K+ currents of encephalitogenic memory T cells decrease with encephalitogenicity while interleukin-2 (IL-2) receptor expression remains stable during IL-2 dependent cell expansion

Previous studies revealed a linkage between increased K+ current and lymphocyte activation upon non-specific stimulation with mitogenic lectins and antibodies. So far no information is available about the behaviour of K+ currents in specifically autoantigen-stimulated lymphocytes. Therefore, we have investigated K+ currents in encephalitogenic T line cells, specifically stimulated with myelin basic protein, using the whole-cell patch-clamp technique. In parallel, the T cell activation marker interleukin-2 (IL-2) receptor was measured quantitatively by flow cytometry. Outward currents were observed in response to depolarizing voltage steps from a holding potential of -80 mV. The peak current density increased with more positive membrane potentials, where the current threshold was about -40mV and the maximum conductance was 1.22 nS/pF. This current was characterized by a fast activation and a fast inactivation with half maximal inactivation at -67 mV. The sensitivity of the peak current to Ki channel blocking agents was as follows: 4-aminopyridine (4-AP) had a half blocking concentration of 0.4mM and a maximal block of 83.7% at 10 mM 4-AP, tetraethylammonium caused a block of 6% at 0.1mM, 15% at 1 mM and 40% at 10mM, charybdotoxin blocked 90% at 100 nM, whereas iberiotoxin had no effect (all values at a damped membrane potential of +30 mV). The encephalitogenic T cells used in our study reach their highest encephalitogenic potency on day 3 to 4 after the onset of restimulation. Furthermore, K+ currents were measured during the whole course of an in vitro restimulation cycle. The peak currents normalized to cell capacitance reached their maximum on day 2 (326+/- 52.8pA/pF, n=4) and decreased thereafter as follows: day 3: 139.7+/-7.87pA/pF (n = 27), day 4: 85.4 +/- 8.95 pA/pF (n = 28) and day 5. 40.9 +/- 7.45 pA/pF (n = 17). The activation and inactivation characteristics of the current and its responses to selective blockers were similar at all days after restimulation. In contrast to the K+ current, IL-2 receptor expression was maintained on >95% of cells until day 6 after restimulation.