Chondrocytes in articular cartilage utilize mechanical signals from their environment to regulate their metabolic activity. However, the sequence of events involved in the transduction of mechanical signals to a biochemical signal is not fully understood. It has been proposed that an increase in the concentration of intracellular calcium ion ([Ca²⁺]ᵢ) is one of the earliest events in the process of cellular mechanical signal transduction. With use of fluorescent confocal microscopy, [Ca²⁺]ᵢ was monitored in isolated articular chondrocytes subjected to controlled deformation with the edge of a glass micropipette. Mechanical stimulation resulted in an immediate and transient increase in [Ca²⁺]ᵢ. The initiation of Ca²⁺ waves was abolished by removing Ca²⁺ from the extracellular media and was significantly inhibited by the presence of gadolinium ion (10 μM) or amiloride (1 mM), which have previously been reported to block mechanosensitive ion channels. Inhibitors of intracellular Ca²⁺ release (dantrolene and 8-diethylaminooctyl 3,4,5-trimethoxybenzoate hydrochloride) or cytoskeletal disrupting agents (cytochalasin D and colchicine) had no significant effect on the characteristics of the Ca²⁺ waves. These findings suggest that a possible mechanism of Ca²⁺ mobilization in this case is a self-reinforcing influx of Ca²⁺ from the extracellular media, initiated by a Ca²⁺-permeable mechanosensitive ion channel. Our results indicate that a transient increase in intracellular Ca²⁺ concentration may be one of the earliest events involved in the response of chondrocytes to mechanical stress and support the hypothesis that deformation-induced Ca²⁺ waves are initiated through mechanosensitive ion channels.