T lymphocytes express muscarinic acetylcholine receptors (mAChRs) involved in regulating their proliferation, differentiation and cytokine release. Activation of M1, M3 or M5 mAChRs increases the intracellular Ca2 + concentration ([Ca2 +]i) through inositol-1,4,5-phosphate (IP3)-mediated Ca2 + release from endoplasmic reticulum Ca2 + stores. In addition, T lymphocytes express Ca2 +-release activated Ca2 + (CRAC) channels to induce Ca2 + influx and to regulate diverse immune functions. Our aim in the present study was to assess the role of CRAC channels during mAChR activation in the Ca2 +-dependent transduction that contributes to the regulation of T cell function.Main methods:
Changes in [Ca2 +]i following mAChR activation on human leukemic T cells, CCRF-CEM (CEM), were monitored using fura-2, based on the ratio of 510 nm fluorescences elicited by excitation at 340 nm and 380 nm (R340/380).Key findings:
We demonstrate that CEM cells express mainly M3 and M5 mAChRs, but little the M1 subtype, and that oxotremorine-M (Oxo-M), an mAChR agonist, induces an initial transient increase in [Ca2 +]i followed by repetitive [Ca2 +]i oscillations. Removing extracellular Ca2 + or pharmacological blockade of CRAC channels abolished the [Ca2 +]i oscillations without affecting the initial [Ca2 +]i transient induced by Oxo-M. Moreover, CRAC channel blockade also suppressed Oxo-M-induced c-fos and interleukin-2 expression.Significance:
These results suggest that upon M3 or M5 mAChR activation, IP3-mediated Ca2 + release induces extracellular Ca2 + influx through CRAC channels, which generates repetitive [Ca2 +]i oscillations and, in turn, enhances c-fos gene expression in T lymphocytes.