Destabilization of Phase-separated Structures in Local Anesthetic-containing Model Biomembranes

Abstract

We observed the phase behaviors, and the changes of membrane fluidity in lipid membranes containing local anesthetics (LAs), lidocaine and tetracaine. The phase separation in DOPC/DPPC binary membranes had not been changed significantly by addition of LAs, whereas that in DOPC/DPPC/ Chol ternary membranes was suppressed by LAs. In order to evaluate the membrane fluidity, Laurdan generalized polarization values of membranes were measured. The fluidity of DOPC-rich phase in the ternary membrane became lower as LAs concentration had increased. Our results indicated the correlation between physical property change on membrane and pharmacological functions of LAs.Local anesthetics (LAs) are essential drugs for modern medicine that are often used to suppress patients' pain. LAs are known to target functional proteins, voltage-dependent sodium channels, which are located on cellular membranes. The mechanism how LAs shield pain signals was believed to be that LAs interact with the channel protein directly, and deactivate it.1 However, binding sites on the channel proteins had not been elucidated completely. In addition, according to past reports, LAs exhibit toxicity against human cells. The presence of LAs sometimes induces fatal side effects such as allergy, liver malfunction, and analgesia. 2 The mechanisms by which these side effects are induced by LAs are also not fully understood. Recently, however, some groups are expecting that these mechanisms can be explained by the change of the physical properties of lipid membranes where the proteins are buried induced by addition of LAs. 3Cell membranes, which are composed of lipid bilayer structure, are known to relate with signal transductions via membrane dynamics such as vesicular formation. In fact, some studies are providing correlation between physical property changes on biomembrane and behavior of channel proteins. 4 Moreover, a "lipid raft model" was proposed by Simons and Ikonen in 1997. 5,6 Lipid raft is a region where a large amount of saturated lipids and cholesterols are included and some functional molecules such as ion channels and membrane receptors can be localized, and lipid raft is speculated as a platform for signal transduction.In order to reveal the mechanism of raft formation, liposome, which is mainly composed of phospholipids, is used as a mimetic model of a biomembrane system. Liposome has similar structure to actual biomembranes, so we can directly observe similar phenomena on liposome with optical microscopy. Especially, raft formation can be regarded as the phase separation between saturated lipid and cholesterol-rich and unsaturated lipid-rich phases in a model membrane system, so from a biological aspect, it is important to discuss phase behavior of model membrane. 7In this study, we observed the phase separation in lipid membranes containing typical LAs, lidocaine and tetracaine, by optical microscopy. In order to understand the phase separation, the membrane fluidity is estimated by Laurdan generalized polarization measurement. Finally, w...

Thermal Stability of Phase-Separated Domains in Multicomponent Lipid Membranes with Local Anesthetics

“…Tetracaine showed more significant effects than lidocaine on the physical properties of the lipid membranes, such as the miscibility temperature, line tension at the L o /L d interface, and main transition temperature. These findings are consistent with the results of a previous study, which reported that tetracaine suppresses phase separation at room temperature more strongly than lidocaine [ 25 ]. Moreover, Paiva et al showed that tetracaine has a stronger effect than lidocaine on the transition temperature of raft-mimetic membranes [ 23 ].…”

“…Furthermore, we have observed the suppression of phase separation on raft-mimetic liposomes composed of unsaturated and saturated lipids and Chol via microscopy. Since the fluidity of the L d phase decreases on the addition of LAs and the fluidity gap between the L o and L d phases reduces, these events result in the suppression of phase separation [ 25 ]. Conversely, in the case of binary lipid mixtures consisting of unsaturated and saturated lipids, the fluidity difference between the S o and L d phases is not adequately reduced.…”

“…Conversely, in the case of binary lipid mixtures consisting of unsaturated and saturated lipids, the fluidity difference between the S o and L d phases is not adequately reduced. Consequently, phase separation between S o and L d phases is not suppressed by LAs [ 25 ].…”

TRP Channels as Sensors of Chemically-Induced Changes in Cell Membrane Mechanical Properties

“…Once lipid rafts are destabilized, the order, structure, and movement of the VGSC subunits forming the structure of the channel gates may be altered [146]. Different LAs have different effects on the membrane, as for instance, tetracaine induces higher miscibility temperature, line tension at the ordered/disordered phase interface, and lipid main transition temperature and phase separation than lidocaine [147]. Additionally, other molecules such as n-alcohol general anesthetics are reported to correlate increasing molecule hydrophobicity, producing a decrease in lipid transition temperature, with an increase in the strength of anesthesia [148,149].…”

Chirality-Dependent Interaction of d- and l-Menthol with Biomembrane Models

“…Some membrane proteins, such as ion channels, are localized on lipid rafts. The activity of a channel is controlled by specific molecules, and changes in activity resulting from molecular binding in a channel generates a sensation, such as warmth, cold, or anaesthesia [ 10 ]. This binding molecule may not only affect the channel, but also the lipid bilayer, thus altering the physical properties of the membrane.…”