The Role of Ion Channels in Differentiating Chondrocytes

Abstract

Cartilage is an important load-bearing component of the skeleton of vertebrates. This tissue has similar composition to other members of the connective tissue family: major constituents are cells, the chondrocytes, and the surrounding extracellular matrix; however, it is also unique being avascular. Chondrocytes are non-excitable cells and little is known about their plasma membrane ion channels. The aim of our study was to identify ion channels and establish their roles in differentiating chondrocytes.Our in vitro chondrogenesis model system is a high density mesenchymal cell culture, in which chondroprogenitor cells are isolated from limb buds of chicken embryos. Using whole-cell patch-clamp we have detected voltage-dependent ionic currents in these differentiating cells, whose amplitude depended on the time elapsed since isolation. An outward current was present in chondrocytes within 1-2 days of isolation, while an inward current gradually replaced it about 2 days after isolation. Using ion substitution experiments we identified the channels responsible for the currents as voltage-gated K+ and Na+ channels, respectively.The average amplitude of the Na+ current in cells during days 3-4 following isolation was −294 +/− 22 pA at 0 mV. The current inactivated with a time constant of tau = 0.59 +/− 0.04 ms. The voltage-dependence of steady state activation and inactivation were also determined yielding V1/2 values of −38 and −72 mV. Tetrodotoxin reversibly blocked the current with a Kd = 12 nM. The results of planned molecular biological experiments combined with our biophysical and pharmacological data will be used to identify the channel. The characterization of the K+ channel is presently underway.In many cell types changes in ion channel expression are associated with differentiation, thus our long-term aim is the clarification of the role of these channels in chondrogenesis and its potential clinical consequences.