Transmembrane ion conductance in human B lymphocyte activation
Brent, L.H.; Butler, J.L.; Woods, W.T.; Bubien, J.K.
Journal of Immunology 145(8): 2381-2389
1990
ISSN/ISBN: 0022-1767 PMID: 1698853 Document Number: 353037
Human B lymphocytes were examined to determine whether transmembrane ion conductance plays a role in cell activation. Mitogens (anti-human IgM F(ab')2 fragment (anti-.mu.) and PMA) were used to stimulate B lymphocytes. Mitogen-induced DNA synthesis was inhibited by tetraethylammonium-Cl (TEA), 4-aminopyridine (4AP), verapamil, and diltiazem in a dose-dependent manner. This inhibition was not due to reduction in cell viability as determined by trypan blue exclusion. Mitogen-induced increases in RNA synthesis were partially inhibited by TEA and 4AP and were more completely inhibited by verapamil and diltiazem. Mitogen-induced cell volume increases were not affected by TEA or 4AP but were completely inhibited by verapamil and diltiazem. B lymphocytes stimulated with anti-.mu. expressed G1 phase cell surface antigens in the presence of TEA or 4AP, but failed to do so in the presence of verapamil or diltiazem. Substitution of PMA for anti-.mu. as the mitogen did not alter the effects of TEA or 4AP. However, verapamil inhibited PMA-induced expression of G1 phase cell surface markers although diltiazem did not. The patch clamp technique was used to directly examine plasma membrane ionic currents in whole-cell, cell-attached, and inside-out patch configurations. Activation of B lymphocytes with either anti-.mu. or the Ca2+ ionophore, A23187, inhibited opening of one type of channel in cell-attached patches. In inside-out patches, this channel type conducted current when the batch [Ca2+] was low (6 .times. 10-8 M) but failed to conduct current when the bath [CA2+] was increased above 1 .times. 10-6 M. The results of these experiments are consistent with the hypothesis that activation of B lymphocytes induces alterations in plasma membrane ion conductance. Single channel studies suggest that activation induced increases in [Ca2+]i may directly inhibit a specific set of plasma membrane ion channels as one mechanism by which transmembrane ion flux is altered.