Probes for INS

Pancreatic islet cells are critical for maintaining normal blood glucose levels, and their malfunction underlies diabetes development and progression. We used single-cell RNA sequencing to determine the transcriptomes of 1,492 human pancreatic α, β, δ, and PP cells from non-diabetic and type 2 diabetes organ donors. We identified cell-type-specific genes and pathways as well as 245 genes with disturbed expression in type 2 diabetes. Importantly, 92% of the genes have not previously been associated with islet cell function or growth. Comparison of gene profiles in mouse and human α and β cells revealed species-specific expression. All data are available for online browsing and download and will hopefully serve as a resource for the islet research community.

KCNQ2/3 channels, ubiquitously expressed neuronal potassium channels, have emerged as indispensable regulators of brain network activity. Despite their critical role in brain homeostasis, the mechanisms by which KCNQ2/3 dysfunction lead to hypersychrony are not fully known. Here, we show that deletion of KCNQ2/3 channels changed PV+ interneurons', but not SST+ interneurons', firing properties. We also find that deletion of either KCNQ2/3 or KCNQ2 channels from PV+ interneurons led to elevated homeostatic potentiation of fast excitatory transmission in pyramidal neurons. Pvalb-Kcnq2 null-mice showed increased seizure susceptibility, suggesting that decreases in interneuron KCNQ2/3 activity remodels excitatory networks, providing a new function for these channels.

Colony-stimulating factor 1 (CSF1) controls the growth and differentiation of macrophages.CSF1R signaling has been implicated in the maintenance of the intestinal stem cell niche and differentiation of Paneth cells, but evidence of expression of CSF1R within the crypt is equivocal. Here we show that CSF1R-dependent macrophages influence intestinal epithelial differentiation and homeostasis. In the intestinallamina propria CSF1R mRNA expression is restricted to macrophages which are intimately associated with the crypt epithelium, and is undetectable in Paneth cells. Macrophage ablation following CSF1R blockade affects Paneth cell differentiation and leads to a reduction of Lgr5+ intestinal stem cells. The disturbances to the crypt caused by macrophage depletion adversely affect the subsequent differentiation of intestinal epithelial cell lineages. Goblet cell density is enhanced, whereas the development of M cells in Peyer's patches is impeded. We suggest that modification of the phenotype or abundance of macrophages in the gut wall alters the development of the intestinal epithelium and the ability to sample gut antigens.

A wealth of data has elucidated the mechanisms by which sensory inputs are encoded in the neocortex, but how these processes are regulated by the behavioral relevance of sensory information is less understood. Here, we focus on neocortical layer 1 (L1), a key location for processing of such top-down information. Using Neuron-Derived Neurotrophic Factor(NDNF) as a selective marker of L1 interneurons (INs) and in vivo 2-photon calcium imaging, electrophysiology, viral tracing, optogenetics, and associative memory, we find that L1 NDNF-INs mediate a prolonged form of inhibition in distal pyramidal neuron dendrites that correlates with the strength of the memory trace. Conversely, inhibition from Martinotti cells remains unchanged after conditioning but in turn tightly controls sensory responses in NDNF-INs. These results define a genetically addressable form of dendritic inhibition that is highly experience dependent and indicate that in addition to disinhibition, salient stimuli are encoded at elevated levels of distal dendritic inhibition.

During neural development, self-avoidance ensures that a neuron's processes arborize to evenly fill a particular spatial domain. At the individual cell level, self-avoidance is promoted by genes encoding cell-surface molecules capable of generating thousands of diverse isoforms, such as Dscam1 (Down syndrome cell adhesion molecule 1) in Drosophila Isoform choice differs between neighboring cells, allowing neurons to distinguish "self" from "nonself". In the mouse retina, Dscam promotes self-avoidance at the level of cell types, but without extreme isoform diversity. Therefore, we hypothesize that DSCAM is a general self-avoidance cue that "masks" other cell type-specific adhesion systems to prevent overly exuberant adhesion. Here, we provide in vivo and in vitro evidence that DSCAM masks the functions of members of the cadherin superfamily, supporting this hypothesis. Thus, unlike the isoform-rich molecules tasked with self-avoidance at the individual cell level, here the diversity resides on the adhesive side, positioning DSCAM as a generalized modulator of cell adhesion during neural development.

Presynaptic inhibition (PSI) of primary sensory neurons is implicated in controlling gain and acuity in sensory systems. Here, we define circuit mechanisms and functions of PSI of cutaneous somatosensory neuron inputs to the spinal cord. We observed that PSI can be evoked by different sensory neuron populations and mediated through at least two distinct dorsal horn circuit mechanisms. Low-threshold cutaneousafferents evoke a GABAA-receptor-dependent form of PSI that inhibits similar afferent subtypes, whereas small-diameter afferentspredominantly evoke an NMDA-receptor-dependent form of PSI that inhibits large-diameter fibers. Behaviorally, loss of either GABAAreceptors (GABAARs) or NMDA receptors (NMDARs) in primary afferents leads to tactile hypersensitivity across skin types, and loss of GABAARs, but not NMDARs, leads to impaired texture discrimination. Post-weaning age loss of either GABAARs or NMDARs in somatosensory neurons causes systemic behavioral abnormalities, revealing critical roles of two distinct modes of PSI of somatosensory afferents in adolescence and throughout adulthood.

This study provides an assessment of the Fluidigm C1 platform for RNA sequencing of single mouse pancreatic islet cells. The system combines microfluidic technology and nanoliter-scale reactions. We sequenced 622 cells, allowing identification of 341 islet cells with high-quality gene expression profiles. The cells clustered into populations of α-cells (5%), β-cells (92%), δ-cells (1%), and pancreatic polypeptide cells (2%). We identified cell-type-specific transcription factors and pathways primarily involved in nutrient sensing and oxidation and cell signaling. Unexpectedly, 281 cells had to be removed from the analysis due to low viability, low sequencing quality, or contamination resulting in the detection of more than one islet hormone. Collectively, we provide a resource for identification of high-quality gene expression datasets to help expand insights into genes and pathways characterizing islet cell types. We reveal limitations in the C1 Fluidigm cell capture process resulting in contaminated cells with altered gene expression patterns. This calls for caution when interpreting single-cell transcriptomics data using the C1 Fluidigm system.

The incidence of non-alcoholic fatty liver disease (NAFLD) increases with age. Cellular senescence refers to a state of irreversible cell-cycle arrest combined with the secretion of proinflammatory cytokines and mitochondrial dysfunction. Senescent cells contribute to age-related tissue degeneration. Here we show that the accumulation of senescent cells promotes hepatic fat accumulation and steatosis. We report a close correlation between hepatic fat accumulation and markers of hepatocyte senescence. The elimination of senescent cells by suicide gene-meditated ablation of p16Ink4a-expressing senescent cells in INK-ATTAC mice or by treatment with a combination of the senolytic drugs dasatinib and quercetin (D+Q) reduces overall hepatic steatosis. Conversely, inducing hepatocyte senescence promotes fat accumulation in vitro and in vivo. Mechanistically, we show that mitochondria in senescent cells lose the ability to metabolize fatty acids efficiently. Our study demonstrates that cellular senescence drives hepatic steatosis and elimination of senescent cells may be a novel therapeutic strategy to reduce steatosis.

Anorexigenic pro-opiomelanocortin (Pomc)/alpha-melanocyte stimulating hormone (αMSH) neurons of the hypothalamic melanocortin system function as key regulators of energy homeostasis, also controlling somatic growth across different species. However, the mechanisms of melanocortin-dependent growth control still remain ill-defined. Here, we reveal a thus-far-unrecognized structural and functional connection between Pomc neurons and the somatotropic hypothalamo-pituitary axis. Excessive feeding of larval zebrafish causes leptin resistance and reduced levels of the hypothalamic satiety mediator pomca. In turn, this leads to reduced activation of hypophysiotropic somatostatin (Sst)-neurons that express the melanocortin receptor Mc4r, elevated growth hormone (GH) expression in the pituitary, and enhanced somatic growth. Mc4r expression and αMSH responsiveness are conserved in Sst-expressing hypothalamic neurons of mice. Thus, acquired leptin resistance and attenuation of pomca transcription in response to excessive caloric intake may represent an ancient mechanism to promote somatic growth when food resources are plentiful.

Sparse populations of neurons in the dentate gyrus (DG) of the hippocampus are causally implicated in the encoding of contextual fear memories. However, engram-specific molecular mechanisms underlying memory consolidation remain largely unknown. Here we perform unbiased RNA sequencing of DG engram neurons 24 h after contextual fear conditioning to identify transcriptome changes specific to memory consolidation. DG engram neurons exhibit a highly distinct pattern of gene expression, in which CREB-dependent transcription features prominently (P = 6.2 × 10-13), including Atf3 (P = 2.4 × 10-41), Penk (P = 1.3 × 10-15), and Kcnq3 (P = 3.1 × 10-12). Moreover, we validate the functional relevance of the RNAseq findings by establishing the causal requirement of intact CREB function specifically within the DG engram during memory consolidation, and identify a novel group of CREB target genes involved in the encoding of long-term memory