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Probes for INS

ACD can configure probes for the various manual and automated assays for INS for RNAscope Assay, or for Basescope Assay compatible for your species of interest.

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Intra-Tumoral Nerve-Tracing in a Novel Syngeneic Model of High-Grade Serous Ovarian Carcinoma

Cells

2021 Dec 10

Barr, JL;Kruse, A;Restaino, AC;Tulina, N;Stuckelberger, S;Vermeer, SJ;Williamson, CS;Vermeer, DW;Madeo, M;Stamp, J;Bell, M;Morgan, M;Yoon, JY;Mitchell, MA;Budina, A;Omran, DK;Schwartz, LE;Drapkin, R;Vermeer, PD;
PMID: 34944001 | DOI: 10.3390/cells10123491

Dense tumor innervation is associated with enhanced cancer progression and poor prognosis. We observed innervation in breast, prostate, pancreatic, lung, liver, ovarian, and colon cancers. Defining innervation in high-grade serous ovarian carcinoma (HGSOC) was a focus since sensory innervation was observed whereas the normal tissue contains predominantly sympathetic input. The origin, specific nerve type, and the mechanisms promoting innervation and driving nerve-cancer cell communications in ovarian cancer remain largely unknown. The technique of neuro-tracing enhances the study of tumor innervation by offering a means for identification and mapping of nerve sources that may directly and indirectly affect the tumor microenvironment. Here, we establish a murine model of HGSOC and utilize image-guided microinjections of retrograde neuro-tracer to label tumor-infiltrating peripheral neurons, mapping their source and circuitry. We show that regional sensory neurons innervate HGSOC tumors. Interestingly, the axons within the tumor trace back to local dorsal root ganglia as well as jugular-nodose ganglia. Further manipulations of these tumor projecting neurons may define the neuronal contributions in tumor growth, invasion, metastasis, and responses to therapeutics.
Central Control Circuit for Context-Dependent Micturition

Cell.

2016 Sep 22

Hou XH, Hyun M, Taranda J, Huang KW, Todd E, Feng D, Atwater E, Croney D, Zeidel ML, Osten P, Sabatini BL.
PMID: 27662084 | DOI: 10.1016/j.cell.2016.08.073

Urine release (micturition) serves an essential physiological function as well as a critical role in social communication in many animals. Here, we show a combined effect of olfaction and social hierarchy on micturition patterns in adult male mice, confirming the existence of a micturition control center that integrates pro- and anti-micturition cues. Furthermore, we demonstrate that a cluster of neurons expressing corticotropin-releasing hormone (Crh) in the pontine micturition center (PMC) is electrophysiologically distinct from their Crh-negative neighbors and sends glutamatergic projections to the spinal cord. The activity of PMC Crh-expressing neurons correlates with and is sufficient to drive bladder contraction, and when silenced impairs micturition behavior. These neurons receive convergent input from widespread higher brain areas that are capable of carrying diverse pro- and anti-micturition signals, and whose activity modulates hierarchy-dependent micturition. Taken together, our results indicate that PMC Crh-expressing neurons are likely the integration center for context-dependent micturition behavior.

Dissociable Structural and Functional Hippocampal Outputs via Distinct Subiculum Cell Classes

Cell.

2018 Apr 10

Cembrowski MS, Phillips MG, DiLisio SF, Shields BC, Winnubst J, Chandrashekar J, Bas E, Spruston N.
PMID: 29681453 | DOI: 10.1016/j.cell.2018.03.031

The mammalian hippocampus, comprised of serially connected subfields, participates in diverse behavioral and cognitive functions. It has been postulated that parallel circuitry embedded within hippocampal subfields may underlie such functional diversity. We sought to identify, delineate, and manipulate this putatively parallel architecture in the dorsal subiculum, the primary output subfield of the dorsal hippocampus. Population and single-cell RNA-seq revealed that the subiculum can be divided into two spatially adjacent subregions associated with prominent differences in pyramidal cell gene expression. Pyramidal cells occupying these two regions differed in their long-range inputs, local wiring, projection targets, and electrophysiological properties. Leveraging gene-expression differences across these regions, we use genetically restricted neuronal silencing to show that these regions differentially contribute to spatial working memory. This work provides a coherent molecular-, cellular-, circuit-, and behavioral-level demonstration that the hippocampus embeds structurally and functionally dissociable streams within its serial architecture.

A neural network for intermale aggression to establish social hierarchy

Nature Neuroscience

2018 May 25

Stagkourakis S, Spigolon G, Williams P, Protzmann J, Fisone G, Broberger C.
PMID: - | DOI: 10.1038/s41593-018-0153-x

Intermale aggression is used to establish social rank. Several neuronal populations have been implicated in aggression, but the circuit mechanisms that shape this innate behavior and coordinate its different components (including attack execution and reward) remain elusive. We show that dopamine transporter-expressing neurons in the hypothalamic ventral premammillary nucleus (PMvDAT neurons) organize goal-oriented aggression in male mice. Activation of PMvDATneurons triggers attack behavior; silencing these neurons interrupts attacks. Regenerative PMvDAT membrane conductances interacting with recurrent and reciprocal excitation explain how a brief trigger can elicit a long-lasting response (hysteresis). PMvDAT projections to the ventrolateral part of the ventromedial hypothalamic and the supramammillary nuclei control attack execution and aggression reward, respectively. Brief manipulation of PMvDAT activity switched the dominance relationship between males, an effect persisting for weeks. These results identify a network structure anchored in PMvDAT neurons that organizes aggressive behavior and, as a consequence, determines intermale hierarchy.

Adnp-mutant mice with cognitive inflexibility, CaMKIIα hyperactivity, and synaptic plasticity deficits

Molecular psychiatry

2023 Jun 26

Cho, H;Yoo, T;Moon, H;Kang, H;Yang, Y;Kang, M;Yang, E;Lee, D;Hwang, D;Kim, H;Kim, D;Kim, JY;Kim, E;
PMID: 37365244 | DOI: 10.1038/s41380-023-02129-5

ADNP syndrome, involving the ADNP transcription factor of the SWI/SNF chromatin-remodeling complex, is characterized by developmental delay, intellectual disability, and autism spectrum disorders (ASD). Although Adnp-haploinsufficient (Adnp-HT) mice display various phenotypic deficits, whether these mice display abnormal synaptic functions remain poorly understood. Here, we report synaptic plasticity deficits associated with cognitive inflexibility and CaMKIIα hyperactivity in Adnp-HT mice. These mice show impaired and inflexible contextual learning and memory, additional to social deficits, long after the juvenile-stage decrease of ADNP protein levels to ~10% of the newborn level. The adult Adnp-HT hippocampus shows hyperphosphorylated CaMKIIα and its substrates, including SynGAP1, and excessive long-term potentiation that is normalized by CaMKIIα inhibition. Therefore, Adnp haploinsufficiency in mice leads to cognitive inflexibility involving CaMKIIα hyperphosphorylation and excessive LTP in adults long after its marked expressional decrease in juveniles.
SCAMPR, a single-cell automated multiplex pipeline for RNA quantification and spatial mapping

Cell reports methods

2022 Oct 24

Ali Marandi Ghoddousi, R;Magalong, VM;Kamitakahara, AK;Levitt, P;
PMID: 36313803 | DOI: 10.1016/j.crmeth.2022.100316

Spatial gene expression, achieved classically through in situ hybridization, is a fundamental tool for topographic phenotyping of cell types in the nervous system. Newly developed techniques allow for visualization of multiple mRNAs at single-cell resolution and greatly expand the ability to link gene expression to tissue topography, yet there are challenges in efficient quantification and analysis of these high-dimensional datasets. We have therefore developed the single-cell automated multiplex pipeline for RNA (SCAMPR), facilitating rapid and accurate segmentation of neuronal cell bodies using a dual immunohistochemistry-RNAscope protocol and quantification of low- and high-abundance mRNA signals using open-source image processing and automated segmentation tools. Proof of principle using SCAMPR focused on spatial mapping of gene expression by peripheral (vagal nodose) and central (visual cortex) neurons. The analytical effectiveness of SCAMPR is demonstrated by identifying the impact of early life stress on gene expression in vagal neuron subtypes.
Sodium-calcium exchanger-3 regulates pain "wind-up": From human psychophysics to spinal mechanisms

Neuron

2022 Jun 10

Trendafilova, T;Adhikari, K;Schmid, AB;Patel, R;Polgár, E;Chisholm, KI;Middleton, SJ;Boyle, K;Dickie, AC;Semizoglou, E;Perez-Sanchez, J;Bell, AM;Ramirez-Aristeguieta, LM;Khoury, S;Ivanov, A;Wildner, H;Ferris, E;Chacón-Duque, JC;Sokolow, S;Saad Boghdady, MA;Herchuelz, A;Faux, P;Poletti, G;Gallo, C;Rothhammer, F;Bedoya, G;Zeilhofer, HU;Diatchenko, L;McMahon, SB;Todd, AJ;Dickenson, AH;Ruiz-Linares, A;Bennett, DL;
PMID: 35705078 | DOI: 10.1016/j.neuron.2022.05.017

Repeated application of noxious stimuli leads to a progressively increased pain perception; this temporal summation is enhanced in and predictive of clinical pain disorders. Its electrophysiological correlate is "wind-up," in which dorsal horn spinal neurons increase their response to repeated nociceptor stimulation. To understand the genetic basis of temporal summation, we undertook a GWAS of wind-up in healthy human volunteers and found significant association with SLC8A3 encoding sodium-calcium exchanger type 3 (NCX3). NCX3 was expressed in mouse dorsal horn neurons, and mice lacking NCX3 showed normal, acute pain but hypersensitivity to the second phase of the formalin test and chronic constriction injury. Dorsal horn neurons lacking NCX3 showed increased intracellular calcium following repetitive stimulation, slowed calcium clearance, and increased wind-up. Moreover, virally mediated enhanced spinal expression of NCX3 reduced central sensitization. Our study highlights Ca2+ efflux as a pathway underlying temporal summation and persistent pain, which may be amenable to therapeutic targeting.
EASI-FISH for thick tissue defines lateral hypothalamus spatio-molecular organization

Cell

2021 Dec 22

Wang, Y;Eddison, M;Fleishman, G;Weigert, M;Xu, S;Wang, T;Rokicki, K;Goina, C;Henry, FE;Lemire, AL;Schmidt, U;Yang, H;Svoboda, K;Myers, EW;Saalfeld, S;Korff, W;Sternson, SM;Tillberg, PW;
PMID: 34875226 | DOI: 10.1016/j.cell.2021.11.024

Determining the spatial organization and morphological characteristics of molecularly defined cell types is a major bottleneck for characterizing the architecture underpinning brain function. We developed Expansion-Assisted Iterative Fluorescence In Situ Hybridization (EASI-FISH) to survey gene expression in brain tissue, as well as a turnkey computational pipeline to rapidly process large EASI-FISH image datasets. EASI-FISH was optimized for thick brain sections (300 μm) to facilitate reconstruction of spatio-molecular domains that generalize across brains. Using the EASI-FISH pipeline, we investigated the spatial distribution of dozens of molecularly defined cell types in the lateral hypothalamic area (LHA), a brain region with poorly defined anatomical organization. Mapping cell types in the LHA revealed nine spatially and molecularly defined subregions. EASI-FISH also facilitates iterative reanalysis of scRNA-seq datasets to determine marker-genes that further dissociated spatial and morphological heterogeneity. The EASI-FISH pipeline democratizes mapping molecularly defined cell types, enabling discoveries about brain organization.
NG2 glia-derived GABA release tunes inhibitory synapses and contributes to stress-induced anxiety

Nature communications

2021 Sep 30

Zhang, X;Liu, Y;Hong, X;Li, X;Meshul, CK;Moore, C;Yang, Y;Han, Y;Li, WG;Qi, X;Lou, H;Duan, S;Xu, TL;Tong, X;
PMID: 34593806 | DOI: 10.1038/s41467-021-25956-y

NG2 glia, also known as oligodendrocyte precursor cells (OPCs), play an important role in proliferation and give rise to myelinating oligodendrocytes during early brain development. In contrast to other glial cell types, the most intriguing aspect of NG2 glia is their ability to directly sense synaptic inputs from neurons. However, whether this synaptic interaction is bidirectional or unidirectional, or its physiological relevance has not yet been clarified. Here, we report that NG2 glia form synaptic complexes with hippocampal interneurons and that selective photostimulation of NG2 glia (expressing channelrhodopsin-2) functionally drives GABA release and enhances inhibitory synaptic transmission onto proximal interneurons in a microcircuit. The mechanism involves GAD67 biosynthesis and VAMP-2 containing vesicular exocytosis. Further, behavioral assays demonstrate that NG2 glia photoactivation triggers anxiety-like behavior in vivo and contributes to chronic social defeat stress.
mGlu1 potentiation enhances prelimbic somatostatin interneuron activity to rescue schizophrenia-like physiological and cognitive deficits

Cell reports

2021 Nov 02

Maksymetz, J;Byun, NE;Luessen, DJ;Li, B;Barry, RL;Gore, JC;Niswender, CM;Lindsley, CW;Joffe, ME;Conn, PJ;
PMID: 34731619 | DOI: 10.1016/j.celrep.2021.109950

Evidence for prefrontal cortical (PFC) GABAergic dysfunction is one of the most consistent findings in schizophrenia and may contribute to cognitive deficits. Recent studies suggest that the mGlu1 subtype of metabotropic glutamate receptor regulates cortical inhibition; however, understanding the mechanisms through which mGlu1 positive allosteric modulators (PAMs) regulate PFC microcircuit function and cognition is essential for advancing these potential therapeutics toward the clinic. We report a series of electrophysiology, optogenetic, pharmacological magnetic resonance imaging, and animal behavior studies demonstrating that activation of mGlu1 receptors increases inhibitory transmission in the prelimbic PFC by selective excitation of somatostatin-expressing interneurons (SST-INs). An mGlu1 PAM reverses cortical hyperactivity and concomitant cognitive deficits induced by N-methyl-d-aspartate (NMDA) receptor antagonists. Using in vivo optogenetics, we show that prelimbic SST-INs are necessary for mGlu1 PAM efficacy. Collectively, these findings suggest that mGlu1 PAMs could reverse cortical GABAergic deficits and exhibit efficacy in treating cognitive dysfunction in schizophrenia.
Spatially patterned excitatory neuron subtypes and projections of the claustrum

eLife

2021 Aug 16

Erwin, SR;Bristow, BN;Sullivan, KE;Kendrick, RM;Marriott, B;Wang, L;Clements, J;Lemire, AL;Jackson, J;Cembrowski, MS;
PMID: 34397382 | DOI: 10.7554/eLife.68967

The claustrum is a functionally and structurally complex brain region, whose very spatial extent remains debated. Histochemical-based approaches typically treat the claustrum as a relatively narrow anatomical region that primarily projects to the neocortex, whereas circuit-based approaches can suggest a broader claustrum region containing projections to the neocortex and other regions. Here, in the mouse, we took a bottom-up and cell-type-specific approach to complement and possibly unite these seemingly disparate conclusions. Using single-cell RNA-sequencing, we found that the claustrum comprises two excitatory neuron subtypes that are differentiable from the surrounding cortex. Multicolor retrograde tracing in conjunction with 12-channel multiplexed in situ hybridization revealed a core-shell spatial arrangement of these subtypes, as well as differential downstream targets. Thus, the claustrum comprises excitatory neuron subtypes with distinct molecular and projection properties, whose spatial patterns reflect the narrower and broader claustral extents debated in previous research. This subtype-specific heterogeneity likely shapes the functional complexity of the claustrum.
The Claustrum Supports Resilience to Distraction

Curr Biol.

2018 Aug 16

Atlan G, Terem A, Peretz-Rivlin N, Sehrawat K, Gonzales BJ, Pozner G, Tasaka G, Goll Y, Refaeli R, Zviran O, Lim BK, Groysman M, Goshen I, Mizrahi A, Nelken I, Citri A.
PMID: 30122531 | DOI: 10.1016/j.cub.2018.06.068

A barrage of information constantly assaults our senses, of which only a fraction is relevant at any given point in time. However, the neural circuitry supporting the suppression of irrelevant sensory distractors is not completely understood. The claustrum, a circuit hub with vast cortical connectivity, is an intriguing brain structure, whose restrictive anatomy, thin and elongated, has precluded functional investigation. Here, we describe the use of Egr2-CRE mice to access genetically defined claustral neurons. Utilizing conditional viruses for anterograde axonal labeling and retrograde trans-synaptic tracing, we validated this transgenic model for accessing the claustrum and extended the known repertoire of claustral input/output connectivity. Addressing the function of the claustrum, we inactivated CLEgr2+ neurons, chronically as well as acutely, in mice performing an automated two-alternative forced-choice behavioral task. Strikingly, inhibition of CLEgr2+ neurons did not significantly impact task performance under varying delay times and cue durations, but revealed a selective role for the claustrum in supporting performance in the presence of an irrelevant auditory distractor. Further investigation of behavior, in the naturalistic maternal pup-retrieval task, replicated the result of sensitization to an auditory distractor following inhibition of CLEgr2+ neurons. Initiating investigation into the underlying mechanism, we found that activation of CLEgr2+ neurons modulated cortical sensory processing, suppressing tone representation in the auditory cortex. This functional study, utilizing selective genetic access, implicates the claustrum in supporting resilienceto distraction, a fundamental aspect of attention.

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Description
sense
Example: Hs-LAG3-sense
Standard probes for RNA detection are in antisense. Sense probe is reverse complent to the corresponding antisense probe.
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Example: Mm-Htt-intron2
Probe targets the indicated intron in the target gene, commonly used for pre-mRNA detection
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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
EnEmProbe targets exons n and m
En-EmProbe 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

Enabling research, drug development (CDx) and diagnostics

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