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

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