Regulation of stress-induced sleep perturbations by dorsal raphe VGLUT3 neurons in male mice.

Exposure to stressors has profound effects on sleep that have been linked to serotonin (5-HT) neurons of the dorsal raphe nucleus (DR). However, the DR also comprises glutamatergic neurons expressing vesicular glutamate transporter type 3 (DRVGLUT3), leading us to examine their role. Cell-type-specific tracing revealed that DRVGLUT3 neurons project to brain areas regulating arousal and stress. We found that chemogenetic activation of DRVGLUT3 neurons mimics stress-induced sleep perturbations. Furthermore, deleting VGLUT3 in the DR attenuated stress-induced sleep perturbations, especially after social defeat stress. In the DR, VGLUT3 is found in subsets of 5-HT and non-5-HT neurons. We observed that both populations are activated by acute stress, including those projecting to the ventral tegmental area. However, deleting VGLUT3 in 5-HT neurons minimally affected sleep regulation. These findings suggest that VGLUT3 expression in the DR drives stress-induced sleep perturbations, possibly involving non-5-HT DRVGLUT3 neurons.

Microglial TNFα controls daily changes in synaptic GABAARs and sleep slow waves.

Microglia sense the changes in their environment. How microglia actively translate these changes into suitable cues to adapt brain physiology is unknown. We reveal an activity-dependent regulation of cortical inhibitory synapses by microglia, driven by purinergic signaling acting on P2RX7 and mediated by microglia-derived TNFα. We demonstrate that sleep induces microglia-dependent synaptic enrichment of GABAARs in a manner dependent on microglial TNFα and P2RX7. We further show that microglia-specific depletion of TNFα alters slow waves during NREM sleep and blunt memory consolidation in sleep-dependent learning tasks. Together, our results reveal that microglia orchestrate sleep-intrinsic plasticity of synaptic GABAARs, sculpt sleep slow waves, and support memory consolidation.

Hybrid core-shell particles for mRNA systemic delivery.

mRNA based infectious disease vaccines have opened the venue for development of novel nucleic acids-based therapeutics. For all mRNA therapeutics dedicated delivery systems are required, where different functionalities and targeting abilities need to be optimized for the respective applications. One option for advanced formulations with tailored properties are lipid-polymer hybrid nanoparticles with complex nanostructure, which allow to combine features of several already well described nucleic acid delivery systems. Here, we explored hyaluronic acid (HA) as coating of liposome-mRNA complexes (LRCs) to investigate effects of the coating on surface charge, physicochemical characteristics and biological activity. HA was electrostatically attached to positively charged complexes, forming hybrid LRCs (HLRCs). At different N/P ratios, physico-chemical characterization of the two sets of particles showed similarity in size (around 200 nm) and mRNA binding abilities, while the presence of the HA shell conferred a negative surface charge to otherwise positive complexes. High transfection efficiency of LRCs and HLRCs in vitro has been obtained in THP-1 and human monocytes derived from PBMC, an interesting target cell population for cancer and immune related pathologies. In mice, quantitative biodistribution of radiolabeled LRC and HLRC particles, coupled with bioluminescence studies to detect the protein translation sites, hinted towards both particles' accumulation in the hepatic reticuloendothelial system (RES). mRNA translated proteins though was found mainly in the spleen, a major source for immune cells, with preference for expression in macrophages. The results showed that surface modifications of liposome-mRNA complexes can be used to fine-tune nanoparticle physico-chemical characteristics. This provides a tool for assembly of stable and optimized nanoparticles, which are prerequisite for future therapeutic interventions using mRNA-based nanomedicines.

Overcoming Pavlovian bias in semantic space.

Action is invigorated in the presence of reward-predicting stimuli and inhibited in the presence of punishment-predicting stimuli. Although valuable as a heuristic, this Pavlovian bias can also lead to maladaptive behaviour and is implicated in addiction. Here we explore whether Pavlovian bias can be overcome through training. Across five experiments, we find that Pavlovian bias is resistant to unlearning under most task configurations. However, we demonstrate that when subjects engage in instrumental learning in a verbal semantic space, as opposed to a motoric space, not only do they exhibit the typical Pavlovian bias, but this Pavlovian bias diminishes with training. Our results suggest that learning within the semantic space is necessary, but not sufficient, for subjects to unlearn their Pavlovian bias, and that other task features, such as gamification and spaced stimulus presentation may also be necessary. In summary, we show that Pavlovian bias, whilst robust, is susceptible to change with experience, but only under specific environmental conditions.

Peripheral natural killer cells in chronic hepatitis B patients display multiple molecular features of T cell exhaustion.

Antiviral effectors such as natural killer (NK) cells have impaired functions in chronic hepatitis B (CHB) patients. The molecular mechanism responsible for this dysfunction remains poorly characterised. We show that decreased cytokine production capacity of peripheral NK cells from CHB patients was associated with reduced expression of NKp30 and CD16, and defective mTOR pathway activity. Transcriptome analysis of patients NK cells revealed an enrichment for transcripts expressed in exhausted T cells suggesting that NK cell dysfunction and T cell exhaustion employ common mechanisms. In particular, the transcription factor TOX and several of its targets were over-expressed in NK cells of CHB patients. This signature was predicted to be dependent on the calcium-associated transcription factor NFAT. Stimulation of the calcium-dependent pathway recapitulated features of NK cells from CHB patients. Thus, deregulated calcium signalling could be a central event in both T cell exhaustion and NK cell dysfunction occurring during chronic infections.