Probes for INS

Pharyngeal pouches, a series of outpocketings that bud from the foregut endoderm, are essential to the formation of craniofacial skeleton as well as several important structures like parathyroid and thymus. However, whether pharyngeal pouch progenitors exist in the developing gut tube remains unknown. Here, taking advantage of cell lineage tracing and transgenic ablation technologies, we identified a population of nkx2.3+ pouch progenitors in zebrafish embryos and demonstrated an essential requirement of ectodermal BMP2b for their specification.At early somite stages, nkx2.3+ cells located at lateral region of pharyngeal endoderm give rise to the pouch epithelium except a subpopulation expressing pdgfαa rather than nkx2.3. A small-scale screen of chemical inhibitors reveals that BMP signaling is necessary to specify these progenitors. Loss-of-function analyses show that BMP2b, expressed in the pharyngeal ectoderm, actives Smad effectors in endodermal cells to induce nkx2.3+ progenitors. Collectively, our study provides in vivo evidence for the existence of pouch progenitors and highlights the importance of BMP2b signaling in progenitor specification.

Major depressive disorder (MDD) patients display a common but often variable set of symptoms making successful, sustained treatment difficult to achieve. Separate depressive symptoms may be encoded by differential changes in distinct circuits in the brain, yet how discrete circuits underlie behavioral subsets of depression and how they adapt in response to stress has not been addressed. We identify two discrete circuits of parvalbumin-positive (PV) neurons in the ventral pallidum (VP) projecting to either the lateral habenula or ventral tegmental area contributing to depression. We find that these populations undergo different electrophysiological adaptations in response to social defeat stress, which are normalized by antidepressant treatment. Furthermore, manipulation of each population mediates either social withdrawal or behavioral despair, but not both. We propose that distinct components of the VP PV circuit can subserve related, yet separate depressive-like phenotypes in mice, which could ultimately provide a platform for symptom-specific treatments of depression.

Homeostatic control of breathing, heart rate, and body temperature relies on circuits within the brainstem modulated by the neurotransmitter serotonin (5-HT). Mounting evidence points to specialized neuronal subtypes within the 5-HT system, which have borne out in functional studies, including the modulation of distinct facets of homeostatic control. These functional differences, read out at the organismal level, are likely subserved by differences among 5-HT neuron subtypes at the cellular and molecular levels, including differences in the capacity to co-express other neurotransmitters such as glutamate, GABA, thyrotropin releasing hormone, and substance P encoded by the Tachykinin-1 (Tac1) gene. Here we characterize in mice a 5-HT neuron subtype identified by expression of Tac1 and the transcription factor gene Pet1, thus referred to as the Tac1-Pet1 neuron subtype. Transgenic cell labeling showed Tac1-Pet1 soma resident largely in the caudal medulla. Chemogenetic (CNO-hM4Di) perturbation of Tac1-Pet1 neuron activity resulted in blunting of the respiratory CO2 chemoreflex, which normally augments ventilation in response to hypercapnic acidosis to restore normal pH and PCO2 Tac1-Pet1 axonal boutons were found localized to brainstem areas implicated in respiratory modulation, with highest density in motor nuclei. These findings demonstrate that the activity of a Pet1 neuron subtype with potential to release both 5-HT and substance P is necessary for normal respiratory dynamics, likely via motor outputs that maintain airway patency and engage muscles of respiration. These Tac1-Pet1 neurons may complement the activity of Egr2-Pet1 neurons, previously established in respiratory chemoreception, but which do not innervate respiratory motor nuclei.

SIGNIFICANCE STATEMENT:

5-HT neurons modulate outputs as diverse as body temperature, respiration, aggression, and mood. We characterize a 5-HT neuron subtype defined by expression of Tachykinin1 and Pet1 (Tac1-Pet1 neurons) which projects to respiratory motor nuclei, and when silenced, blunts the ventilatory response to inhaled carbon dioxide. We employ genetic tools to access this subset of 5-HT neurons to query function, anatomy, and connectivity. Localization of synaptic boutons from Tac1-Pet1 neurons, primarily within motor regions, contrasts with those from previously described Egr2-Pet1 neurons, which are chemosensitive and reside in the raphe magnus and project primarily to chemosensory integration, but not motor, regions of the brainstem.

In the arcuate nucleus of the hypothalamus (ARH) satiety signaling (anorexigenic) pro-opiomelanocortin (POMC)-expressing and hunger signaling (orexigenic) agouti-related peptide (AgRP)-expressing neurons are key components of the neuronal circuits that control food intake and energy homeostasis. Here, we assessed whether the catecholamine noradrenalin directly modulates the activity of these neurons in mice. Perforated patch clamp recordings showed that noradrenalin changes the activity of these functionally antagonistic neurons in opposite ways, increasing the activity of the orexigenic NPY/AgRP neurons and decreasing the activity of the anorexigenic POMC neurons. Cell type-specific transcriptomics and pharmacological experiments revealed that the opposing effect on these neurons is mediated by the activation of excitatory α1A - and β- adrenergic receptors in NPY/AgRP neurons, while POMC neurons are inhibited via α2A - adrenergic receptors. Thus, the coordinated differential modulation of the key hypothalamic neurons in control of energy homeostasis assigns noradrenalin an important role to promote feeding.

Light Sheet Fluorescence Microscopy (LSFM) of whole organs, in particular the brain, offers a plethora of biological data imaged in 3D. This technique is however often hindered by cumbersome non-automated analysis methods. Here we describe an approach to fully automate the analysis by integrating with data from the Allen Institute of Brain Science (AIBS), to provide precise assessment of the distribution and action of peptide-based pharmaceuticals in the brain. To illustrate this approach, we examined the acute central nervous system effects of the glucagon-like peptide-1 (GLP-1) receptor agonist liraglutide. Peripherally administered liraglutide accessed the hypothalamus and brainstem, and led to activation in several brain regions of which most were intersected by projections from neurons in the lateral parabrachial nucleus. Collectively, we provide a rapid and unbiased analytical framework for LSFM data which enables quantification and exploration based on data from AIBS to support basic and translational discovery.