Intervertebral disc cells can be cultured in vitro. Several culturing systems in a mechanically active environment have been developed to study the relationship between mechanical stimulations and biochemical events. The aim of this study was to assess the phenotype of rabbit intervertebral disc cells from the anulus fibrosus (AF) region cultured on flexible substrate before and after application of cyclic tensile stretch (CTS) and to control culture conditions during application of CTS. CTS was applied with a pressure-operated instrument, inducing the deformation of flexible-bottomed culture plates (Flexercell) at 20% and 5% stretch, at a frequency of 1 Hz, during 30 minutes to 24 hours. A significant decrease in culture medium volume and temperature was observed (52% and 2.1 degrees C at 20% stretch and 24 hours' application of CTS). These phenomena were inhibited by adding culture medium around culture wells and by a culture medium temperature control system. Like AF cells cultured in plastic wells, AF cells cultured on flexible substrate expressed collagen type II, but collagen type I mRNA was not detected. In both culture conditions, neosynthesized proteoglycans had the same aggregating properties. CTS at 20% stretch during 12 hours did not induce cell detachment from the substrate and did not modify aggregating properties of neosynthesized proteoglycans; AF cells continued to express collagen type II but not collagen type I mRNA. In conclusion, the Flexercell system appears to be appropriate for studying, at the cellular level, the metabolic responses to CTS.