Role of ryanodine receptor channels in Ca²⁺ oscillations of porcine tracheal smooth muscle

Acetylcholine (ACh) induces repetitive, propagating intracellular Ca²⁺ concentration ([Ca²⁺](i)) oscillations in porcine tracheal smooth muscle (TSM) cells. Using real-time confocal microscopy, we examined the role of sarcoplasmic reticulum (SR) Ca²⁺ release through inositol 1,4,5- trisphosphate (IP₃) receptor and ryanodine receptor (RyR) channels in ACh- induced [Ca²⁺](i) oscillations. In β-escin permeabilized TSM cells, exposure to ACh in the presence of GTP also resulted in [Ca²⁺](i) oscillations. [Ca²⁺](i) oscillations could not be initiated by IP₃ alone; however, an elevation of [Ca²⁺](i) was observed. During ongoing [Ca²⁺](i) oscillations, exposure to heparin, an IP₃ receptor antagonist, caused a slowing of oscillation frequency but not complete inhibition. In contrast, ruthenium red, a RyR antagonist, completely abolished ACh-induced [Ca²⁺](i) oscillations. Reverse transcriptase-polymerase chain reaction of TSM mRNA demonstrated the expression of RyR-2 and RyR-3 isoforms of the RyR. These results indicate that SR Ca²⁺ release through RyR channels is critical for ACh-induced [Ca²⁺](i) oscillations in porcine TSM cells.