Thiopental Elevates Steady-State Levels of Intracellular Ca2+ and Zn2+ in Rat Thymic Lymphocytes
Toxicity test of thiopental on rat thymic lymphocytes
AbstractThiopental is an ultra-short-acting barbiturate and has been used commonly in the induction phase of general anesthesia. However, the toxic effect of thiopental is not completely clear. The effect of thiopental on intracellular Ca2+ ([Ca2+]i) levels was investigated in non-excitable cells. Experiments were carried out using a flow-cytometric technique, rat thymic lymphocytes (as non-excitable cells), and appropriate fluorescent probes. Treatment of cells with 300 µM thiopental increased Fluo-3 fluorescence intensity, indicating elevation of [Ca2+]i. This increase was partially attenuated by a chelator of intracellular Zn2+. Thus, thiopental elevated both [Ca2+]i and intracellular Zn2+ ([Zn2+]i) levels. Under intracellular Zn2+-free conditions, 100–300 µM thiopental was still able to induce a statistically significant increase in [Ca2+]i, whereas removal of extracellular Ca2+ greatly reduced the increase in [Ca2+]i induced by this dose of thiopental. Therefore, the thiopental-induced increase in [Ca2+]i was mainly due to an increased influx of Ca2+. Treatment of cells with 300 µM thiopental increased FluoZin-3 fluorescence intensity, indicating the presence of [Zn2+]i, both in the presence and absence of extracellular Zn2+. The thiopental-induced elevation of [Zn2+]i was due to an increase in both influx of Zn2+ and intracellular Zn2+ release. Concanavalin A (10 µg/mL) augmented Fluo-3 fluorescence in the presence of an intracellular Zn2+ chelator. The combination of concanavalin A and 100–300 µM thiopental synergistically increased [Ca2+]i. Results suggest that thiopental increases [Ca2+]i in both quiescent and activated lymphocytes, possibly resulting in modulation of immune system function.
 Yamakage, M., Hirshman, C., Croxton, T., 1995. Inhibitory effects of thiopental, ketamine, and propofol on voltage-dependent calcium2+ channels in porcine tracheal smooth muscle cells. Anesthesiology: The Journal of the American Society of Anesthesiologists 83, 1274–1282.
 Zhan, R.Z., Fujiwara, N., Endoh, H., Yamakura, T., Taga, K., Fukuda, S., Shimoji, K., 1998. Thiopental inhibits increases in [Ca2+]i induced by membrane depolarization, NMDA receptor activation, and ischemia in rat hippocampal and cortical slices. Anesthesiology: The Journal of the American Society of Anesthesiologists 89, 56–466.
 Kimura, M., Shibukawa, Y., Momose, Y., Sugaya, M., Yamamura, S., Suzuki, T., Hatakeyama, N., Yamazaki, M., 2007. Effects of thiopental on Ca2+ currents and intracellular Ca2+ transient in single atrial cells from guinea pig. Pharmacology 80, 33–39.
 Lecharny, J.B., Salord, F., Henzel, D., Desmonts, J.M., Mantz, J., 1995. Effects of thiopental, halothane and isoflurane on the calcium-dependent and-independent release of GABA from striatal synaptosomes in the rat. Brain Research 670, 308–312.
 Miao, N., Nagao, K., Lynch, C., 1998. Thiopental and methohexital depress Ca2+ entry into and glutamate release from cultured neurons. Anesthesiology: The Journal of the American Society of Anesthesiologists 88, 1643–1653.
 Lewis, R.S., 2001. Calcium signaling mechanisms in T lymphocytes. Annual Review of Immunology 19, 497–521.
 Feske, S., 2007. Calcium signalling in lymphocyte activation and disease. Nature Reviews Immunology 7, 690–702.
 Hirano, T., Murakami, M., Fukada, T., Nishida, K., Yamasaki, S., Suzuki, T., 2008. Roles of zinc and zinc signaling in immunity: zinc as an intracellular signaling molecule. Advances in Immunology 97, 149–176.
 Hasan, R., Rink, L., Haase, H., 2013. Zinc signals in neutrophil granulocytes are required for the formation of neutrophil extracellular traps. Innate Immunity 19, 253–264.
 Jiang, S., Chow, S.C., Nicotera, P., Orrenius, S., 1994. Intracellular Ca2+
signals activate apoptosis in thymocytes: studies using the Ca2+-ATPase
inhibitor thapsigargin. Experimental Cell Research 212, 84–92.
 Provinciali, M., Di Stefano, G., Fabris, N., 1995. Dose-dependent opposite
effect of zinc on apoptosis in mouse thymocytes. International Journal of
Immunopharmacology 17, 735–744.
 Telford, W.G., Fraker, P.J., 1995. Preferential induction of apoptosis in
mouse CD4+ CD8+ αβTCRIoCD3εIo thymocytes by zinc. Journal of Cellular
Physiology 164, 259–270.
 McConkey, D.J., Orrenius, S., 1997. The role of calcium in the regulation of
apoptosis. Biochemical and Biophysical Research Communications 239,
 Keel, M., Mica, L., Stover, J., Stocker, R., Trentz, O., & Härter, L., 2005.
Thiopental-induced apoptosis in lymphocytes is independent of CD95
activation. Anesthesiology: The Journal of the American Society of
Anesthesiologists 103, 576–584.
 Roesslein, M., Schibilsky, D., Muller, L., Goebel, U., Schwer, C., Humar, M.,
Schmidt, R., Geiger, K.K., Pahl, H.L., Pannen, B.H., & Loop, T., 2008.
Thiopental protects human T lymphocytes from apoptosis in vitro via the
expression of heat shock protein 70. Journal of Pharmacology and
Experimental Therapeutics 325, 217–225.
 Gee, K.R., Zhou, Z.L., Ton-That, D., Sensi, S.L., Weiss, J.H., 2002.
Measuring zinc in living cells: A new generation of sensitive and selective
fluorescent probes. Cell Calcium 31, 245–251.
 Kao, J.P., Harootunian, A.T., Tsien, R.Y., 1989. Photochemically generated
cytosolic calcium pulses and their detection by fluo-3. Journal of Biological
Chemistry 264, 8179–8184.
 Sakanashi, Y., Oyama, T.M., Matsuo, Y., Oyama, T.B., Nishimura, Y., Ishida,
S., Okano, Y., Oyama, Y., 2009. Zn2+, derived from cell preparation, partly
attenuates Ca2+-dependent cell death induced by A23187, calcium
ionophore, in rat thymocytes. Toxicology in Vitro 23, 338–345.
 Mousa, W. F., Enoki, T., Fukuda, K., 2000. Thiopental induces contraction
of rat aortic smooth muscle through Ca2+ release from the sarcoplasmic
reticulum. Anesthesia & Analgesia 91, 62–67.
 Henkel, C.C., Asbun, J., Ceballos, G., Castillo, M.D.C., Castillo, E.F., 2001.
Relationship between extra and intracellular sources of calcium and the
contractile effect of thiopental in rat aorta. Canadian Journal of Physiology
and Pharmacology 79, 407–414.
 O'Flynn, K., Linch, D.C., Tatham, P.E.R., 1984. The effect of mitogenic
lectins and monoclonal antibodies on intracellular free calcium
concentration in human T-lymphocytes. Biochemical Journal 219, 661–666.
 Delogu, G., Antonucci, A., Moretti, S., Marandola, M., Tellan, G., Signore,
M., Famularo, G., 2004. Oxidative stress and mitochondrial glutathione in
human lymphocytes exposed to clinically relevant anesthetic drug
concentrations. Journal of Clinical Anesthesia 16, 189–194.
 Dwyer, J.M., Johnson, C., 1981. The use of concanavalin A to study the
immunoregulation of human T cells. Clinical and Experimental Immunology
 Maret, W., 1994. Oxidative metal release from metallothionein via zinc-
thiol/disulfide interchange. Proceedings of the National Academy of
Sciences 91, 237–241.
 Taeger, K., Lueg, J., Finsterer, U., Roedig, G., Weninger, E., Peter, K., 1986.
Thiopental levels in the plasma during induction of anesthesia. Anasthesie
Intensivtherapie Notfallmedizin 21, 169–174.
Copyright (c) 2020 Journal of Tropical Pharmacy and Chemistry
This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License.