Lotun, A;Li, D;Xu, H;Su, Q;Tuncer, S;Sanmiguel, J;Mooney, M;Baer, CE;Ulbrich, R;Eyles, SJ;Strittmatter, L;Hayward, LJ;Gessler, DJ;Gao, G;
PMID: 37149081 | DOI: 10.1016/j.pneurobio.2023.102460
Myelinating oligodendrocytes are essential for neuronal communication and homeostasis of the central nervous system (CNS). One of the most abundant molecules in the mammalian CNS is N-acetylaspartate (NAA), which is catabolized into L-aspartate and acetate by the enzyme aspartoacylase (ASPA) in oligodendrocytes. The resulting acetate moiety is thought to contribute to myelin lipid synthesis. In addition, affected NAA metabolism has been implicated in several neurological disorders, including leukodystrophies and demyelinating diseases such as multiple sclerosis. Genetic disruption of ASPA function causes Canavan disease, which is hallmarked by increased NAA levels, myelin and neuronal loss, large vacuole formation in the CNS, and early death in childhood. Although NAA's direct role in the CNS is inconclusive, in peripheral adipose tissue, NAA-derived acetate has been found to modify histones, a mechanism known to be involved in epigenetic regulation of cell differentiation. We hypothesize that a lack of cellular differentiation in the brain contributes to the disruption of myelination and neurodegeneration in diseases with altered NAA metabolism, such as Canavan disease. Our study demonstrates that loss of functional Aspa in mice disrupts myelination and shifts the transcriptional expression of neuronal and oligodendrocyte markers towards less differentiated stages in a spatiotemporal manner. Upon re-expression of ASPA, these oligodendrocyte and neuronal lineage markers are either improved or normalized, suggesting that NAA breakdown by Aspa plays an essential role in the maturation of neurons and oligodendrocytes. Also, this effect of ASPA re-expression is blunted in old mice, potentially due to limited ability of neuronal, rather than oligodendrocyte, recovery.
Sanchez-Navarro, MJ;Borner, T;Reiner, BC;Crist, RC;Samson, WK;Yosten, GLC;Stein, L;Hayes, MR;
PMID: 37242151 | DOI: 10.3390/nu15102268
The g-protein coupled receptor GPR-160, recently identified as a putative receptor for the cocaine and amphetamine-regulated transcript (CART) peptide, shows abundant expression in the energy-balance control nuclei, including the dorsal vagal complex (DVC). However, its physiological role in the control of food intake has yet to be fully explored. Here, we performed a virally mediated, targeted knockdown (KD) of Gpr160 in the DVC of male rats to evaluate its physiological role in control of feeding. Our results indicate that DVC Gpr160 KD affects meal microstructure. Specifically, DVC Gpr160 KD animals consumed more frequent, but shorter meals during the dark phase and showed decreased caloric intake and duration of meals during the light phase. Cumulatively, however, these bidirectional effects on feeding resulted in no difference in body weight gain. We next tested the role of DVC GPR-160 in mediating the anorexigenic effects of exogenous CART. Our results show that DVC Gpr160 KD partially attenuates CART's anorexigenic effects. To further characterize Gpr160+ cells in the DVC, we utilized single-nucleus RNA sequencing data to uncover abundant GPR-160 expression in DVC microglia and only minimal expression in neurons. Altogether, our results suggest that DVC CART signaling may be mediated by Gpr160+ microglia, which in turn may be modulating DVC neuronal activity to control food intake.
Matrix biology : journal of the International Society for Matrix Biology
Neupane, S;Berardinelli, SJ;Cameron, DC;Grady, RC;Komatsu, DE;Percival, CJ;Takeuchi, M;Ito, A;Liu, TW;Nairn, AV;Moremen, KW;Haltiwanger, RS;Holdener, BC;
PMID: 35167946 | DOI: 10.1016/j.matbio.2022.02.002
Many extracellular matrix (ECM) associated proteins that influence ECM properties have Thrombospondin type 1 repeats (TSRs) which are modified with O-linked fucose. The O-fucose is added in the endoplasmic reticulum to folded TSRs by the enzyme Protein O-fucosyltransferase-2 (POFUT2) and is proposed to promote efficient trafficking of substrates. The importance of this modification for function of TSR-proteins is underscored by the early embryonic lethality of mouse embryos lacking Pofut2. To overcome early lethality and investigate the impact of the Pofut2 knockout on the secretion of POFUT2 substrates and on extracellular matrix properties in vivo, we deleted Pofut2 in the developing limb mesenchyme using Prrx1-Cre recombinase. Loss of Pofut2 in the limb mesenchyme caused significant shortening of the limbs, long bones and tendons and stiff joint resembling the musculoskeletal dysplasias in human and in mice with mutations in ADAMTS or ADAMTSL proteins. Limb shortening was evident at embryonic day 14.5 where loss of O-fucosylation led to an accumulation of fibrillin 2 (FBN2), decreased BMP and IHH signaling, and increased TGF-β signaling. Consistent with these changes we saw a decrease in the size of the hypertrophic zone with lower levels of Collagen-X. Unexpectedly, we observed minimal effects of the Pofut2 knockout on secretion of two POFUT2 substrates, CCN2 or ADAMTS17, in the developing bone. In contrast, CCN2 and two other POFUT2 substrates important for bone development, ADAMTS6 and 10, showed a decrease in secretion from POFUT2-null HEK293T cells in vitro. These combined results suggest that the impact of the Pofut2 mutation is cell-type specific. In addition, these observations raise the possibility that the O-fucose modification on TSRs extends beyond promoting efficient trafficking of POFUT2 substrates and has the potential to influence their function in the extracellular environment.
Mutation in the Ciliary Protein C2CD3 Reveals Organ-Specific Mechanisms of Hedgehog Signal Transduction in Avian Embryos
Journal of Developmental Biology
Brooks, E;Bonatto Paese, C;Carroll, A;Struve, J;Nagy, N;Brugmann, S;
| DOI: 10.3390/jdb9020012
Primary cilia are ubiquitous microtubule-based organelles that serve as signaling hubs for numerous developmental pathways, most notably the Hedgehog (Hh) pathway. Defects in the structure or function of primary cilia result in a class of diseases called ciliopathies. It is well known that primary cilia participate in transducing a Hh signal, and as such ciliopathies frequently present with phenotypes indicative of aberrant Hh function. Interestingly, the exact mechanisms of cilia-dependent Hh signaling transduction are unclear as some ciliopathic animal models simultaneously present with gain-of-Hh phenotypes in one organ system and loss-of-Hh phenotypes in another. To better understand how Hh signaling is perturbed across different tissues in ciliopathic conditions, we examined four distinct Hh-dependent signaling centers in the naturally occurring avian ciliopathic mutant talpid2 (ta2). In addition to the well-known and previously reported limb and craniofacial malformations, we observed dorsal-ventral patterning defects in the neural tube, and a shortened gastrointestinal tract. Molecular analyses for elements of the Hh pathway revealed that the loss of cilia impact transduction of an Hh signal in a tissue-specific manner at variable levels of the pathway. These studies will provide increased knowledge into how impaired ciliogenesis differentially regulates Hh signaling across tissues and will provide potential avenues for future targeted therapeutic treatments.
Davis MI, Crittenden JR, Feng AY, Kupferschmidt DA, Naydenov A, Stella N, Graybiel AM, Lovinger DM.
PMID: 29466446 | DOI: 10.1371/journal.pone.0191436
Presynaptic cannabinoid-1 receptors (CB1-R) bind endogenous and exogenous cannabinoids to modulate neurotransmitter release. CB1-Rs are expressed throughout the basal ganglia, including striatum and substantia nigra, where they play a role in learning and control of motivated actions. However, the pattern of CB1-R expression across different striatal compartments, microcircuits and efferent targets, and the contribution of different CB1-R-expressing neurons to this pattern, are unclear. We use a combination of conventional techniques and novel genetic models to evaluate CB1-R expression in striosome (patch) and matrix compartments of the striatum, and in nigral targets of striatal medium spiny projection neurons (MSNs). CB1-R protein and mRNA follow a descending dorsolateral-to-ventromedial intensity gradient in the caudal striatum, with elevated expression in striosomes relative to the surrounding matrix. The lateral predominance of striosome CB1-Rs contrasts with that of the classical striosomal marker, the mu opioid receptor (MOR), which is expressed most prominently in rostromedial striosomes. The dorsolateral-to-ventromedial CB1-R gradient is similar to Drd2 dopamine receptor immunoreactivity and opposite to Substance P. This topology of CB1-R expression is maintained downstream in the globus pallidus and substantia nigra. Dense CB1-R-expressing striatonigral fibers extend dorsally within the substantia nigra pars reticulata, and colocalize with bundles of ventrally extending, striosome-targeted, dendrites of dopamine-containing neurons in the substantia nigra pars compacta (striosome-dendron bouquets). Within striatum, CB1-Rs colocalize with fluorescently labeled MSN collaterals within the striosomes. Cre recombinase-mediated deletion of CB1-Rs from cortical projection neurons or MSNs, and MSN-selective reintroduction of CB1-Rs in knockout mice, demonstrate that the principal source of CB1-Rs in dorsolateral striosomes is local MSN collaterals. These data suggest a role for CB1-Rs in caudal dorsolateral striosome collaterals and striosome-dendron bouquet projections to lateral substantia nigra, where they are anatomically poised to mediate presynaptic disinhibition of both striosomal MSNs and midbrain dopamine neurons in response to endocannabinoids and cannabinomimetics.
Ledwon JK, Turin SY, Gosain AK, Topczewska JM.
PMID: 29630949 | DOI: 10.1016/j.gep.2018.04.002
Fibroblast growth factor (FGF) signaling is essential for many developmental processes and plays a pivotal role in skeletal homeostasis, regeneration and wound healing. FGF signals through one of five tyrosine kinase receptors: Fgfr1a, -1b, -2, -3, -4. To characterize the expression of zebrafish fgfr3 from the larval stage to adulthood, we used RNAscope in situ hybridization on paraffin sections of the zebrafish head. Our study revealed spatial and temporal distribution of fgfr3 transcript in chondrocytes of the head cartilages, osteoblasts involved in bone formation, ventricular zone of the brain, undifferentiated mesenchymal cells of the skin, and lens epithelium of the eye. In general, the expression pattern of zebrafish fgfr3 is similar to the expression observed in higher vertebrates.
Carbonaro, M;Wang, K;Huang, H;Frleta, D;Patel, A;Pennington, A;Desclaux, M;Moller-Tank, S;Grindley, J;Altarejos, J;Zhong, J;Polites, G;Poueymirou, W;Jaspers, S;Kyratsous, C;Zambrowicz, B;Murphy, A;Lin, JC;Macdonald, LE;Daly, C;Sleeman, M;Thurston, G;Li, Z;
PMID: 37058568 | DOI: 10.1126/sciadv.adf4490
Liver steatosis is an increasing health issue with few therapeutic options, partly because of a paucity of experimental models. In humanized liver rodent models, abnormal lipid accumulation in transplanted human hepatocytes occurs spontaneously. Here, we demonstrate that this abnormality is associated with compromised interleukin-6 (IL-6)-glycoprotein 130 (GP130) signaling in human hepatocytes because of incompatibility between host rodent IL-6 and human IL-6 receptor (IL-6R) on donor hepatocytes. Restoration of hepatic IL-6-GP130 signaling, through ectopic expression of rodent IL-6R, constitutive activation of GP130 in human hepatocytes, or humanization of an Il6 allele in recipient mice, substantially reduced hepatosteatosis. Notably, providing human Kupffer cells via hematopoietic stem cell engraftment in humanized liver mice also corrected the abnormality. Our observations suggest an important role of IL-6-GP130 pathway in regulating lipid accumulation in hepatocytes and not only provide a method to improve humanized liver models but also suggest therapeutic potential for manipulating GP130 signaling in human liver steatosis.
bioRxiv : the preprint server for biology
François, M;Delgado, IC;Lafond, A;Lewis, EM;Kuromaru, M;Hassouna, R;Deng, S;Thaker, VV;Dölen, G;Zeltser, LM;
PMID: 36824966 | DOI: 10.1101/2023.02.15.528679
Females are more sensitive to social exclusion, which could contribute to their heightened susceptibility to anxiety disorders. Chronic social isolation stress (CSIS) for at least 7 weeks after puberty induces anxiety-related behavioral adaptations in female mice. Here, we show that Arginine vasopressin receptor 1a ( Avpr1a )-expressing neurons in the central nucleus of the amygdala (CeA) mediate these sex-specific effects, in part, via projections to the caudate putamen. Loss of function studies demonstrate that AVPR1A signaling in the CeA is required for effects of CSIS on anxiety-related behaviors in females but has no effect in males or group housed females. This sex-specificity is mediated by AVP produced by a subpopulation of neurons in the posterodorsal medial nucleus of the amygdala that project to the CeA. Estrogen receptor alpha signaling in these neurons also contributes to preferential sensitivity of females to CSIS. These data support new therapeutic applications for AVPR1A antagonists in women.
Nikitin, P;Musina, G;Pekov, S;Kuzin, A;Popov, I;Belyaev, A;Kobyakov, G;Usachev, D;Nikolaev, V;Mikhailov, V;
| DOI: 10.3390/cancers15010145
Diffuse gliomas continue to be an important problem in neuro-oncology. To solve it, studies have considered the issues of molecular pathogenesis from the intratumoral heterogeneity point. Here, we carried out a comparative dynamic analysis of the different cell populations’ content in diffuse gliomas of different molecular profiles and grades, considering the cell populations’ functional properties and the relationship with patient survival, using flow cytometry, immunofluorescence, multiparametric fluorescent in situ hybridization, polymerase chain reaction, and cultural methods. It was shown that an increase in the IDH-mutant astrocytomas and oligodendrogliomas malignancy is accompanied by an increase in stem cells’ proportion and mesenchymal cell populations’ appearance arising from oligodendrocyte-progenitor-like cells with cell plasticity and cells’ hypoxia response programs’ activation. In glioblastomas, malignancy increase is accompanied by an increase in both stem and definitive cells with mesenchymal differentiation, while proneuronal glioma stem cells are the most likely the source of mesenchymal glioma stem cells, which, in hypoxic conditions, further give rise to mesenchymal-like cells. Clinical confirmation was a mesenchymal-like cell and mesenchymal glioma stem cell number, and the hypoxic and plastic molecular programs’ activation degree had a significant effect on relapse-free and overall survival. In general, we built a multi-vector model of diffuse gliomas’ pathogenetic tracing up to the practical plane.
Variation in phenotypes from a Bmp-Gata3 genetic pathway is modulated by Shh signaling
Swartz, ME;Lovely, CB;Eberhart, JK;
PMID: 34033651 | DOI: 10.1371/journal.pgen.1009579
We sought to understand how perturbation of signaling pathways and their targets generates variable phenotypes. In humans, GATA3 associates with highly variable defects, such as HDR syndrome, microsomia and choanal atresia. We previously characterized a zebrafish point mutation in gata3 with highly variable craniofacial defects to the posterior palate. This variability could be due to residual Gata3 function, however, we observe the same phenotypic variability in gata3 null mutants. Using hsp:GATA3-GFP transgenics, we demonstrate that Gata3 function is required between 24 and 30 hpf. At this time maxillary neural crest cells fated to generate the palate express gata3. Transplantation experiments show that neural crest cells require Gata3 function for palatal development. Via a candidate approach, we determined if Bmp signaling was upstream of gata3 and if this pathway explained the mutant's phenotypic variation. Using BRE:d2EGFP transgenics, we demonstrate that maxillary neural crest cells are Bmp responsive by 24 hpf. We find that gata3 expression in maxillary neural crest requires Bmp signaling and that blocking Bmp signaling, in hsp:DN-Bmpr1a-GFP embryos, can phenocopy gata3 mutants. Palatal defects are rescued in hsp:DN-Bmpr1a-GFP;hsp:GATA3-GFP double transgenic embryos, collectively demonstrating that gata3 is downstream of Bmp signaling. However, Bmp attenuation does not alter phenotypic variability in gata3 loss-of-function embryos, implicating a different pathway. Due to phenotypes observed in hypomorphic shha mutants, the Sonic Hedgehog (Shh) pathway was a promising candidate for this pathway. Small molecule activators and inhibitors of the Shh pathway lessen and exacerbate, respectively, the phenotypic severity of gata3 mutants. Importantly, inhibition of Shh can cause gata3 haploinsufficiency, as observed in humans. We find that gata3 mutants in a less expressive genetic background have a compensatory upregulation of Shh signaling. These results demonstrate that the level of Shh signaling can modulate the phenotypes observed in gata3 mutants.
Voronova A, Yuzwa SA, Wang BS, Zahr S, Syal C, Wang J, Kaplan DR, Miller FD.
PMID: 28472653 | DOI: 10.1016/j.neuron.2017.04.018
During development, newborn interneurons migrate throughout the embryonic brain. Here, we provide evidence that these interneurons act in a paracrine fashion to regulate developmental oligodendrocyte formation. Specifically, we show that medial ganglionic eminence (MGE) interneurons secrete factors that promote genesis of oligodendrocytes from glially biased cortical precursors in culture. Moreover, when MGE interneurons are genetically ablated in vivo prior to their migration, this causes a deficit in cortical oligodendrogenesis. Modeling of the interneuron-precursor paracrine interaction using transcriptome data identifies the cytokine fractalkine as responsible for the pro-oligodendrocyte effect in culture. This paracrine interaction is important in vivo, since knockdown of the fractalkine receptor CX3CR1 in embryonic cortical precursors, or constitutive knockout of CX3CR1, causes decreased numbers of oligodendrocyte progenitor cells (OPCs) and oligodendrocytes in the postnatal cortex. Thus, in addition to their role in regulating neuronal excitability, interneurons act in a paracrine fashion to promote the developmental genesis of oligodendrocytes.
Yao, Y;Chen, J;Li, X;Chen, ZF;Li, P;
PMID: 36750092 | DOI: 10.1016/j.cub.2023.01.019
Increased ventilation is a critical process that occurs when the body responds to a hypoxic environment. Sighs are long, deep breaths that prevent alveolar collapse, and their frequency is significantly increased by hypoxia. In this study, we first show that sighing is induced by hypoxia as a function of increased hypoxic severity and that hypoxia-induced sighing is capable of increasing the oxygen saturation in a mouse model. We next found that the gastrin-releasing peptide (Grp) expressing neurons in the nucleus of the solitary tract (NTS) are important in mediating hypoxia-induced sighing. Retrograde tracing from these Grp neurons reveals their direct afferent input from the petrosal ganglion neurons that innervate the carotid body, the major peripheral chemoreceptor that senses blood oxygen. Acute hypoxia preferentially activates these Grp neurons in the NTS. Photoactivation of these neurons through their projections in the inspiratory rhythm generator in the ventral medulla induces sighing, whereas genetic ablation or chemogenetic silencing of these neurons specifically diminishes the sighs, but not other respiratory responses, induced by hypoxia. Finally, the mice with reduced sighing in hypoxia exhibit an elevated heart-rate increase, which may compensate for maintaining the blood oxygen level. Therefore, we identified a neural circuit that connects the carotid body to the breathing control center in the ventral medulla with a specific function for hypoxia-induced sighing, which restores the oxygen level.
