Mapping of clonal lineages across developmental stages in human neural differentiation

Cell stem cell

2023 Mar 14

You, Z;Wang, L;He, H;Wu, Z;Zhang, X;Xue, S;Xu, P;Hong, Y;Xiong, M;Wei, W;Chen, Y;
PMID: 36933556 | DOI: 10.1016/j.stem.2023.02.007

The cell lineages across developmental stages remain to be elucidated. Here, we developed single-cell split barcoding (SISBAR) that allows clonal tracking of single-cell transcriptomes across stages in an in vitro model of human ventral midbrain-hindbrain differentiation. We developed "potential-spective" and "origin-spective" analyses to investigate the cross-stage lineage relationships and mapped a multi-level clonal lineage landscape depicting the whole differentiation process. We uncovered many previously uncharacterized converging and diverging trajectories. Furthermore, we demonstrate that a transcriptome-defined cell type can arise from distinct lineages that leave molecular imprints on their progenies, and the multilineage fates of a progenitor cell-type represent the collective results of distinct rather than similar clonal fates of individual progenitors, each with distinct molecular signatures. Specifically, we uncovered a ventral midbrain progenitor cluster as the common clonal origin of midbrain dopaminergic (mDA) neurons, midbrain glutamatergic neurons, and vascular and leptomeningeal cells and identified a surface marker that can improve graft outcomes.

Glutamatergic and GABAergic neurons in pontine central gray mediate opposing valence-specific behaviors through a global network

Neuron

2023 Feb 28

Xiao, C;Wei, J;Zhang, GW;Tao, C;Huang, JJ;Shen, L;Wickersham, IR;Tao, HW;Zhang, LI;
PMID: 36893756 | DOI: 10.1016/j.neuron.2023.02.012

Extracting the valence of environmental cues is critical for animals' survival. How valence in sensory signals is encoded and transformed to produce distinct behavioral responses remains not well understood. Here, we report that the mouse pontine central gray (PCG) contributes to encoding both negative and positive valences. PCG glutamatergic neurons were activated selectively by aversive, but not reward, stimuli, whereas its GABAergic neurons were preferentially activated by reward signals. The optogenetic activation of these two populations resulted in avoidance and preference behavior, respectively, and was sufficient to induce conditioned place aversion/preference. Suppression of them reduced sensory-induced aversive and appetitive behaviors, respectively. These two functionally opponent populations, receiving a broad range of inputs from overlapping yet distinct sources, broadcast valence-specific information to a distributed brain network with distinguishable downstream effectors. Thus, PCG serves as a critical hub to process positive and negative valences of incoming sensory signals and drive valence-specific behaviors with distinct circuits.

A Sleep-Specific Midbrain Target for Sevoflurane Anesthesia

Advanced science (Weinheim, Baden-Wurttemberg, Germany)

2023 Mar 24

Yi, T;Wang, N;Huang, J;Wang, Y;Ren, S;Hu, Y;Xia, J;Liao, Y;Li, X;Luo, F;Ouyang, Q;Li, Y;Zheng, Z;Xiao, Q;Ren, R;Yao, Z;Tang, X;Wang, Y;Chen, X;He, C;Li, H;Hu, Z;
PMID: 36961096 | DOI: 10.1002/advs.202300189

Sevoflurane has been the most widely used inhaled anesthetics with a favorable recovery profile; however, the precise mechanisms underlying its anesthetic action are still not completely understood. Here the authors show that sevoflurane activates a cluster of urocortin 1 (UCN1+ )/cocaine- and amphetamine-regulated transcript (CART+ ) neurons in the midbrain involved in its anesthesia. Furthermore, growth hormone secretagogue receptor (GHSR) is highly enriched in sevoflurane-activated UCN1+ /CART+ cells and is necessary for sleep induction. Blockade of GHSR abolishes the excitatory effect of sevoflurane on UCN1+ /CART+ neurons and attenuates its anesthetic effect. Collectively, their data suggest that anesthetic action of sevoflurane necessitates the GHSR activation in midbrain UCN1+ /CART+ neurons, which provides a novel target including the nucleus and receptor in the field of anesthesia.

A Novel Single Vector Intersectional AAV Strategy for Interrogating Cellular Diversity and Brain Function

bioRxiv : the preprint server for biology

2023 Feb 08

Hughes, AC;Pollard, BG;Xu, B;Gammons, JW;Chapman, P;Bikoff, JB;Schwarz, LA;
PMID: 36798174 | DOI: 10.1101/2023.02.07.527312

As the discovery of cellular diversity in the brain accelerates, so does the need for functional tools that target cells based on multiple features, such as gene expression and projection target. By selectively driving recombinase expression in a feature-specific manner, one can utilize intersectional strategies to conditionally promote payload expression only where multiple features overlap. We developed Conditional Viral Expression by Ribozyme Guided Degradation (ConVERGD), a single-construct intersectional targeting strategy that combines a self-cleaving ribozyme with traditional FLEx switches. ConVERGD offers benefits over existing platforms, such as expanded intersectionality, the ability to accommodate larger and more complex payloads, and a vector design that is easily modified to better facilitate rapid toolkit expansion. To demonstrate its utility for interrogating neural circuitry, we employed ConVERGD to target an unexplored subpopulation of norepinephrine (NE)-producing neurons within the rodent locus coeruleus (LC) identified via single-cell transcriptomic profiling to co-express the stress-related endogenous opioid gene prodynorphin ( Pdyn ). These studies showcase ConVERGD as a versatile tool for targeting diverse cell types and reveal Pdyn -expressing NE + LC neurons as a small neuronal subpopulation capable of driving anxiogenic behavioral responses in rodents.

Single-Nucleus RNA Sequencing of Developing and Mature Superior Colliculus Identifies Neuronal Diversity and Candidate Mediators of Circuit Assembly

bioRxiv : the preprint server for biology

2023 Feb 07

Ayupe, AC;Choi, JS;Beckedorff, F;Mccartan, R;Levay, K;Park, KK;
PMID: 36778361 | DOI: 10.1101/2023.02.01.526254

The superior colliculus (SC) is a sensorimotor structure in the midbrain that integrates input from multiple sensory modalities to initiate motor commands. It undergoes well-characterized steps of circuit assembly during development, rendering the mouse SC a popular model to study establishment and refinement of neural connectivity. Here we performed single nucleus RNA-sequencing analysis of the mouse SC isolated at various developmental time points. Our study provides a transcriptomic landscape of the cell types that comprise the SC across murine development with particular emphasis on neuronal heterogeneity. We used these data to identify Pax7 as a marker for an anatomically homogeneous population of GABAergic neurons. Lastly, we report a repertoire of genes differentially expressed across the different postnatal ages, many of which are known to regulate axon guidance and synapse formation. Our data provide a valuable resource for interrogating the mechanisms of circuit development, and identifying markers for manipulating specific SC neuronal populations and circuits.

Distinct serotonergic pathways to the amygdala underlie separate behavioral features of anxiety

Nature neuroscience

2022 Dec 01

Yu, XD;Zhu, Y;Sun, QX;Deng, F;Wan, J;Zheng, D;Gong, W;Xie, SZ;Shen, CJ;Fu, JY;Huang, H;Lai, HY;Jin, J;Li, Y;Li, XM;
PMID: 36446933 | DOI: 10.1038/s41593-022-01200-8

Anxiety-like behaviors in mice include social avoidance and avoidance of bright spaces. Whether these features are distinctly regulated is unclear. We demonstrate that in mice, social and anxiogenic stimuli, respectively, increase and decrease serotonin (5-HT) levels in basal amygdala (BA). In dorsal raphe nucleus (DRN), 5-HT∩vGluT3 neurons projecting to BA parvalbumin (DRN5-HT∩vGluT3-BAPV) and pyramidal (DRN5-HT∩vGluT3-BAPyr) neurons have distinct intrinsic properties and gene expression and respond to anxiogenic and social stimuli, respectively. Activation of DRN5-HT∩vGluT3→BAPV inhibits 5-HT release via GABAB receptors on serotonergic terminals in BA, inducing social avoidance and avoidance of bright spaces. Activation of DRN5-HT∩vGluT3→BA neurons inhibits two subsets of BAPyr neurons via 5-HT1A receptors (HTR1A) and 5-HT1B receptors (HTR1B). Pharmacological inhibition of HTR1A and HTR1B in BA induces avoidance of bright spaces and social avoidance, respectively. These findings highlight the functional significance of heterogenic inputs from DRN to BA subpopulations in the regulation of separate anxiety-related behaviors.

Distribution and Activation of Melanin-Concentrating Hormone Receptor-1 at Dopaminergic, GABAergic, and Glutamatergic Neurons in the Ventral Tegmental Area

Carleton University

2022 Dec 16

Spencer, C;
| DOI: 10.22215/etd/2022-15217

Melanin-concentrating hormone (MCH) is an orexigenic neuropeptide that acts through its receptor (MCHR1) to promote positive energy balance by increasing food intake and decreasing energy expenditure. MCH has been shown to inhibit dopamine release from the mesocorticolimbic dopamine pathway originating in the ventral tegmental area (VTA), and a hyperdopaminergic state underlies hyperactivity observed in animals lacking MCH or MCHR1. However, it is not known if the inhibitory effect of MCH on dopaminergic tone could be due to direct regulation of dopaminergic VTA neurons. We used a combination of molecular, neuroanatomical, and electrophysiological techniques to assess MCHR1 expression and activation in the VTA. MCH neurons project to the VTA, which comprises nerve terminals that contain MCH and may represent MCH release sites. Consistent with this, we detected MCHR1 expression on major VTA cell types, including those that are dopaminergic, GABAergic, and glutamatergic. Functional MCHR1 activation may regulate dopamine release via two mechanisms, one by acutely and directly inhibiting dopaminergic VTA neurons, and the other by disinhibiting glutamatergic afferents to dopaminergic VTA neurons. While we have not measured dopamine release in this present thesis, we postulate that MCH may acutely suppress dopamine release, while concurrently engaging local glutamatergic signaling to restore dopamine levels. These results signify that the VTA is a novel target for MCH-mediated physiology, including for the maintenance of energy homeostasis

Hedgehog-interacting protein acts in the habenula to regulate nicotine intake

Proceedings of the National Academy of Sciences of the United States of America

2022 Nov 15

Caligiuri, SPB;Howe, WM;Wills, L;Smith, ACW;Lei, Y;Bali, P;Heyer, MP;Moen, JK;Ables, JL;Elayouby, KS;Williams, M;Fillinger, C;Oketokoun, Z;Lehmann, VE;DiFeliceantonio, AG;Johnson, PM;Beaumont, K;Sebra, RP;Ibanez-Tallon, I;Kenny, PJ;
PMID: 36346845 | DOI: 10.1073/pnas.2209870119

Hedgehog-interacting protein (HHIP) sequesters Hedgehog ligands to repress Smoothened (SMO)-mediated recruitment of the GLI family of transcription factors. Allelic variation in HHIP confers risk of chronic obstructive pulmonary disease and other smoking-related lung diseases, but underlying mechanisms are unclear. Using single-cell and cell-type-specific translational profiling, we show that HHIP expression is highly enriched in medial habenula (MHb) neurons, particularly MHb cholinergic neurons that regulate aversive behavioral responses to nicotine. HHIP deficiency dysregulated the expression of genes involved in cholinergic signaling in the MHb and disrupted the function of nicotinic acetylcholine receptors (nAChRs) through a PTCH-1/cholesterol-dependent mechanism. Further, CRISPR/Cas9-mediated genomic cleavage of the <i>Hhip</i> gene in MHb neurons enhanced the motivational properties of nicotine in mice. These findings suggest that HHIP influences vulnerability to smoking-related lung diseases in part by regulating the actions of nicotine on habenular aversion circuits.

Postnatal age-differential ASD-like transcriptomic, synaptic, and behavioral deficits in Myt1l-mutant mice

Cell reports

2022 Sep 20

Kim, S;Oh, H;Choi, SH;Yoo, YE;Noh, YW;Cho, Y;Im, GH;Lee, C;Oh, Y;Yang, E;Kim, G;Chung, WS;Kim, H;Kang, H;Bae, Y;Kim, SG;Kim, E;
PMID: 36130507 | DOI: 10.1016/j.celrep.2022.111398

Myelin transcription factor 1 like (Myt1l), a zinc-finger transcription factor, promotes neuronal differentiation and is implicated in autism spectrum disorder (ASD) and intellectual disability. However, it remains unclear whether Myt1l promotes neuronal differentiation in vivo and its deficiency in mice leads to disease-related phenotypes. Here, we report that Myt1l-heterozygous mutant (Myt1l-HT) mice display postnatal age-differential ASD-related phenotypes: newborn Myt1l-HT mice, with strong Myt1l expression, show ASD-like transcriptomic changes involving decreased synaptic gene expression and prefrontal excitatory synaptic transmission and altered righting reflex. Juvenile Myt1l-HT mice, with markedly decreased Myt1l expression, display reverse ASD-like transcriptomes, increased prefrontal excitatory transmission, and largely normal behaviors. Adult Myt1l-HT mice show ASD-like transcriptomes involving astrocytic and microglial gene upregulation, increased prefrontal inhibitory transmission, and behavioral deficits. Therefore, Myt1l haploinsufficiency leads to ASD-related phenotypes in newborn mice, which are temporarily normalized in juveniles but re-appear in adults, pointing to continuing phenotypic changes long after a marked decrease of Myt1l expression in juveniles.

Early postnatal serotonin modulation prevents adult-stage deficits in Arid1b-deficient mice through synaptic transcriptional reprogramming

Nature communications

2022 Aug 27

Kim, H;Kim, D;Cho, Y;Kim, K;Roh, JD;Kim, Y;Yang, E;Kim, SS;Ahn, S;Kim, H;Kang, H;Bae, Y;Kim, E;
PMID: 36030255 | DOI: 10.1038/s41467-022-32748-5

Autism spectrum disorder is characterized by early postnatal symptoms, although little is known about the mechanistic deviations that produce them and whether correcting them has long-lasting preventive effects on adult-stage deficits. ARID1B, a chromatin remodeler implicated in neurodevelopmental disorders, including autism spectrum disorder, exhibits strong embryonic- and early postnatal-stage expression. We report here that Arid1b-happloinsufficient (Arid1b+/-) mice display autistic-like behaviors at juvenile and adult stages accompanied by persistent decreases in excitatory synaptic density and transmission. Chronic treatment of Arid1b+/- mice with fluoxetine, a selective serotonin-reuptake inhibitor, during the first three postnatal weeks prevents synaptic and behavioral deficits in adults. Mechanistically, these rescues accompany transcriptomic changes, including upregulation of FMRP targets and normalization of HDAC4/MEF2A-related transcriptional regulation of the synaptic proteins, SynGAP1 and Arc. These results suggest that chronic modulation of serotonergic receptors during critical early postnatal periods prevents synaptic and behavioral deficits in adult Arid1b+/- mice through transcriptional reprogramming.

Control of non-REM sleep by ventrolateral medulla glutamatergic neurons projecting to the preoptic area

Nature communications

2022 Aug 12

Teng, S;Zhen, F;Wang, L;Schalchli, JC;Simko, J;Chen, X;Jin, H;Makinson, CD;Peng, Y;
PMID: 35961989 | DOI: 10.1038/s41467-022-32461-3

Understanding the neural mechanisms underlying sleep state transitions is a fundamental goal of neurobiology and important for the development of new treatments for insomnia and other sleep disorders. Yet, brain circuits controlling this process remain poorly understood. Here we identify a population of sleep-active glutamatergic neurons in the ventrolateral medulla (VLM) that project to the preoptic area (POA), a prominent sleep-promoting region, in mice. Microendoscopic calcium imaging demonstrate that these VLM glutamatergic neurons display increased activity during the transitions from wakefulness to Non-Rapid Eye Movement (NREM) sleep. Chemogenetic silencing of POA-projecting VLM neurons suppresses NREM sleep, whereas chemogenetic activation of these neurons promotes NREM sleep. Moreover, we show that optogenetic activation of VLM glutamatergic neurons or their projections in the POA initiates NREM sleep in awake mice. Together, our findings uncover an excitatory brainstem-hypothalamic circuit that controls the wake-sleep transitions.

Examining ventral subiculum and basolateral amygdala projections to the nucleus accumbens shell: Differential expression of VGLuT1, VGLuT2 and VGaT in the rat

Neuroscience letters

2022 Aug 26

Jin, S;Maddern, XJ;Campbell, EJ;Lawrence, AJ;
PMID: 36038028 | DOI: 10.1016/j.neulet.2022.136858

Projections to the striatum are well-identified. For example, in the ventral striatum, two major inputs to the medial nucleus accumbens shell include the ventral subiculum and basolateral amygdala. However, the chemical phenotype(s) of these projection neurons remain unclear. In this study, we examined amygdalostriatal and corticostriatal connectivity in rats using injections of the retrograde tracer cholera toxin b into the nucleus accumbens shell. To determine the neurotransmitter identity of projection neurons, we combined retrograde tracing with RNAscope in-situ hybridization, using mRNA probes against vesicular transporters associated with glutamatergic (VGluT1 - Slc17a7, VGluT2 - Slc17a6) or GABAergic (VGaT - Slc32a1) neurotransmission. Confocal imaging was used to examine vesicular transporter mRNA expression in the ventral subiculum and basolateral amygdala inputs to the nucleus accumbens shell. Both projections contained mostly VGluT1-expressing neurons. Interestingly, almost a quarter of ventral subiculum to nucleus accumbens shell projections co-expressed VGluT1 and VGluT2 compared to a relatively small number (∼3%) that were co-expressed in basolateral amygdala to nucleus accumbens shell afferents. However, almost a quarter of basolateral amygdala to nucleus accumbens shell projections were VGaT-positive. These findings highlight the diverse proportions of glutamatergic and GABAergic afferents in two major projections to the nucleus accumbens shell and raise important questions for functional studies.

Pages

	Description
sense Example: Hs-LAG3-sense	Standard probes for RNA detection are in antisense. Sense probe is reverse complent to the corresponding antisense probe.
Intron# Example: Mm-Htt-intron2	Probe targets the indicated intron in the target gene, commonly used for pre-mRNA detection
Pool/Pan Example: Hs-CD3-pool (Hs-CD3D, Hs-CD3E, Hs-CD3G)	A mixture of multiple probe sets targeting multiple genes or transcripts
No-XSp Example: Hs-PDGFB-No-XMm	Does not cross detect with the species (Sp)
XSp Example: Rn-Pde9a-XMm	designed to cross detect with the species (Sp)
O# Example: Mm-Islr-O1	Alternative design targeting different regions of the same transcript or isoforms
CDS Example: Hs-SLC31A-CDS	Probe targets the protein-coding sequence only
EnEm	Probe targets exons n and m
En-Em	Probe targets region from exon n to exon m
Retired Nomenclature
tvn Example: Hs-LEPR-tv1	Designed to target transcript variant n
ORF Example: Hs-ACVRL1-ORF	Probe targets open reading frame
UTR Example: Hs-HTT-UTR-C3	Probe targets the untranslated region (non-protein-coding region) only
5UTR Example: Hs-GNRHR-5UTR	Probe targets the 5' untranslated region only
3UTR Example: Rn-Npy1r-3UTR	Probe targets the 3' untranslated region only
Pan Example: Pool	A mixture of multiple probe sets targeting multiple genes or transcripts

Search form

Probes for SLC17A6

Content for comparison

Gene

Product

Research area

Category

Pages

Advanced Cell Diagnostics

Bio-Techne

Advanced Cell Diagnostics China