A neuron-glia lipid metabolic cycle couples daily sleep to mitochondrial homeostasis.

Sleep is thought to be restorative to brain energy homeostasis, but it is not clear how this is achieved. We show here that Drosophila glia exhibit a daily cycle of glial mitochondrial oxidation and lipid accumulation that is dependent on prior wake and requires the Drosophila APOE orthologs NLaz and GLaz, which mediate neuron-glia lipid transfer. In turn, a full night of sleep is required for glial lipid clearance, mitochondrial oxidative recovery and maximal neuronal mitophagy. Knockdown of neuronal NLaz causes oxidative stress to accumulate in neurons, and the neuronal mitochondrial integrity protein, Drp1, is required for daily glial lipid accumulation. These data suggest that neurons avoid accumulation of oxidative mitochondrial damage during wake by using mitophagy and passing damage to glia in the form of lipids. We propose that a mitochondrial lipid metabolic cycle between neurons and glia reflects a fundamental function of sleep relevant for brain energy homeostasis.

Anandamide Metabolites Protect against Seizures through the TRP Channel Water Witch in Drosophila melanogaster.

Endocannabinoids protect against seizures, but their mechanism of action is still unclear, as they can have effects independent of known cannabinoid receptors. Using Drosophila melanogaster, which lacks canonical cannabinoid receptors, we report that the endocannabinoids anandamide and 2-arachidonoylglycerol protect against seizures in multiple fly seizure models. Surprisingly, inhibition of anandamide catabolism renders flies insensitive to protection by anandamide, indicating that anandamide metabolites are responsible for seizure protection. Consistent with this finding, arachidonic acid, a direct metabolite of anandamide, protects against seizures. To identify downstream effectors, we test for a role of transient receptor potential (TRP) channels and find that the TRPV1 antagonist capsazepine blocks the protective effect of anandamide. Also, a targeted genetic screen of TRP channels identifies water witch as a mediator of protection by anandamide. Using a Drosophila model, we reveal the role of arachidonic acid in seizure protection and identify a cannabinoid-receptor-1/2-independent mechanism of endocannabinoid seizure protection.

Photoaffinity Ligand for the Inhalational Anesthetic Sevoflurane Allows Mechanistic Insight into Potassium Channel Modulation.

Sevoflurane is a commonly used inhaled general anesthetic. Despite this, its mechanism of action remains largely elusive. Compared to other anesthetics, sevoflurane exhibits distinct functional activity. In particular, sevoflurane is a positive modulator of voltage-gated Shaker-related potassium channels (Kv1.x), which are key regulators of action potentials. Here, we report the synthesis and validation of azisevoflurane, a photoaffinity ligand for the direct identification of sevoflurane binding sites in the Kv1.2 channel. Azisevoflurane retains major sevoflurane protein binding interactions and pharmacological properties within in vivo models. Photoactivation of azisevoflurane induces adduction to amino acid residues that accurately reported sevoflurane protein binding sites in model proteins. Pharmacologically relevant concentrations of azisevoflurane analogously potentiated wild-type Kv1.2 and the established mutant Kv1.2 G329T. In wild-type Kv1.2 channels, azisevoflurane photolabeled Leu317 within the internal S4-S5 linker, a vital helix that couples the voltage sensor to the pore region. A residue lining the same binding cavity was photolabeled by azisevoflurane and protected by sevoflurane in the Kv1.2 G329T. Mutagenesis of Leu317 in WT Kv1.2 abolished sevoflurane voltage-dependent positive modulation. Azisevoflurane additionally photolabeled a second distinct site at Thr384 near the external selectivity filter in the Kv1.2 G329T mutant. The identified sevoflurane binding sites are located in critical regions involved in gating of Kv channels and related ion channels. Azisevoflurane has thus emerged as a new tool to discover inhaled anesthetic targets and binding sites and investigate contributions of these targets to general anesthesia.