General Anesthetics Lower Critical Temperatures in Plasma Membrane Vesicles and Disrupt Antigen Induced Signaling in RBL-2H3 Cells

Abstract

There is a long standing debate whether general anesthetics act through non-specific perturbation of bilayer physical properties or through binding to specific sites within ion channels. In this study, we investigate how a series of liquid general anesthetics impact phase transition temperatures in giant plasma membrane vesicles (GPMVs) isolated from RBL-2H3 mast cells. We have chosen to use a series of alcohol general anesthetics with a range of chain lengths that have well characterized membrane partitioning properties and anesthetic potency. We find that all alcohols investigated lower critical temperatures in GPMVs. When alcohol concentrations are scaled by anesthetic potency, the magnitude of depression is equivalent within error. All compounds depress critical temperatures by 4±1°C when added to vesicles at their anesthetic dose. This is larger than depressions previously measured for the gel-liquid transition in vesicles. Since the functional effects of general anesthetic treated cells are well characterized in the ion channel context, we have chosen to investigate the effects of alcohols on another signaling pathway thought to be sensitive to membrane physical properties. RBL cells undergo immune signaling when IgE antibodies bound to FceRI receptors are cross-linked with a multivalent antigen, and early signaling steps are sensitive to plasma membrane lipids. We find that treatment with alcohols general anesthetics disrupts signaling as assayed by measuring de-granulation, a functional output downstream of receptor activation. Current work is investigating functional outputs at earlier signaling steps including calcium responses and receptor phosphorylation. We are investigating receptor mobility and signaling responses in cells treated with alcohol general anesthetics using super-resolution localization microscopy. By correlating the critical temperature lowering effects of alcohol with cell signaling events, we hope to better understand how membrane physical properties influence signaling processes.