Abstract
Several reports have documented that thapsigargin is a potent inhibitor of the SR Ca2+ ATPase isolated from cardiac or skeletal muscle. We have characterized the specificity of this agent in intact rat cardiac myocytes using cells maintained in the whole cell voltage clamp configuration. We have shown that thapsigargin decreases the magnitude of the Ca2+ transient and the twitch by about 80% while it slows the decay rate for these responses. These changes were not accompanied by any alterations in sarcolemmal currents or in the trigger Ca2+ generated by the inward calcium current. Taken together these results reveal that the action of thapsigargin is restricted to the SR Ca2+ ATPase in intact cardiac myocytes. Furthermore, it is demonstrated unambiguously that SR intracellular Ca2+ stores are an absolute requirement for the development of contractile tension in rat heart myocytes. It is shown that thapsigargin is a valuable probe to examine the importance of SR pools of Ca2+ and the role of the Ca2+ ATPase in intact myocytes as well as in genetically altered heart cells.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Campbell, A. M., Kessler, P. D., Sagara, Y., Inesi, G. and Farabrough, D. A. (1991) Nucleotide sequences of avian cardiac and brain SR/ER Ca2+-ATPases and functional comparisons with fast twitch Ca2+-ATPase.J. Biol. Chem. 266:16050–16055.
Channell, M. B., Cheng, H. and Lederer, W. J. (1994) Spatial non-uniformities in [Ca2+] i during excitation-contraction coupling in cardiac myocytes.Biophysical Journal 67:1956.
Cannell, M. B., Cheng, H. and Lederer, W. J. (1995) The control of calcium release in heart muscle.Science 268:1045–1049.
Chaudhari, N. and Beam, K. G. (1989) The dysgenesis mutation in mice leads to arrest of the genetic program for muscle differentiation.Dev. Biol. 133:457–467.
Christensen, S. B., Larsen, I. K. and Rasmussen, U. (1982) Thapsigargin and thapsigargicin, two histamine liberating sesquiterpine lactones fromThapsia gargancia.J. Org. Chem. 47:649–652.
Corin, S., Levitt, L. K., O'Mahoney, J., Joya, J. E., Hardeman, E. C. and Wade, R. (1995) Delineation of a slow twitch myofiber-specific transcriptional element usingin vivo somatic gene transfer.Proc. Natl. Acad. Sci. (USA) 92:6185–6189.
Huang, C.-F., Flucher, B. E., Schmidt, M. M., Stroud, S. K. and Schmidt, J. (1994) Depolarization-transcription signals in skeletal muscle use calcium flux through L channels, but bypass the sarcoplasmic reticulum.Neuron 13:167–177.
Inesi, G. and Sagara, Y. (1994) Specific inhibitors of intracellular Ca2+ transport ATPases.J. Membr. Biol. 141:1–6.
Jackson, T. R., Patterson, S. I., Thastrup, O. and Hanley, M. R. (1988) A novel tumour promotor, thapsigargin, transiently increases cytoplasmic free Ca2+ without generation of inositol phosphates in NG115-40L neuronal cells.Biochem. J. 253:81–86.
Kijima, Y., Ogunbunmi, E. and Fleischer, S. (1991) Drug action of thapsigargin on the Ca2+ pump protein of sarcoplasmic reticulum.J. Biol. Chem. 266:22912–22918.
Kirby, M. S., Sagara, Y., Gaa, S. T., Inesi, G., Lederer, W. J. and Rogers, T. B. (1992) Thapsigargin inhibits contraction and Ca2+ transient in cardiac cells by specific inhibition of the sarcoplasmic reticulum Ca2+ pump.J. Biol. Chem. 267:12545–12551.
Leinwald, L. A. and Leiden, J. M. (1991) Gene transfer into cardiac myocytes in vivo.Trends Cardiovasc. Med. 1:271–276.
Lytton, J., Westlin, M. and Hanley, M. R. (1991) Thapsigargin inhibits the sarcoplasmic or endoplasmic reticulum Ca-ATPase family of calcium pumps.J. Biol Chem. 266:17067–17071.
Rasmussen, U., Christensen, S. B. and Sandberg, F. (1978) Thapsigargin and thapsigargicin, two new histamine liberators fromThapsia garganica.Acta Phamaceut. Suec. 15:133–140.
Sagara, Y., Fernandez-Belda, F., De Meis, L. and Inesi, G. (1992) Characterization of the inhibition of intracellular Ca2+ transport ATPases by thapsigargin.J. Biol. Chem. 267:12606–12613.
Sagara, Y. and Inesi, G. (1991) Inhibition of the sarcoplasmic reticulum Ca2+ transport ATPase by thapsigargin at subnanomolar concentrations.J. Biol. Chem. 266:13503–13506.
Sagara, Y., Wade, J. B. and Inesi, G. (1992) A conformational mechanism for formation of a dead-end complex by sarcoplasmic reticulum ATPase with thapsigargin.J. Biol. Chem. 267:1286–1292.
Thastrup, O., Foder, B. and Scharff, O. (1987) The calcium mobilizing and tumour promoting agent, thapsigargin, elevates the platelet cytoplasmic free calcium concentration to a higher steady state level. A possible mechanism of action for the tumour promotion.Biochem. Biophys. Res. Comm. 142:654–660.
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Rogers, T.B., Inesi, G., Wade, R. et al. Use of thapsigargin to study Ca2+ homeostasis in cardiac cells. Biosci Rep 15, 341–349 (1995). https://doi.org/10.1007/BF01788366
Received:
Issue Date:
DOI: https://doi.org/10.1007/BF01788366