Ryanodine receptors (RyR) have been shown to be involved in the Ca²⁺ response in airway myocytes from various species (Kannan et al. 1997). The aim of this study was to develop a theoretical model of [Ca²⁺]_i variations upon RyR stimulation and to compare its predictions with experimental observations in airway myocytes.

Experimental studies were performed on myocytes freshly isolated from rat trachea. Rats were humanely killed according to national guidelines. [Ca²⁺]_i responses were measured by microspectrofluorimetry using the Ca²⁺-sensitive fluorescent dye indo-1. Statistical comparisons were done by paired or unpaired Student’s t tests as appropriate, and considered as significant when P < 0.05. Values are means ± S.E.M.

The mathematical model describing the mechanism of Ca²⁺ handling upon RyR stimulation included the sarcoplasmic reticulum (SR) and also cytosolic proteins and mitochondria as additional Ca²⁺ buffers. Our model predicts that after Ca²⁺ release from the SR Ca²⁺ is first sequestrated into mitochondria and binds to protein fast binding sites. Ca²⁺ is then shifted to the protein slow binding sites, and finally pumped back into the SR. RyR stimulation by 5 mM caffeine (CAF) for 30 s induced a first transient peak followed by a progressive decay to a plateau phase. When CAF exposure was stopped during the decay phase, [Ca²⁺]_i decay was increased and returned to baseline, indicating an immediate washout of CAF and closure of RyR. Stimulation by CAF for 1 s induced a Ca²⁺ peak (mean value 714 ± 120.8 nM, n = 8) followed by a quick decay, the RyR being closed. A second stimulation 15 s after the first one induced a second Ca²⁺ peak (amplitude 144 ± 93 nM). The baseline value after 30 s exposure to 10 mM cyclopiazonic acid (CPA), a reversible inhibitor of the sarco-endoplasmic Ca²⁺-ATPase (SERCA) was not significantly modified (127 ± 6.7 nM; n = 8) vs. control (118 ± 6 nM; n = 8). When cells were stimulated by CAF 30 s after the beginning of exposure to CPA, the first peak was not modified vs. control (713 ± 124.4 nM), but the Ca²⁺ response to the second stimulation was abolished (25 ± 8.2 nM), indicating that the SERCA was blocked by CPA. However, the exponential decay in [Ca²⁺]_i following Ca²⁺ increase was similar in the absence and in the presence of CPA.

These experimental results indicate in accordance with the predictions of our theoretical model that though Ca²⁺ pumping back by SERCA is active after Ca²⁺ release from SR upon RyR stimulation, it is not primarily involved in [Ca²⁺]_i decrease that may be due to other buffering processes.

All procedures accord with current National guidelines.