Neurotransmitter switching in the adult mammalian brain occurs following photoperiod-induced stress, but the mechanism of regulation is unknown. Here, we demonstrate that elevated activity of dopaminergic neurons in the paraventricular nucleus of the hypothalamus (PaVN) in the adult rat is required for the loss of dopamine expression after long-day photoperiod exposure. The transmitter switch occurs exclusively in PaVN dopaminergic neurons that coexpress vesicular glutamate transporter 2 (VGLUT2), is accompanied by a loss of dopamine type 2 receptors (D2Rs) on corticotrophin-releasing factor (CRF) neurons, and can lead to increased release of CRF. Suppressing activity of all PaVN glutamatergic neurons decreases the number of inhibitory PaVN dopaminergic neurons, indicating homeostatic regulation of transmitter expression in the PaVN.

The dorsal horn of the spinal cord is critical to processing distinct modalities of noxious and innocuous sensation, but little is known of the neuronal subtypes involved, hampering efforts to deduce principles governing somatic sensation. Here we used single-cell RNA sequencing to classify sensory neurons in the mouse dorsal horn. We identified 15 inhibitory and 15 excitatory molecular subtypes of neurons, equaling the complexity in cerebral cortex. Validating our classification scheme in vivo and matching cell types to anatomy of the dorsal horn by spatial transcriptomics reveals laminar enrichment for each of the cell types. Neuron types, when combined, define a multilayered organization with like neurons layered together. Employing our scheme, we find that heat and cold stimuli activate discrete sets of both excitatory and inhibitory neuron types. This work provides a systematic and comprehensive molecular classification of spinal cord sensory neurons, enabling functional interrogation of sensory processing.

The experience of rewarding or aversive stimuli is encoded by distinct afferents to dopamine (DA) neurons of the ventral tegmental area (VTA). Several neuromodulatory systems including oxytocin regulate DA neuron excitability and synaptic transmission that process socially meaningful stimuli. We and others have recently characterized oxytocinergic modulation of activity in mouse VTA DA neurons, but the mechanisms underlying oxytocinergic modulation of synaptic transmission in DA neurons remain poorly understood. Here, we find that oxytocin application or optogenetic release decrease excitatory synaptic transmission, via long lasting, presynaptic, endocannabinoid-dependent mechanisms. Oxytocin modulation of excitatory transmission alters the magnitude of short and long-term depression. We find that only some glutamatergic projections to DA neurons express CB1 receptors. Optogenetic stimulation of three major VTA inputs demonstrates that oxytocin modulation is limited to projections that show evidence of CB1R transcripts. Thus, oxytocin gates information flow into reward circuits in a temporally selective and pathway-specific manner.

We report that 7SL, the RNA component of the signal recognition particle (SRP), is an abundant ncRNA in mature red blood cells (RBCs) of human, mouse, and the frog Xenopus. 7SL RNA in RBCs is not associated with the canonical proteins of the SRP. Instead, it co-immunoprecipitates from a lysate of RBCs with a number of membrane-binding proteins. Human and mouse RBCs also contain a previously undescribed 68 nt RNA, sRN7SL, derived from the "S domain" of 7SL RNA. We discuss the possibility that 7SL RNA is selectively protected from nucleases by association with the RBC membrane. Because 7SL is not associated with the canonical proteins of the SRP, it could represent a non-functional remnant of the protein synthetic machinery. Alternatively, it could play a new, as yet undefined role in RBC metabolism.

Dipeptidyl peptidase 9 (DPP9) is a ubiquitously expressed intracellular prolyl peptidase implicated in immunoregulation. However, its physiological relevance in the immune system remains largely unknown. We investigated the role of DPP9 enzyme in immune system by characterizing DPP9 knock-in mice expressing a catalytically inactive S729A mutant of DPP9 enzyme (DPP9ki/ki mice). DPP9ki/ki mice show reduced number of lymphoid and myeloid cells in fetal liver and postnatal blood but their hematopoietic cells are fully functional and able to reconstitute lymphoid and myeloid lineages even in competitive mixed chimeras. These studies demonstrate that inactivation of DPP9 enzymatic activity does not lead to any perturbations in mouse hematopoiesis.

Traumatic brain injury (TBI) commonly results in injury to the components of the white matter tracts, causing post-injury cognitive deficits. The myelin-producing oligodendrocytes (OLs) are vulnerable to TBI although may plausibly be replaced by proliferating oligodendrocyte progenitor cells (OPCs). The cytokine interleukin-1β (IL-1β) is a key mediator of the complex inflammatory response, and when neutralized in experimental TBI behavioral outcome was improved. To evaluate the role of IL-1β on OL cell death and OPC proliferation, 116 adult male mice subjected to sham injury or the central fluid percussion injury (cFPI) model of traumatic axonal injury, were analyzed at 2, 7 and 14 days post-injury. At 30 min post-injury, mice were randomly assigned to receiving an IL-1β neutralizing or a control antibody. OPC proliferation (5-ethynyl 2´- deoxyuridine (EdU)/Olig2 co-labeling) and mature OL cell loss was evaluated in injured white matter tracts. Microglia immunohistochemistry and ramification using Sholl analysis was also evaluated. Neutralizing IL-1β resulted in attenuated cell death, indicated by cleaved caspase-3 expression, and reduced loss of mature OLs from 2-7 days post-injury in brain-injured animals. IL-1β neutralization also attenuated the early, 2 day post-injury, increase of microglial immunoreactivity and the change in their ramification. However, the proliferation of OPCs in brain-injured animals was not altered. Our data suggest that IL-1β is involved in the TBI-induced loss of OLs and early microglial activation, although not the OPC proliferation. Attenuated oligodendrocyte cell loss may contribute to the improved behavioral outcome observed by IL-1β neutralization in this mouse model of diffuse TBI.

Background Perturbations to embryonic hemodynamics are known to adversely affect cardiovascular development. Vitelline vein ligation (VVL) is a model of reduced placental blood flow used to induce cardiac defects in early chick embryo development. The effect of these hemodynamic interventions on maturing elastic arteries is largely unknown. We hypothesize that hemodynamic changes impact maturation of the dorsal aorta (DA). Results We examined the effects of VVL on hemodynamic properties well into the maturation process and the corresponding changes in aortic dimensions, wall composition, and gene expression. In chick embryos, we found that DA blood velocity was reduced immediately post-surgery at Hamburger-Hamilton stage (HH) 18 and later at HH36, but not in the interim. Throughout this period, DA diameter adapted to maintain a constant shear stress. At HH36, we found that VVL DAs showed a substantial decrease in elastin and modest increase in collagen protein content. In VVL DAs, upregulation of elastic fiber related genes followed the downregulation of flow-dependent genes. Together, these suggest the existence of a compensatory mechanism in response to shear induced delays in maturation. Conclusions The DA's response to hemodynamic perturbations invokes coupled mechanisms for shear regulation and matrix maturation, potentially impacting the course of vascular development.

Protein arginine methylation has been recently identified as an important form of post-translational modification (PTM). It is carried out by the protein arginine methyltransferase (PRMT) family of enzymes, which in mammals consists of nine members. Among them, PRMT1 is the major arginine methyltransferase and participates in transcription, signal transduction, development and cancer. The function of PRMT1 in craniofacial development remains unclear. We generated Wnt1-Cre;Prmt1fl/fl mice with cranial neural crest (CNC)-specific deletion of Prmt1 and compared CNC-derived craniofacial bones from newborn control and Wnt1-Cre;Prmt1fl/fl mice. The size, surface area and volume of the premaxilla, maxilla, palatine bone, frontal bone, and mandible were analyzed using three-dimensional (3D) micro-computed tomography (microCT). We found that Prmt1 deficiency led to alterations in craniofacial bones including the premaxilla, maxilla, palatine bone, frontal bone, and mandible, as well as defects in the incisor and alveolar bone, recapitulating changes seen in Msx1-deficient mice. We further determined that Prmt1 depletion resulted in significant downregulation of Msx1 in calvaria-derived preosteoblast and primordium of frontal bone and mandible. Our study reveals critical roles of PRMT1 in the formation of CNC-derived craniofacial bones and suggests that Prmt1 is an upstream regulator of Msx1 in craniofacial bone development.

Pancreatic ductal adenocarcinoma (PDAC) is resistant to most therapies including single-agent immunotherapy and has a dense desmoplastic stroma, and most patients present with advanced metastatic disease. We reveal that macrophages are the dominant leukocyte population both in human PDAC stroma and autochthonous models, with an important functional contribution to the squamous subtype of human PDAC. We targeted macrophages in a genetic PDAC model using AZD7507, a potent selective inhibitor of CSF1R. AZD7507 caused shrinkage of established tumors and increased mouse survival in this difficult-to-treat model. Malignant cell proliferation diminished, with increased cell death and an enhanced T cell immune response. Loss of macrophages rewired other features of the TME, with global changes in gene expression akin to switching PDAC subtypes. These changes were markedly different to those elicited when neutrophils were targeted via CXCR2. These results suggest targeting the myeloid cell axis may be particularly efficacious in PDAC, especially with CSF1R inhibitors.

Abstract

BACKGROUND:

Converging evidence suggests a role for microglia-mediated neuroinflammation in Parkinson's disease (PD). Animal models of PD can serve as a platform to investigate the role of neuroinflammation in degeneration in PD. However, due to features of the previously available PD models, interpretations of the role of neuroinflammation as a contributor to or a consequence of neurodegeneration have remained elusive. In the present study, we investigated the temporal relationship of neuroinflammation in a model of synucleinopathy following intrastriatal injection of pre-formed alpha-synuclein fibrils (α-syn PFFS).

METHODS:

Male Fischer 344 rats (N = 114) received unilateral intrastriatal injections of α-syn PFFs, PBS, or rat serum albumin with cohorts euthanized at monthly intervals up to 6 months. Quantification of dopamine neurons, total neurons, phosphorylated α-syn (pS129) aggregates, major histocompatibility complex-II (MHC-II) antigen-presenting microglia, and ionized calcium-binding adaptor molecule-1 (Iba-1) immunoreactive microglial soma size was performed in the substantia nigra. In addition, the cortex and striatum were also examined for the presence of pS129 aggregates and MHC-II antigen-presenting microglia to compare the temporal patterns of pSyn accumulation and reactive microgliosis.

RESULTS:

Intrastriatal injection of α-syn PFFs to rats resulted in widespread accumulation of phosphorylated α-syn inclusions in several areas that innervate the striatum followed by significant loss (~ 35%) of substantia nigra pars compacta dopamine neurons within 5-6 months. The peak magnitudes of α-syn inclusion formation, MHC-II expression, and reactive microglial morphology were all observed in the SN 2 months following injection and 3 months prior to nigral dopamine neuron loss. Surprisingly, MHC-II immunoreactivity in α-syn PFF injected rats was relatively limited during the later interval of degeneration. Moreover, we observed a significant correlation between substantia nigra pSyn inclusion load and number of microglia expressing MHC-II. In addition, we observed a similar relationship between α-syn inclusion load and number of microglia expressing MHC-II in cortical regions, but not in the striatum.

CONCLUSIONS:

Our results demonstrate that increases in microglia displaying a reactive morphology and MHC-II expression occur in the substantia nigra in close association with peak numbers of pSyn inclusions, months prior to nigral dopamine neuron degeneration, and suggest that reactive microglia may contribute to vulnerability of SNc neurons to degeneration. The rat α-syn PFF model provides an opportunity to examine the innate immune response to accumulation of pathological α-syn in the context of normal levels of endogenous α-syn and provides insight into the earliest neuroinflammatory events in PD.

Diet has a profound effect on tissue regeneration in diverse organisms, and low caloric states such as intermittent fasting have beneficial effects on organismal health and age-associated loss of tissue function. The role of adult stem and progenitor cells in responding to short-term fasting and whether such responses improve regeneration are not well studied. Here we show that a 24 hr fast augments intestinal stem cell (ISC) function in young and aged mice by inducing a fatty acid oxidation (FAO) program and that pharmacological activation of this program mimics many effects of fasting. Acute genetic disruption of Cpt1a, the rate-limiting enzyme in FAO, abrogates ISC-enhancing effects of fasting, but long-term Cpt1a deletion decreases ISC numbers and function, implicating a role for FAO in ISC maintenance. These findings highlight a role for FAO in mediating pro-regenerative effects of fasting in intestinal biology, and they may represent a viable strategy for enhancing intestinal regeneration.

Tissues that undergo rapid cellular turnover, such as the mammalian haematopoietic system or the intestinal epithelium, are dependent on stem and progenitor cells that proliferate to provide differentiated cells to maintain organismal health. Stem and progenitor cells, in turn, are thought to rely on signals and growth factors provided by local niche cells to support their function and self-renewal. Several cell types have been hypothesized to provide the signals required for the proliferation and differentiation of the intestinal stem cells in intestinal crypts1-6. Here we identify subepithelial telocytes as an important source of Wnt proteins, without which intestinal stem cells cannot proliferate and support epithelial renewal. Telocytes are large but rare mesenchymal cells that are marked by expression of FOXL1 and form a subepithelial plexus that extends from the stomach to the colon. While supporting the entire epithelium, FOXL1+ telocytes compartmentalize the production of Wnt ligands and inhibitors to enable localized pathway activation. Conditional genetic ablation of porcupine (Porcn), which is required for functional maturation of all Wnt proteins, in mouse FOXL1+ telocytes causes rapid cessation of Wnt signalling to intestinal crypts, followed by loss of proliferation of stem and transit amplifying cells and impaired epithelial renewal. Thus, FOXL1+ telocytes are an important source of niche signals to intestinal stem cells.