The avascular nature of articular cartilage means that chondrocytes are dependent on the diffusion of O₂ from the synovial fluid and subchondral bone. As a result, cartilage has a relatively low ambient O₂ tension estimated at between 5% and 7%. In addition, synovial O₂ tensions can fall to very low levels in inflammatory conditions, which can further lower O₂ levels within the matrix. These low O₂ tensions can affect ion homeostasis. For example, they alter pH homeostasis, with intracellular acidification associated with inhibition of Na⁺-H⁺ exchange (Milner et al. 2005). Intracellular Ca²⁺ also represents an important cell parameter, shown to be linked to pH levels (Fairfax et al. 2003). In the present study therefore, we investigated the effect of O₂ on Ca²⁺ homeostasis. Cartilage slices were obtained postmortem from bovine metacarpophalangeal joints. Chondrocytes were isolated overnight by collagenase digestion at either 20% (termed normoxia) or 1% O₂ (termed hypoxia), and maintained at these levels during all subsequent procedures. Intracellular Ca²⁺ ([Ca²⁺]_i) was then determined fluorimetrically using fura-2 (5 μM; EM 510nm; EX 340nm/380nm). Ca²⁺ levels are given as the 340nm/380nm signal ratio (R) (Sanchez & Wilkins, 2004). Ca²⁺ homeostasis was shown to depend on Ca²⁺ entry via the plasma membrane, thapsigargin-sensitive stores and Ca²⁺ efflux mediated predominantly by the Na⁺-Ca²⁺ exchanger. When cells isolated overnight at 1% (hypoxia) were compared to those isolated at 20% O₂, it was found that [Ca²⁺]_i was elevated, with R typically rising from 1.27 to 1.51. By contrast, shorter term hypoxia (3 hours) had no effect on Ca²⁺ homeostasis, and nor was there a significant difference between cells isolated at 20% and 5% O₂. At 1% O₂, exposure of chondrocytes to either Co²⁺ (100 μM) or antimycin A (5 μM) caused a reduction in [Ca²⁺]_i. For example, with Co²⁺, R fell from 1.59 to 1.36. Hypoxia has been shown to decrease levels of reactive oxygen species (ROS) in chondrocytes, whilst exposure to either Co²⁺ or antimycin A increases them (see accompanying poster). These findings suggest that Ca²⁺ homeostasis in articular chondrocytes is affected by low O₂ tension, with the putative signal being a reduction in ROS. We hypothesise that high levels of ROS mediates a reduction in [Ca²⁺]_i via stimulation of the Na⁺-Ca²⁺ exchange.