EFFECTS OF INTERMITTENT PRESSURE-INDUCED STRAIN ON THE ELECTROPHYSIOLOGY OF CULTURED HUMAN CHONDROCYTES - EVIDENCE FOR THE PRESENCE OF STRETCH-ACTIVATED MEMBRANE ION CHANNELS

1. Cyclical pressurization of cultured chondrocytes results in increas es in cyclic AMP and in the rate of proteoglycan synthesis. Intermitte nt increases in hydrostatic pressure are also associated with hyperpol arization of chondrocyte cell membranes and activation of Ca2+-depende nt K+-ion channels but the physiological basis for this response to me chanical stimulation is unclear. 2. Experiments have been undertaken t o better define the types of ion channels involved and to explore the possibility that the hyperpolarization response associated with cyclic al pressurization of chondrocytes follows activation of stretch-activa ted ion channels. 3. The mean membrane potential of chondrocytes in no n-confluent monolayer cell culture rose from -15.3 +/- 0.24 mV to -21. 1 +/- 0.28 mV (n = 60, P < 0.0001) after intermittent pressurization ( 0.33 Hz, 16 kPa, 20 min). 4. Strain gauge measurements showed that cyc lical pressurization was associated with strain on the base of the cul ture plate, The amplitude of the hyperpolarization response was propor tional to the microstrain to which cells were subjected. 5. Membrane h yperpolarization did not occur when chondrocytes were subjected to cyc lical pressurization in rigid glass culture dishes or plastic dishes p ositioned in the pressurization chamber so as to avoid bending of the base of the culture dish. 6. Indirect evidence that the hyperpolarizat ion response after intermittent pressure-induced strain was associated with stimulation of stretch-activated ion channels was obtained from experiments with gadolinium, amiloride and hexamethylene amiloride, ea ch of which abolished hyperpolarization. 7. Experiments with apamin, c harybdotoxin and iberiotoxin showed that the Ca2+-activated K+ channel s involved in the hyperpolarization response are apamin-sensitive, cha rybdotoxin- and iberiotoxin-resistant, low-conductance channels. 8. So matostatin and cadmium chloride, which block L-type calcium channels, abolished strain-induced chondrocyte hyperpolarization, EGTA, which ch elates extracellular Ca2+, reduced the response to 48% of control valu es, and thapsigargin, which raises intracellular Ca2+ by inhibition of Ca2+-ATPase in eudoplasmic reticulum, caused hyperpolarization indepe ndently with further hyperpolarization after pressure-induced strain, These data indicate that chondrocyte hyperpolarization was dependent o n intracellular Ca2+ concentrations. 9. Further work is required to de termine whether stretch-activated ion channels shown to be associated with chondrocyte hyperpolarization after cyclical pressure-induced str ain are also involved in the signal transduction process that leads to increases in proteoglycan synthesis.