Brain Chimeroids reveal individual susceptibility to neurotoxic triggers.

Interindividual genetic variation affects the susceptibility to and progression of many diseases1,2. However, efforts to study how individual human brains differ in normal development and disease phenotypes are limited by the paucity of faithful cellular human models, and the difficulty of scaling current systems to represent multiple people. Here we present human brain Chimeroids, a highly reproducible, multidonor human brain cortical organoid model generated by the co-development of cells from a panel of individual donors in a single organoid. By reaggregating cells from multiple single-donor organoids at the neural stem cell or neural progenitor cell stage, we generate Chimeroids in which each donor produces all cell lineages of the cerebral cortex, even when using pluripotent stem cell lines with notable growth biases. We used Chimeroids to investigate interindividual variation in the susceptibility to neurotoxic triggers that exhibit high clinical phenotypic variability: ethanol and the antiepileptic drug valproic acid. Individual donors varied in both the penetrance of the effect on target cell types, and the molecular phenotype within each affected cell type. Our results suggest that human genetic background may be an important mediator of neurotoxin susceptibility and introduce Chimeroids as a scalable system for high-throughput investigation of interindividual variation in processes of brain development and disease.

α2-containing γ-aminobutyric acid type A receptors promote stress resiliency in male mice.

Brain α2-containing GABAA receptors play a critical role in the modulation of anxiety- and fear-like behavior. However, it is unknown whether these receptors also play a role in modulating resilience to chronic stress, and in which brain areas and cell types such an effect would be mediated. We evaluated the role of α2-containing GABAA receptors following chronic social defeat stress using male mice deficient in the α2 subunit globally or conditionally in dopamine D1- or D2-receptor-expressing neurons, e.g., within the nucleus accumbens (NAc). In addition, we examined the effect of the lack of the α2 subunit on intermediates of the glutathione synthesis pathway. We found that α2-containing GABAA receptors on D2-receptor-positive but not on D1-receptor-positive neurons promote resiliency to chronic social defeat stress, as reflected in social interaction tests. The pro-resiliency effects of α2-containing GABAA receptors on D2-receptor-positive neurons do not appear to be directly related to alterations in anxiety-like behavior, as reflected in the elevated plus-maze, light-dark box, and novel open field tests. Increases in indices of oxidative stress-reflected by increases in cystathionine levels and reductions in GSH/GSSG ratios-were found in the NAc and prefrontal cortex but not in the hippocampus of mice lacking α2-containing GABAA receptors. We conclude that α2-containing GABAA receptors within specific brain areas and cell populations promote stress resiliency independently of direct effects on anxiety-like behaviors. A potential mechanism contributing to this increased resiliency is the protection that α2-containing GABAA receptors provide against oxidative stress in NAc and the prefrontal cortex.

Postmitotic Prox1 Expression Controls the Final Specification of Cortical VIP Interneuron Subtypes.

Throughout development, neuronal identity is controlled by key transcription factors that determine the unique properties of a cell. During embryogenesis, the transcription factor Prox1 regulates VIP-positive cortical interneuron migration, survival, and connectivity. Here, we explore the role of Prox1 as a regulator of genetic programs that guide the final specification of VIP interneuron subtypes in early postnatal life. Synaptic in vitro electrophysiology in male and female mice shows that postnatal Prox1 removal differentially affects the dynamics of excitatory inputs onto VIP bipolar and multipolar subtypes. RNA sequencing reveals that one of the downstream targets of Prox1 is the postsynaptic protein Elfn1, a constitutive regulator of presynaptic release probability. Further genetic, pharmacological, and electrophysiological experiments demonstrate that removing Prox1 reduces Elfn1 function in VIP multipolar but not in bipolar cells. Finally, overexpression experiments and analysis of native Elfn1 mRNA expression reveal that Elfn1 levels are differentially controlled at the post-transcriptional stage. Thus, in addition to activity-dependent processes that contribute to the developmental trajectory of VIP cells, genetic programs engaged by Prox1 control the final differentiation of multipolar and bipolar subtypes.SIGNIFICANCE STATEMENT The transcription factor Prox1 generates functional diversification of cortical VIP interneuron subtypes in early postnatal life, thus expanding the inhibitory repertoire of the cortex.

Developmental divergence of sensory stimulus representation in cortical interneurons.

Vasocative-intestinal-peptide (VIP+) and somatostatin (SST+) interneurons are involved in modulating barrel cortex activity and perception during active whisking. Here we identify a developmental transition point of structural and functional rearrangements onto these interneurons around the start of active sensation at P14. Using in vivo two-photon Ca2+ imaging, we find that before P14, both interneuron types respond stronger to a multi-whisker stimulus, whereas after P14 their responses diverge, with VIP+ cells losing their multi-whisker preference and SST+ neurons enhancing theirs. Additionally, we find that Ca2+ signaling dynamics increase in precision as the cells and network mature. Rabies virus tracings followed by tissue clearing, as well as photostimulation-coupled electrophysiology reveal that SST+ cells receive higher cross-barrel inputs compared to VIP+ neurons at both time points. In addition, whereas prior to P14 both cell types receive direct input from the sensory thalamus, after P14 VIP+ cells show reduced inputs and SST+ cells largely shift to motor-related thalamic nuclei.

Maternal overnutrition during critical developmental periods leads to different health adversities in the offspring: relevance of obesity, addiction and schizophrenia.

Maternal overnutrition during sensitive periods of early development increases the risk for obesity and neuropsychiatric disorders later in life. However, it still remains unclear during which phases of early development the offspring is more vulnerable. Here, we investigate the effects of maternal high-fat diet (MHFD) at different stages of pre- or postnatal development and characterize the behavioral, neurochemical and metabolic phenotypes. We observe that MHFD exposure at pre-conception has no deleterious effects on the behavioral and metabolic state of the offspring. Late gestational HFD exposure leads to more prominent addictive-like behaviors with reduced striatal dopamine levels compared to early gestational HFD. Conversely, offspring exposed to MHFD during lactation display the metabolic syndrome and schizophrenia-like phenotype. The latter, is manifested by impaired sensory motor gating, and latent inhibition as well as enhanced sensitivity to amphetamine. These effects are accompanied by higher striatal dopamine levels. Together, our data suggest that MHFD exposure during specific stages of development leads to distinct neuropathological alterations that determine the severity and nature of poor health outcome in adulthood, which may provide insight in identifying effective strategies for early intervention.

The mesoSPIM initiative: open-source light-sheet microscopes for imaging cleared tissue.

Light-sheet microscopy is an ideal technique for imaging large cleared samples; however, the community is still lacking instruments capable of producing volumetric images of centimeter-sized cleared samples with near-isotropic resolution within minutes. Here, we introduce the mesoscale selective plane-illumination microscopy initiative, an open-hardware project for building and operating a light-sheet microscope that addresses these challenges and is compatible with any type of cleared or expanded sample ( www.mesospim.org ).

Calbindin-D28K Limits Dopamine Release in Ventral but Not Dorsal Striatum by Regulating Ca2+ Availability and Dopamine Transporter Function.

The calcium-binding protein calbindin-D28K, or calb1, is expressed at higher levels by dopamine (DA) neurons originating in the ventral tegmental area (VTA) than in the adjacent substantia nigra pars compacta (SNc). Calb1 has received attention for a potential role in neuroprotection in Parkinson's disease. The underlying physiological roles for calb1 are incompletely understood. We used cre-loxP technology to knock down calb1 in mouse DA neurons to test whether calb1 governs axonal release of DA in the striatum, detected using fast-scan cyclic voltammetry ex vivo. In the ventral but not dorsal striatum, calb1 knockdown elevated DA release and modified the spatiotemporal coupling of Ca2+ entry to DA release. Furthermore, calb1 knockdown enhanced DA uptake but attenuated the impact of DA transporter (DAT) inhibition by cocaine on underlying DA release. These data reveal that calb1 acts through a range of mechanisms underpinning both DA release and uptake to limit DA transmission in the ventral but not dorsal striatum.