Figure 4: (A) Electrophysiological recordings of whole-cell K+ currents from purified human T cells, in the presence of Sub P (10 nM) and various Kv1.3 channel blockers. Typical whole-cell K+ currents were obtained by stepping the membrane from a holding potential of −80 mV to +60 mV (500 ms) in 10-mV increments. The effect saturated after ∼8 min. Quinidine (10 μM) and ChTX (10 nM) inhibited the maximal amplitude of the K+ currents by 90 and 46%, respectively. Apamin, as a negative control, had no inhibitory effect on the K+ currents. The peak amplitudes of the control (□, n = 13), apamin (5 nM; ▪, n = 8), ChTX (10 nM; ○, n = 11), Sub P (10 nM; •, n = 10), MgTx (10 nM; ▵, n = 5), and quinidine (10 μM; ▴, n = 9) were plotted against the voltage step and fitted using the Boltzmann distribution. Statistical difference is expressed by SE. (B) Physical association between Kv1.3 channels and β1 integrins in human T cells. Top, The extract of membrane fraction prepared from purified normal human T cells was subjected to parallel immunoprecipitations (IP) with either anti–β1 integrin (CD29), anti-Kv1.3, or preimmune (P.I.) negative control antibodies. Immunoblotting of these precipitated immune complexes, as well as of a portion of the lysate (Input, 10%), was carried out using an anti–β1 integrin antibody. The β1 integrin is detected in membrane extracts precipitated with either the anti–β1 integrin or the anti-Kv1.3 antibody, as well as in the lysate. Bottom, Similar immunoprecipitation as in the top panel, and blotting with the anti-Kv1.3 antibody. The Kv1.3 protein is detected in membrane extracts precipitated with either the anti-Kv1.3 or the anti–β1 integrin antibody. Two different gels from two independent experiments are shown to illustrate the immunoprecipitation of Kv1.3 either by anti-Kv1.3 antibodies (left) or by anti-CD29 antibodies (right).

Mentions: Representative whole-cell currents are shown in Fig. 4 A. The membrane of untreated human T cells was stepped from a holding potential of −80 mV to +60 mV, in 10-mV increments. After application of Sub P (10 nM), the maximal K+ currents were reduced by 39%, from 241 ± 14 pA (n = 10) to 148 ± 24 pA (n = 10). A 100-fold higher Sub P concentration (1 μM) reduced the current by ∼45%, to 109 ± 11 pA (n = 10).

Levite M, Cahalon L, Peretz A, Hershkoviz R, Sobko A, Ariel A, Desai R, Attali B, Lider O - J. Exp. Med. (2000)

Bottom Line: In support of this notion, we found that the proadhesive effects of the chemokine macrophage-inflammatory protein 1beta, the neuropeptide calcitonin gene-related peptide (CGRP), as well as elevated [K(+)](o) levels, are blocked by specific Kv1.3 channel blockers, and that the unique physiological ability of substance P to inhibit T cell adhesion correlates with Kv1.3 inhibition.This study shows that T cells can be activated and driven to integrin function by a pathway that does not involve any of its specific receptors (i.e., by elevated [K(+)](o)).In addition, our results suggest that undesired T cell integrin function in a series of pathological conditions can be arrested by molecules that block the Kv1.3 channels.

Affiliation: Department of Immunology, The Weizmann Institute of Science, Rehovot 76100, Israel. mia.levite@weizmann.ac.il

Abstract: Elevated extracellular K(+) ([K(+)](o)), in the absence of "classical" immunological stimulatory signals, was found to itself be a sufficient stimulus to activate T cell beta1 integrin moieties, and to induce integrin-mediated adhesion and migration. Gating of T cell voltage-gated K(+) channels (Kv1.3) appears to be the crucial "decision-making" step, through which various physiological factors, including elevated [K(+)](o) levels, affect the T cell beta1 integrin function: opening of the channel leads to function, whereas its blockage prevents it. In support of this notion, we found that the proadhesive effects of the chemokine macrophage-inflammatory protein 1beta, the neuropeptide calcitonin gene-related peptide (CGRP), as well as elevated [K(+)](o) levels, are blocked by specific Kv1.3 channel blockers, and that the unique physiological ability of substance P to inhibit T cell adhesion correlates with Kv1.3 inhibition. Interestingly, the Kv1.3 channels and the beta1 integrins coimmunoprecipitate, suggesting that their physical association underlies their functional cooperation on the T cell surface. This study shows that T cells can be activated and driven to integrin function by a pathway that does not involve any of its specific receptors (i.e., by elevated [K(+)](o)). In addition, our results suggest that undesired T cell integrin function in a series of pathological conditions can be arrested by molecules that block the Kv1.3 channels.