AJ, Berk BC, Alexander RW: Increased sodium-hydrogen exchange and intracellular pH in spontaneously hypertensive rat vascular smooth muscle cells (Abstr). Clin Res 35: 445A, 1987
Schubert D, Harris AJ, Devine CE, Heinemann S: Characterization of an unique muscle cell line. J Cell Biol 61: 398–413, 1974
Putnam RW: Down regulation of pH-regulating transport systems in BC3H-1 cells. Annals NY Acad Sci 574: 354–369, 1989
Putnam RW, Grubbs RD: Steady state pH1, buffering power and effect of CO2 in a smooth muscle-like cell line. Am J Physiol 258 (Cell Physiol 27): C461-C469, 1990
Cobbold PH, Rink TJ: Fluorescence and bioluminescence measurement of cytoplasmic-free calcium. Biochem 1248: 313–328, 1987
Grynkiewicz G, Poenie M, Tsien RY: A new generation of Ca2+ indicators with greatly improved fluorescence properties. J Biol Chem 260: 3440–3450, 1985
Boron WF, De Weer P: Intracellular pH transients in squid giant axons caused by CO2, NH3, and metabolic inhibitors. J Gen Physiol 67: 91–112, 1976
Putnam RW: pH regulatory transport systems in a smooth muscle-like cell line. Am J Physiol 258 (Cell Physiol 27): C470-C479, 1990
Mix LL, Dinerstein RJ, Villereal ML: Mitogens and melittin stimulate an increase in intracellular free calcium concentration in human fibroblasts. Biochem Biophys Res Comm 119: 69–75, 1984
Goldenhofen K, Hermstein N: Differentiation of calcium activation mechanisms in vascular smooth muscle by selective suppression with verapamil and D600. Blood Vessels 12: 21–37, 1975
Ighoroje AD, Spurway NC: How does vascular muscle in the isolated rabbit ear adapt its tone after alkaline or acid loads? (Abstr) J Physiol 357: 105P, 1984
Izzard AS, Heagerty AM: The measurement of internal pH in resistance arterioles: evidence that intracellular pH is more alkaline in SHR than WKY animals. J Hypertension 7: 173–180, 1989
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Putnam, R.W., Douglas, P.B. Effect of changes of pHi on intracellular calcium in a smooth muscle-like cell line. Mol Cell Biochem 99, 89–95 (1990). https://doi.org/10.1007/BF00230338
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF00230338