Trapping of Syntaxin1a in Presynaptic Nanoclusters by a Clinically Relevant General Anesthetic

Abstract

Propofol is the most commonly used general anesthetic in humans. Our understanding of its mechanism of action has focused on its capacity to potentiate inhibitory systems in the brain. However, it is unknown whether other neural mechanisms are involved in general anesthesia. Here, we demonstrate that the synaptic release machinery is also a target. Using single-particle tracking photoactivation localization microscopy, we show that clinically relevant concentrations of propofol and etomidate restrict syntaxin1A mobility on the plasma membrane, whereas non-anesthetic analogs produce the opposite effect and increase syntaxin1A mobility. Removing the interaction with the t-SNARE partner SNAP-25 abolishes propofol-induced syntaxin1A confinement, indicating that syntaxin1A and SNAP-25 together form an emergent drug target. Impaired syntaxin1A mobility and exocytosis under propofol are both rescued by co-expressing a truncated syntaxin1A construct that interacts with SNAP-25. Our results suggest that propofol interferes with a step in SNARE complex formation, resulting in non-functional syntaxin1A nanoclusters.

Need for speed: Super-resolving the dynamic nanoclustering of syntaxin-1 at exocytic fusion sites

“…Syntaxin-1 is organized as nanoclusters in the NMJs of Drosophila larvae (Bademosi et al, 2017;Ullrich et al, 2015). To understand how activity-dependent neurotransmitter release alters the nanocluster organization of syntaxin-1 in the presynaptic membrane, Bademosi et al (2017Bademosi et al ( , 2018aBademosi et al ( , 2018b) developed a new protocol to image syntaxin-1-mEos2 in the NMJs of fixed and live third instar Drosophila larvae (Fig. 2).…”

“…Propofol is one of the most commonly used general anesthetics in humans. A recent study found that a clinically relevant concentration of propofol restricts the mobility of syntaxin-1 and increases nanoclustering of syntaxin-1 molecules on the plasma membrane of PC12 cells and NMJs of Drosophila larvae (Bademosi et al, 2018b). Interestingly, preventing the interaction between syntaxin-1 and SNAP-25 and SNARE assembly blocked this effect, suggesting that propofol can interfere with the building up of a release site and render cells fusion incompetent.…”

“…This technique allows for the quantification of the size distribution of protein clusters on the plasma membrane (Klar and Hell, 1999;Sieber et al, 2006;Sieber et al, 2007;van den Bogaart et al, 2011). PALM allows for the characterization of the size and distribution of protein clusters in fixed and live cells and allows single particle tracking (sptPALM) in live cells (Bademosi et al, 2018a;Bademosi et al, 2017;Bademosi et al, 2018b;Manley et al, 2008;Vanhauwaert et al, 2017;Yang et al, 2012). Unlike PALM, stochastic optical reconstruction microscopy (STORM) uses photoswitchable organic fluorophores to immunolabel endogenous proteins (Bar-On et al, 2012;Rust et al, 2006).…”

Mechanisms of sleep and general anaesthesia in Drosophila melanogaster: the unresponsive brain

“…Syntaxin1A can be tagged with a fluorescent probe and the dynamics of single molecules, such as location, and displacement, can be observed (Kusumi et al, 2014). Alterations in synaptic activity have been associated with changes in syntaxin1A mobility (Kasula et al, 2016, Gandasi and Barg, 2014, Bademosi et al, 2017. In the presence of clinically relevant doses of propofol and etomidate, syntaxin1A mobility is restricted (Bademosi et al, 2017).…”

“…On the other hand, more recently, evidence has implicated the synaptic release machinery as a presynaptic target of anaesthetics. A variety of anaesthetics have been shown to decrease neurotransmission, an effect mediated by proteins involved in the release of vesicles from the nerve terminal (Herring et al, 2011, Herring et al, 2009, Xie et al, 2013b, Bademosi et al, 2017. Lipid solubility (X-axis) correlates strongly with anaesthetic potency (Y-axis).…”