Probes for INS

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.

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.

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

Midbrain dopamine neurons project to numerous targets throughout the brain to modulate various behaviors and brain states. Within this small population of neurons exists significant heterogeneity based on physiology, circuitry, and disease susceptibility. Recent studies have shown that dopamine neurons can be subdivided based on gene expression; however, the extent to which genetic markers represent functionally relevant dopaminergic subpopulations has not been fully explored. Here we performed single-cell RNA-sequencing of mouse dopamine neurons and validated studies showing that Neurod6 and Grp are selective markers for dopaminergic subpopulations. Using a combination of multiplex fluorescent in situ hybridization, retrograde labeling, and electrophysiology in mice of both sexes, we defined the anatomy, projection targets, physiological properties, and disease vulnerability of dopamine neurons based on Grp and/or Neurod6 expression. We find that the combinatorial expression of Grp and Neurod6 defines dopaminergic subpopulations with unique features. Grp+/Neurod6+ dopamine neurons reside in the ventromedial VTA, send projections to the medial shell of the nucleus accumbens, and have non-canonical physiological properties. Grp+/Neurod6- DA neurons are found in the VTA as well as in the ventromedial portion of the SNc where they project selectively to the dorsomedial striatum. Grp-/Neurod6+ DA neurons represent a smaller VTA subpopulation, which is preferentially spared in a 6-OHDA model of Parkinson’s disease. Together our work provides detailed characterization of Neurod6 and Grp expression in the midbrain and generates new insights into how these markers define functionally relevant dopaminergic subpopulations with distinct projection patterns, physiology, and disease vulnerability.

Significance Statement Recent single cell gene profiling studies have uncovered new subpopulations of midbrain dopamine neurons defined by their specific genetic signature. How these genetically-defined cell types map onto known dopaminergic circuits and functionally-defined cell types is unknown. This study elucidates the anatomy, circuitry, physiologic properties, and disease susceptibility of subpopulations of midbrain DA neurons defined by their expression of two genetic markers. This work not only advances our understanding of the dopaminergic system by providing new information about the properties of specific dopamine neuron subpopulations, it also demonstrates that unbiased genetic classification of neurons can reveal functionally relevant cell types.

The locus coeruleus (LC)-norepinephrine (NE) system modulates a range of salient brain functions, including memory and response to stress. The LC-NE system is regulated by neurochemically diverse inputs, including a range of neuropeptides such as arginine-vasopressin (AVP). Whilst the origins of many of these LC inputs, their synaptic connectivity with LC neurons, and their contribution to LC-mediated brain functions, have been well characterized, this is not the case for the AVP-LC system. Therefore, our aims were to define the types of synapses formed by AVP+ fibers with LC neurons using immunohistochemistry together with confocal and transmission electron microscopy (TEM), the origins of such inputs, using retrograde tracers, and the plasticity of the LC AVP system in response to stress and spatial learning, using the maternal separation (MS) and Morris water maze (MWM) paradigms respectively, in rat. Confocal microscopy revealed that AVP+ fibers contacting tyrosine hydroxylase (TH)+ LC neurons were also immunopositive for vesicular glutamate transporter 2, a marker of presynaptic glutamatergic axons. TEM confirmed that AVP+ axons formed Gray type I (asymmetric) synapses with TH+ dendrites thus confirming excitatory synaptic connections between these systems. Retrograde tracing revealed that these LC AVP+ fibers originate from hypothalamic vasopressinergic magnocellular neurosecretory neurons (AVPMN). MS induced a significant increase in the density of LC AVP+ fibers. Finally, AVPMNN circuit upregulation by water-deprivation improved MWM performance while increased Fos expression was found in LC and efferent regions such as hippocampus and prefrontal cortex, suggesting that AVPMMN projections to LC could integrate homeostatic responses modifying neuroplasticity.