Activation of nicotinic acetylcholine receptors expressed in quail fibroblasts: effects on intracellular calcium

1 The aim of these experiments was to determine the ability of the muscle‐type nicotinic acetylcholine receptor (AChR) stably expressed in quail fibroblasts (QF18 cells) to elevate intracellular calcium ([Ca2+]i) upon activation. Ratiometric confocal microscopy, with the calcium‐sensitive fluorescent dye Indo‐1 was used. 2 Application of the nicotinic agonist, suberyldicholine (SDC), to the transfected QF18 cells caused an increase in [Ca2+]i. Control [Ca2+]i levels in QF18 cells were found to be 164 ± 22 nM (mean ± s.e.mean; n = 40 cells) rising to 600 ± 81 nM on addition of SDC (10 μm; n=15 cells), whereas no increase in [Ca2+]i was seen in non‐transfected control QT6 fibroblasts (before: 128 ± 9 nM, n = 40; after: 113±13 nM, n= 15). 3 The increase in [Ca2+]i caused by application of SDC was dose‐dependent, with an EC50 value of 12.7 ± 5.9 μm (n=14). 4 The responses to SDC in QF18 cells were blocked by prior application of α‐bungarotoxin (200 nM), by the addition of Ca2+ (100 μm), by removal of Na+ ions from the extracellular solution, or by the voltage‐sensitive calcium channel blockers nifedipine and ω‐conotoxin, which act with IC50 values of 100 nM and 100 pM respectively. 5 We conclude that activation of the nicotinic AChRs leads to a Na+‐dependent depolarization and hence activation of endogenous voltage‐sensitive Ca2+ channels in the plasma membrane and an increase in [Ca2+]i. There is no significant entry of Ca2+ through the nicotinic receptor itself.