Gadolinium Ions Block Mechanosensitive Channels by Altering the Packing and Lateral Pressure of Anionic Lipids

Effects of polyvalent ions on the lateral packing of phospholipids have been known for decades, but the physiological consequences have not been systematically studied. Gd³⁺ is a relatively nonspecific agent that blocks mechano-gated channels with a variable affinity. In this study, we show that the large mechanosensitive channel MscL of Escherichia coli is effectively blocked by Gd³⁺ only when reconstituted with negatively charged phospholipids (e.g., PS). Taking this lead, we studied effects of Gd³⁺ on monolayers and unilamellar vesicles made of natural brain PS, DMPS, and its mixtures with DMPC. In monolayer experiments, we found that μM Gd³⁺ present in the subphase leads to ∼8% lateral compaction of brain PS (at 35 mN/m). Gd³⁺ more strongly shrinks and rigidifies DMPS films causing a spontaneous liquid expanded-to-compact transition to the limiting 40 Ų/mol. Pressure-area isotherms of uncharged DMPC were unaffected by Gd³⁺, and neutralization of DMPS surface by low pH did not produce strong compaction. Upshifts of surface potential isotherms of DMPS monolayers reflected changes in the diffuse double layer due to neutralization of headgroup charges by Gd³⁺, whereas the increased packing density produced up to a 200 mV change in the interfacial dipole potential. The slopes of surface potential versus reciprocal area predicted that Gd³⁺ induced a modest (∼18%) increase in the magnitude of the individual lipid dipoles in DMPS. Isothermal titration calorimetry indicated that binding of Gd³⁺ to DMPS liposomes in the gel state is endothermic, whereas binding to liquid crystalline liposomes produces heat consistent with the isothermal liquid-to-gel phase transition induced by the ion. Both titration curves suggested a K_b of ∼10⁶ M⁻¹. We conclude that anionic phospholipids serve as high-affinity receptors for Gd³⁺ ions, and the ion-induced compaction generates a lateral pressure increase estimated as tens of mN/m. This pressure can “squeeze” the channel and shift the equilibrium toward the closed state.