Probes for INS

The ventral pallidum (VP) lies at the interface between sensory, motor, and cognitive processing-with a particular role in mounting behavioral responses to rewards. Though the VP is predominantly GABAergic, glutamate neurons were recently identified, though their relative abundances and respective roles are unknown. Here, we show that VP glutamate neurons are concentrated in the rostral ventromedial VP and project to qualitatively similar targets as do VP GABA neurons. At the functional level, we used optogenetics to show that activity in VP GABA neurons can drive positive reinforcement, particularly through projections to the ventral tegmental area (VTA). On the other hand, activation of VP glutamate neurons leads to behavioral avoidance, particularly through projections to the lateral habenula. These findings highlight cell-type and projection-target specific roles for VP neurons in behavioral reinforcement, dysregulation of which could contribute to the emergence of negative symptoms associated with drug addiction and other neuropsychiatric disease.

Molecular characterization of neurons across brain regions has revealed new taxonomies for understanding functional diversity even among classically defined neuronal populations. Neuronal diversity has become evident within the brain serotonin (5-HT) system, which is far more complex than previously appreciated. However, until now it has been difficult to define subpopulations of 5-HT neurons based on molecular phenotypes. We demonstrate that the MET receptor tyrosine kinase (MET) is specifically expressed in a subset of 5-HT neurons within the caudal part of the dorsal raphe nuclei (DRC) that is encompassed by the classic B6 serotonin cell group. Mapping from embryonic day 16 through adulthood reveals that MET is expressed almost exclusively in the DRC as a condensed, paired nucleus, with an additional sparse set of MET+ neurons scattered within the median raphe. Retrograde tracing experiments reveal that MET-expressing 5-HT neurons provide substantial serotonergic input to the ventricular/subventricular region that contains forebrain stem cells, but do not innervate the dorsal hippocampus or entorhinal cortex. Conditional anterograde tracing experiments show that 5-HT neurons in the DRC/B6 target additional forebrain structures such as the medial and lateral septum and the ventral hippocampus. Molecular neuroanatomical analysis identifies 14 genes that are enriched in DRC neurons, including 4 neurotransmitter/neuropeptide receptors and 2 potassium channels. These analyses will lead to future studies determining the specific roles that 5-HTMET+ neurons contribute to the broader set of functions regulated by the serotonergic system.

Cerebrovascular disease is the third most common cause of death in developed countries, but our understanding of the cells that compose the cerebral vasculature is limited. Here, using vascular single-cell transcriptomics, we provide molecular definitions for the principal types of blood vascular and vessel-associated cells in the adult mouse brain. We uncover the transcriptional basis of the gradual phenotypic change (zonation) along the arteriovenous axis and reveal unexpected cell type differences: a seamless continuum for endothelial cells versus a punctuated continuum for mural cells. We also provide insight into pericyte organotypicity and define a population of perivascular fibroblast-like cells that are present on all vessel types except capillaries. Our work illustrates the power of single-cell transcriptomics to decode the higher organizational principles of a tissue and may provide the initial chapter in a molecular encyclopaedia of the mammalian vasculature.

Neutrophils require directional cues to navigate through the complex structure of venular walls and into inflamed tissues. Here we applied confocal intravital microscopy to analyze neutrophil emigration in cytokine-stimulated mouse cremaster muscles. We identified differential and non-redundant roles for the chemokines CXCL1 and CXCL2, governed by their distinct cellular sources. CXCL1 was produced mainly by TNF-stimulated endothelial cells (ECs) and pericytes and supported luminal and sub-EC neutrophil crawling. Conversely, neutrophils were the main producers of CXCL2, and this chemokine was critical for correct breaching of endothelial junctions. This pro-migratory activity of CXCL2 depended on the atypical chemokine receptor 1 (ACKR1), which is enriched within endothelial junctions. Transmigrating neutrophils promoted a self-guided migration response through EC junctions, creating a junctional chemokine "depot" in the form of ACKR1-presented CXCL2 that enabled efficient unidirectional luminal-to-abluminal migration. Thus, CXCL1 and CXCL2 act in a sequential manner to guide neutrophils through venular walls as governed by their distinct cellular sources.