Cellular Pharmacology of Bupivacaine in Lung Fibroblasts: Role of Intracellular Calcium Signalling and Chelation

Bupivacaine, a commonly used aminoamide local anaesthetic, exerts cellular effects beyond anaesthesia. However, its effects on calcium (Ca²⁺) signalling and the physiological responses of lung fibroblasts remain insufficiently characterised. This study aimed to determine whether bupivacaine disrupts intracellular Ca²⁺ homeostasis and induces cytotoxicity in IMR-90 human foetal lung fibroblasts via Ca²⁺-dependent mechanisms. Cell viability was assessed using the CCK-8 assay, and intracellular Ca²⁺ levels ([Ca²⁺]ᵢ) were monitored in fura-2-loaded cells. Pharmacological inhibitors were used to dissect the signalling pathways involved. Bupivacaine (20-60 μM) caused a concentration-dependent rise in [Ca²⁺]ᵢ and a corresponding decline in cell viability. These effects were significantly attenuated by pre-treatment with the Ca²⁺ chelator BAPTA-AM, indicating a Ca²⁺-dependent cytotoxic mechanism. Mechanistically, bupivacaine induced Ca²⁺ influx through store-operated Ca²⁺ entry (SOCE), involving protein kinase C (PKC), and triggered Ca²⁺ release from the endoplasmic reticulum (ER) via a phospholipase C (PLC)-dependent pathway. Mn²⁺ quenching assays confirmed SOCE as the primary route of Ca²⁺ entry. Bupivacaine disrupts Ca²⁺ signalling through both extracellular influx and intracellular release, contributing to pulmonary cytotoxicity. These findings underscore the relevance of Ca²⁺ homeostasis in anaesthetic-related lung injury and suggest that Ca²⁺ chelation may be a potential protective strategy.