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Summary: The paraventricular nucleus of the thalamus (PVT) projects axons to multiple areas, mediates a wide range of behaviors, and exhibits regional heterogeneity in both functions and axonal projections. Still, questions regarding the cell types present in the PVT and the extent of their differences remain inadequately addressed. We applied single-cell RNA sequencing to depict the transcriptomic characteristics of mouse PVT neurons. We found that one of the most significant variances in the PVT transcriptome corresponded to the anterior-posterior axis. While the single-cell transcriptome classified PVT neurons into five types, our transcriptomic and histological analyses showed continuity among the cell types. We discovered that anterior and posterior subpopulations had nearly non-overlapping projection patterns, while another population showed intermediate patterns. In addition, these subpopulations responded differently to appetite-related neuropeptides, with their activation showing opposing effects on food consumption. Our studies unveiled the contrasts and the continuity of PVT neurons that underpin their function.

Conditioned taste aversion (CTA) is a form of one-trial learning dependent on basolateral amygdala projection neurons (BLApn). Its underlying cellular and molecular mechanisms remain poorly understood. RNAseq from BLApn identified changes in multiple candidate learning-related transcripts including the expected immediate early gene Fos and Stk11, a master kinase of the AMP-related kinase pathway with important roles in growth, metabolism and development, but not previously implicated in learning. Deletion of Stk11 in BLApn blocked memory prior to training, but not following it and increased neuronal excitability. Conversely, BLApn had reduced excitability following CTA. BLApn knockout of a second learning-related gene, Fos, also increased excitability and impaired learning. Independently increasing BLApn excitability chemogenetically during CTA also impaired memory. STK11 and C-FOS activation were independent of one another. These data suggest key roles for Stk11 and Fos in CTA long-term memory formation, dependent at least partly through convergent action on BLApn intrinsic excitability.

Abstract The broadly-distributed, non-topographic projections to and from the olfactory cortex may suggest a flat, non-hierarchical organization in odor information processing. Layer 2 principal neurons in the anterior piriform cortex (APC) can be divided into 2 subtypes: semilunar (SL) and superficial pyramidal (SP) cells. Although it is known that SL and SP cells receive differential inputs from the olfactory bulb (OB), little is known about their projections to other olfactory regions. Here, we examined axonal projections of SL and SP cells using a combination of mouse genetics and retrograde labeling. Retrograde tracing from the OB or posterior piriform cortex (PPC) showed that the APC projects to these brain regions mainly through layer 2b cells, and dual-labeling revealed many cells extending collaterals to both target regions. Furthermore, a transgenic mouse line specifically labeling SL cells showed that they send profuse axonal projections to olfactory cortical areas, but not to the OB. These findings support a model in which information flow from SL to SP cells and back to the OB is mediated by a hierarchical feedback circuit, whereas both SL and SP cells broadcast information to higher olfactory areas in a parallel manner.

Transcription regulation is a key aspect of cellular identity established during development and maintained into adulthood. Molecular and biochemical assays that probe the genome are critical tools in exploring mechanisms of transcription regulation and cell type identity. The mammalian brain is composed of a huge diversity of cell types with distinct properties and functions. To understand these specific roles, it is necessary to selectively target cell populations for study. However, the need to selectively study restricted cell populations poses a challenge in neurobiology. It is often difficult to collect sufficient cellular input for many standard biochemical and molecular assays. Recently, important advances have been made to scale assays down, opening up new frontiers to explore molecular mechanisms in neurons. Concurrently, methodologies for preparing neurons for such assays has advanced taking into consideration specific methods to preserve the cell biology meant to be assayed. Here we describe a method for preparing live neurons from adult brain tissue for the Assay for Transposase Accessible Chromatin (ATAC).

Understanding the principles governing neuronal diversity is a fundamental goal for neuroscience. Here, we provide an anatomical and transcriptomic database of nearly 200 genetically identified cell populations. By separately analyzing the robustness and pattern of expression differences across these cell populations, we identify two gene classes contributing distinctly to neuronal diversity. Short homeobox transcription factors distinguish neuronal populations combinatorially, and exhibit extremely low transcriptional noise, enabling highly robust expression differences. Long neuronal effector genes, such as channels and cell adhesion molecules, contribute disproportionately to neuronal diversity, based on their patterns rather than robustness of expression differences. By linking transcriptional identity to genetic strains and anatomical atlases, we provide an extensive resource for further investigation of mouse neuronal cell types.

Synaptic scaling is a form of homeostatic plasticity driven by transcription-dependent changes in AMPA-type glutamate receptor (AMPAR) trafficking. To uncover the pathways involved, we performed a cell-type-specific screen for transcripts persistently altered during scaling, which identified the μ subunit (μ3A) of the adaptor protein complex AP-3A. Synaptic scaling increased μ3A (but not other AP-3 subunits) in pyramidal neurons and redistributed dendritic μ3A and AMPAR to recycling endosomes (REs). Knockdown of μ3A prevented synaptic scaling and this redistribution, while overexpression (OE) of full-length μ3A or a truncated μ3A that cannot interact with the AP-3A complex was sufficient to drive AMPAR to REs. Finally, OE of μ3A acted synergistically with GRIP1 to recruit AMPAR to the dendritic membrane. These data suggest that excess μ3A acts independently of the AP-3A complex to reroute AMPAR to RE, generating a reservoir of receptors essential for the regulated recruitment to the synaptic membrane during scaling up.

There is a continuing need for driver strains to enable cell-type-specific manipulation in the nervous system. Each cell type expresses a unique set of genes, and recapitulating expression of marker genes by BAC transgenesis or knock-in has generated useful transgenic mouse lines. However, since genes are often expressed in many cell types, many of these lines have relatively broad expression patterns. We report an alternative transgenic approach capturing distal enhancers for more focused expression. We identified an enhancer trap probe often producing restricted reporter expression and developed efficient enhancer trap screening with the PiggyBac transposon. We established more than 200 lines and found many lines that label small subsets of neurons in brain substructures, including known and novel cell types. Images and other information about each line are available online (enhancertrap.bio.brandeis.edu).

Cerebellar granule cells constitute the majority of neurons in the brain and are the primary conveyors of sensory and motor-related mossy fiber information to Purkinje cells. The functional capability of the cerebellum hinges on whether individual granule cells receive mossy fiber inputs from multiple precerebellar nuclei or are instead unimodal; this distinction is unresolved. Using cell-type-specific projection mapping with synaptic resolution, we observed the convergence of separate sensory (upper body proprioceptive) and basilar pontine pathways onto individual granule cells and mapped this convergence across cerebellar cortex. These findings inform the long-standing debate about the multimodality of mammalian granule cells and substantiate their associative capacity predicted in the Marr-Albus theory of cerebellar function. We also provide evidence that the convergent basilar pontine pathways carry corollary discharges from upper body motor cortical areas. Such merging of related corollary and sensory streams is a critical component of circuit models of predictive motor control.

Neuronal cell identity is established during development and must be maintained throughout an animal’s life (Fishell G, Heintz N. Neuron 80: 602–612, 2013). Transcription factors critical for establishing neuronal identity can be required for maintaining it (Deneris ES, Hobert O. Nat Neurosci 17: 899–907, 2014). Posttranscriptional regulation also plays an important role in neuronal differentiation (Bian S, Sun T. Mol Neurobiol 44: 359–373, 2011), but its role in maintaining cell identity is less established. To better understand how posttranscriptional regulation might contribute to cell identity, we examined the proprioceptive neurons in the dorsal root ganglion (DRG), a highly specialized sensory neuron class, with well-established properties that distinguish them from other neurons in the ganglion. By conditionally ablating Dicer in mice, using parvalbumin (Pvalb)-driven Cre recombinase, we impaired posttranscriptional regulation in the proprioceptive sensory neuron population. Knockout (KO) animals display a progressive form of ataxia at the beginning of the fourth postnatal week that is accompanied by a cell death within the DRG. Before cell loss, expression profiling shows a reduction of proprioceptor specific genes and an increased expression of nonproprioceptive genes normally enriched in other ganglion neurons. Furthermore, although central connections of these neurons are intact, the peripheral connections to the muscle are functionally impaired. Posttranscriptional regulation is therefore necessary to retain the transcriptional identity and support functional specialization of the proprioceptive sensory neurons. [ABSTRACT FROM AUTHOR]

The dopamine systems of the brain powerfully influence movement and motivation. We demonstrate that striatonigral fibers originating in striosomes form highly unusual bouquet-like arborizations that target bundles of ventrally extending dopamine-containing dendrites and clusters of their parent nigral cell bodies. Retrograde tracing showed that these clustered cell bodies in turn project to the striatum as part of the classic nigrostriatal pathway. Thus, these striosome-dendron formations, here termed "striosome-dendron bouquets," likely represent subsystems with the nigro-striato-nigral loop that are affected in human disorders including Parkinson's disease. Within the bouquets, expansion microscopy resolved many individual striosomal fibers tightly intertwined with the dopamine-containing dendrites and also with afferents labeled by glutamatergic, GABAergic, and cholinergicmarkers and markers for astrocytic cells and fibers and connexin 43 puncta. We suggest that the striosome-dendron bouquets form specialized integrative units within the dopamine-containing nigral system. Given evidence that striosomes receive input from cortical regions related to the control of mood and motivation and that they link functionally to reinforcement and decision-making, the striosome-dendron bouquets could be critical to dopamine-related function in health and disease. [ABSTRACT FROM AUTHOR]

Abstract Background Meckel’s diverticulum is considered the most prevalent congenital anomaly of the gastrointestinal tract. Approximately 4% of patients are symptomatic with complications such as bleeding, intestinal obstruction, and inflammation, while axial torsion of Meckel’s diverticulum is rare, particularly in pregnancy. Case presentation A 31-year-old woman in week 15 of pregnancy complained of epigastric pain, nausea and vomiting. Clinical diagnosis was severe hyperemesis gravidarum. Because the symptoms persisted during hospitalization, CT was performed and revealed dilated small bowel loops with multiple air-fluid levels. In the right mid-abdomen, there was a large part of air containing a cavity connected to the small intestine, which was considered a dilated bowel loop. Emergency laparotomy was performed and axial torsion of a large Meckel’s diverticulum measuring 11 cm was found at a few centimeters proximal to the ileocecal valve. Ileocecal resection including Meckel’s diverticulum was performed. The postoperative course was uneventful. At 40 weeks gestation, she had vaginal delivery of normal baby. Conclusion The physiological and anatomical changes in pregnancy can make a straightforward clinical diagnosis difficult. Prompt diagnosis and management were needed in order to avoid significant maternal and fetal risks. The use of imaging examinations, especially CT examination, with proper timing may be helpful to prevent delay in diagnosis and surgical intervention. Here, we report the case of a patient with axial torsion of Meckel’s diverticulum in pregnancy. To our knowledge, axial torsion of Meckel’s diverticulum in the first trimester of pregnancy has not been reported in the English medical literature.

Keratocystic Odontogenic Tumour (KCOT) is unicystic or multicystic intraosseous benign tumour of odontogenic origin that recurs due to locally destructive behaviour. KCOTs are usually the first manifestation of Nevoid Basal Cell Carcinoma Syndrome (NBCCS), an autosomal dominant disorder also known as Gorlin’s syndrome and they are most frequently observed familial symptom regardless of patients’ nationality. In addition, the recurrence rate and multiplicity of KCOTs is relatively high as compared to that of other sporadic carcinomas. KCOT has been considered as a non-hereditary lesion and its familial onset is an extremely rare event in non-NBCCS cases. Here, we describe previously unreported non-syndromic multiple KCOT cases in identical twins in a Japanese family. The subjects were female Japanese identical twins who were 26 and 27 years old, respectively, at the time of diagnosis for KCOT. They had no major or minor features of NBCCS other than KCOT. Although there were lesions that were likely to be dentigerous cysts based on radiographic findings, one of them was KCOT. This case report highlights the importance of precise diagnosis, choice of surgical method and careful observation for multiplicity or familial onset in sporadic KCOT cases without NBCCS.

Keratocystic odontogenic tumor (KCOT) arises as part of Gorlin syndrome (GS) or as a sporadic lesion. Gene mutations and loss of heterozygosity (LOH) of the hedgehog receptor PTCH1 plays an essential role in the pathogenesis of KCOT. However, some KCOT cases lack evidence for gene alteration of PTCH1, suggesting that other genes in the hedgehog pathway may be affected. PTCH2 and SUFU participate in the occurrence of GS-associated tumors, but their roles in KCOT development are unknown. To elucidate the roles of these genes, we enrolled 36 KCOT patients in a study to sequence their entire coding regions of PTCH1, PTCH2 and SUFU. LOH and immunohistochemical expression of these genes, as well as the downstream targets of hedgehog signaling, were examined using surgically-excised KCOT tissues. PTCH1 mutations, including four novel ones, were found in 9 hereditary KCOT patients, but not in sporadic KCOT patients. A pathogenic mutation of PTCH2 or SUFU was not found in any patients. LOH at PTCH1 and SUFU loci correlated with the presence of epithelial budding. KCOT harboring a germline mutation (Type 1) showed nuclear localization of GLI2 and frequent histological findings such as budding and epithelial islands, as well as the highest recurrence rate. KCOT with LOH but without a germline mutation (Type 2) less frequently showed these histological features, and the recurrence rate was lower. KCOT with neither germline mutation nor LOH (Type 3) consisted of two subgroups, Type 3A and 3B, which were characterized by nuclear and cytoplasmic GLI2 localization, respectively. Type 3B rarely exhibited budding and recurrence, behaving as the most amicable entity. The expression patterns of CCND1 and BCL2 tended to correlate with these subgroups. Our data indicates a significant role of PTCH1 and SUFU in the pathogenesis of KCOT, and the genotype-oriented subgroups constitute entities with different potential aggressiveness.