scholarly journals Chemogenetic activation of astrocytes in the hippocampus and cortex changes the transcriptome of microglia and other cell types

2020 ◽  
Author(s):  
Stéphanie Philtjens ◽  
Marion T. Turnbull ◽  
Brian P. Thedy ◽  
Younghye Moon ◽  
Jungsu Kim
Keyword(s):  
Single Cell ◽  
G Protein ◽  
Low Density Lipoprotein ◽  
Ion Homeostasis ◽  
Density Lipoprotein ◽  
Cell Types ◽  
Designer Drugs ◽  
Specific Cell ◽  
G Protein Coupled

AbstractAstrocytes are the most common glial cell type in the brain, yet, it is still not clear how their activation affects the transcriptome of other brain cells such as microglia and neurons. Engineered G protein-coupled receptors called Designer Receptors Exclusively Activated by Designer Drugs (DREADDS) make it possible to selectively activate specific cell types, such as neurons and astrocytes. By combining the selective activation of astrocytes with single cell RNA sequencing, we were able to study transcriptional changes that occur in response to the activation of astrocytes at the single cell level. Interestingly, our data shows that long-term activation of astrocytes in healthy mice results in dramatic alteration in the transcriptome of astrocytes and microglia. Genes that were differentially expressed in these Gq-DREADD-activated astrocytes were involved in neurogenesis and low density lipoprotein particle biology, while those in the microglia were involved in the response to lipoproteins, and the migration and chemotaxis of immune cells. Furthermore, network analysis showed that Gq-DREADD-mediated activation in astrocytes resulted in an upregulation of genes involved in the G protein-coupled receptor signaling pathway and calcium ion homeostasis. This confirmed the activation of astrocytes through the expressed DREADDS. Our findings show the importance of considering the transcriptomic alteration in microglia and neurons after the activation of astrocytes in in vivo models. Therefore, our data will serve as a resource for the broader neuroscience community.

scholarly journals Chemogenetic Tools for Causal Cellular and Neuronal Biology

2018 ◽  
Vol 98 (1) ◽  
pp. 391-418 ◽  
Author(s):  
Deniz Atasoy ◽  
Scott M. Sternson
Keyword(s):  
G Protein ◽  
Ex Vivo ◽  
Cell Types ◽  
Cell Populations ◽  
Specific Cell ◽  
Cellular Processes ◽  
Distinct Cell ◽  
G Protein Coupled ◽  
Pharmacological Control

Chemogenetic technologies enable selective pharmacological control of specific cell populations. An increasing number of approaches have been developed that modulate different signaling pathways. Selective pharmacological control over G protein-coupled receptor signaling, ion channel conductances, protein association, protein stability, and small molecule targeting allows modulation of cellular processes in distinct cell types. Here, we review these chemogenetic technologies and instances of their applications in complex tissues in vivo and ex vivo.

The G Protein-Coupled Receptor cysLT1 Mediates Gi Protein-Dependent Chemotaxis and Adhesion of CD34+ Cells in Vitro Similar to CXCR4, but Not Progenitor Mobilization in Vivo.

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 1388-1388
Author(s):  
Adriana Drost ◽  
Lena Jaggy ◽  
Lothar Kanz ◽  
Robert Möhle
Keyword(s):  
Bone Marrow ◽  
G Protein ◽  
Actin Polymerization ◽  
Cell Types ◽  
Mediated Effects ◽  
Cd34 Cells ◽  
Gi Protein ◽  
G Protein Coupled

Abstract CysLT1, which is expressed in several cell types during inflammation and allergy, and the homing related chemokine receptor CXCR4 both belong to the family of G protein-coupled receptors (GPCR). Similar to CXCR4, the cysteinyl-leukotriene receptor cysLT1 is strongly expressed in CD34+ cell lines and CD34+ hematopoietic progenitor cells (HPC). We therefore compared the effects mediated by cysLT1 on HPC to those observed after activation of CXCR4. The most potent cysLT1 ligand LTD4 induced chemotaxis and adhesion of CD34+ HPC to endothelial cells, immobilized VCAM-1 and fibronectin, which was comparable to effects induced by the CXCR4 ligand SDF-1. CysLT1- and CXCR4-mediated effects were inhibited by pertussis toxin (PTX), suggesting that both GPCRs employ the same Gi-protein-dependent signaling pathways in CD34+ HPC. This is supported by identical time courses of intracellular calcium fluxes and actin polymerization induced by LTD4 and SDF-1, as measured by time-dependent flow cytometry. Given the striking similarities of cysLT1- and CXCR4-mediated effects in vitro, one might expect also overlapping functions in vivo. We therefore investigated whether blocking of cysLT1 is associated with HPC mobilization. As cysLT1 antagonists are established for therapy and prophylaxis in patients with allergic and exercise-induced asthma, circulating CD34+ progenitors were enumerated after initiation of a treatment with the cysLT1 antagonist montelukast, used as asthma prophylaxis in otherwise healthy subjects. The number of CD34+ cells or white blood counts did not differ significantly from the baseline value 2, 4, 8, 12, and 24 hours after administration of 10 mg montelukast, in contrast to previous studies analyzing stem cell mobilization induced by CXCR4 antagonists. This corresponds with our in vitro findings that LTD4 is produced by bone marrow endothelium and stromal cells only when deprived of hematopoietic cells, in contrast to the constitutive production of SDF-1. We conclude that cysLT1 is not involved in bone marrow retention of HPC during steady-state hematopoiesis, but may modulate HPC homing when its ligands are produced either locally (i.e. bone marrow aplasia) or systemically (i.e. inflammation).

High Levels of Acetylated Low Density Lipoprotein Uptake and Low Tie2 Promoter Activity Distinguishes Sinusoids from Other Vessel Types in the Murine Bone Marrow

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 5465-5465
Author(s):  
Xiaomiao Li ◽  
Zhongbo Hu ◽  
Marda Jorgenson ◽  
William Slayton
Keyword(s):  
Bone Marrow ◽  
Blood Vessels ◽  
Low Density Lipoprotein ◽  
Density Lipoprotein ◽  
Cell Types ◽  
Blood Stream ◽  
Low Density ◽  
Murine Bone Marrow ◽  
Different Types

Abstract Background: The bone marrow contains a variety of blood vessels that have different functions in maintaining the bone marrow as the major blood producing organ in adulthood. For instance, arterioles function to control the flow of blood into bone marrow compartments, and the sinusoids serve as a conduit to the blood stream and niches for megakaryocyte development. Most current studies of the bone marrow vasculature, including studies quantifying changes in the marrow vascular by microvascular density, do not differentiate between different types of marrow vessels. Recognizing the changes in different types of blood vessels has important physiologic implications. Here we report a new method to distinguish sinusoids from arterioles in the murine bone marrow. Methods and Results: We used transgenic mice with GFP expressed downstream of the Tie-2 promoter, combined with in vivo acetylated low-density lipoprotein (Ac-LDL) uptake method to differentiate sinusoids from arterioles. We found that Ac-LDL was specifically endocytosed by sinusoids, and Tie-2 expression was more pronounced in the arteries, arterioles, and transitional capillaries. Combining these two functional endothelial markers and using confocal microscopy to obtain three dimensional images, we identified transitional zones where arterioles emptied into the sinusoids. Conclusions: These results demonstrate that the marrow vasculature and specific endothelial cell types are functionally heterogeneous. Methods to study changes in the marrow vasculature and particularly the vascular niche, a function of sinusoids, need to take into account this heterogeneity.

G-protein-coupled designer receptors – new chemical-genetic tools for signal transduction research

2013 ◽  
Vol 394 (12) ◽  
pp. 1615-1622 ◽  
Author(s):  
Gerald Thiel ◽  
Anke Kaufmann ◽  
Oliver G. Rössler
Keyword(s):  
Signal Transduction ◽  
Signaling Pathway ◽  
G Protein ◽  
Membrane Receptors ◽  
Designer Drugs ◽  
Gpcr Signaling ◽  
Chemical Genetic ◽  
G Protein Coupled

Abstract G-protein-coupled receptors (GPCRs) are the largest group of plasma membrane receptors in nature and are activated by a variety of different ligands. The biological outcome of GPCR stimulation is complex, as a plethora of signaling pathways are activated upon stimulation. These complexity and diversity of GPCR signaling make it difficult to manipulate the signaling pathway of a specific GPCR by natural ligands. To reduce the complexity in experimental settings, specific pharmacological ligands that preferentially activate one signaling pathway have been developed. In addition, G-protein-coupled designer receptors that are unresponsive to endogenous ligands but can be activated by otherwise pharmacologically inert compounds have been designed. These receptors have been termed designer receptors exclusively activated by designer drugs. The lack of constitutive activity of these designer receptors allows their use for in vitro and in vivo studies of GPCR-mediated signal transduction. The analysis of recently generated transgenic mice showed that the expression of G-protein-coupled designer receptors represents a powerful chemical-genetic tool to investigate GPCR signaling and function.

Engineered GPCRs as Tools to Modulate Signal Transduction

Physiology ◽  
2008 ◽  
Vol 23 (6) ◽  
pp. 313-321 ◽  
Author(s):  
Ying Pei ◽  
Sarah C. Rogan ◽  
Feng Yan ◽  
Bryan L. Roth
Keyword(s):  
Signal Transduction ◽  
Signaling Pathways ◽  
G Protein ◽  
G Protein Coupled Receptors ◽  
Designer Drugs ◽  
Modulate Signal ◽  
Synthetic Drug ◽  
G Protein Coupled

Different families of G-protein-coupled receptors (GPCRs) have been engineered to provide exclusive control over the activation of these receptors and thus to understand better the consequences of their signaling in vitro and in vivo. These engineered receptors, named RASSLs (receptors activated solely by synthetic ligands) and DREADDs (designer receptors exclusively activated by designer drugs), are insensitive to their endogenous ligands but can be activated by synthetic drug-like compounds. Currently, the existing RASSLs and DREADDs cover the Gi, Gq, and Gs signaling pathways. These modified GPCRs can be utilized as ideal tools to study GPCR functions selectively in specific cellular populations.

scholarly journals Lipoprotein lipase stimulates the binding and uptake of moderately oxidized low-density lipoprotein by J774 macrophages

1996 ◽  
Vol 314 (2) ◽  
pp. 563-568 ◽  
Author(s):  
Wendy L. HENDRIKS ◽  
Hans van der BOOM ◽  
Leonie C. van VARK ◽  
Louis M. HAVEKES
Keyword(s):  
Lipoprotein Lipase ◽  
Oxidized Ldl ◽  
Low Density Lipoprotein ◽  
Heparan Sulphate ◽  
Density Lipoprotein ◽  
Cell Types ◽  
Low Density ◽  
Dependent Manner ◽  
Dose Dependent

Lipoprotein lipase (LPL) stimulates the uptake of low-density lipoprotein (LDL) and very-low-density lipoprotein (VLDL) in different cell types, including macrophages, through bridging of LPL between lipoproteins and extracellular heparan sulphate proteoglycans (HSPG). Because macrophages produce LPL and because modified lipoproteins are present in the arterial wall in vivo, we wondered whether LPL also enhances the uptake of oxidized LDL by J774 macrophages. LDL samples with different degrees of oxidation, as evaluated by relative electrophoretic mobility (REM) as compared with native LDL are used, as well as native and acetylated LDL. Addition of 5 μg/ml LPL to the J774 cell culture medium stimulated the binding of both native LDL and moderately oxidized LDL (REM < 3.5) 50–100-fold, and their uptake was stimulated approx. 20-fold. The LPL-mediated binding of native LDL and moderately oxidized LDL was dose-dependent. Preincubation of the cells with heparinase (2.4 units/ml) inhibited the stimulatory effect of LPL, indicating that this LPL-mediated stimulation was due to bridging between the lipoproteins and HSPG. The binding to J774 macrophages of severely oxidized LDL (REM = 4.3) was stimulated less than 3-fold by LPL, whereas its uptake was not stimulated significantly. The binding and uptake of acetylated LDL (AcLDL) were not stimulated by LPL, although the LPL-molecule itself does bind to AcLDL. Measurements of the cellular lipid content showed that addition of LPL also stimulated the accumulation in the cells of cholesteryl ester derived from both native LDL and moderately oxidized LDL in a dose-dependent manner. We conclude that our results present experimental evidence for the hypothesis that LPL serves as an atherogenic component in the vessel wall.

scholarly journals The puzzling uniqueness of the heterotrimeric G15 protein and its potential beyond hematopoiesis

2010 ◽  
Vol 44 (5) ◽  
pp. 259-269 ◽  
Author(s):  
Flavia Giannone ◽  
Giorgio Malpeli ◽  
Veronica Lisi ◽  
Silvia Grasso ◽  
Priyanka Shukla ◽  
...  
Keyword(s):  
G Protein ◽  
G Proteins ◽  
Physiological Role ◽  
Cell Types ◽  
Biochemical Properties ◽  
Membrane Receptors ◽  
General Mechanism ◽  
Specific Cell ◽  
Α Subunit ◽  
G Protein Coupled

Heterotrimeric G proteins transduce the signals of the largest family of membrane receptors (G protein-coupled receptors, GPCRs) hence triggering the activation of a wide variety of physiological responses. G15 is a G protein characterized by a number of functional peculiarities that make its signaling exceptional: 1) it can couple a variety of Gs-, Gi/o-, and Gq-linked receptors to phospholipase C activation; 2) relatively to other G proteins, it is poorly affected by β-arrestin-dependent desensitization, the general mechanism that regulates GPCR function and 3) at the protein level, its expression is only detected in highly specific cell types (hematopoietic and epithelial cells). G15 α-subunit displays unique structural and biochemical properties, and is phylogenetically the most recent and divergent component of the Gαq/11 subfamily. All these aspects shed a mysterious light on G15 biological role, which remains substantially elusive. Thus, far, G15 signaling has been analyzed in the context of hematopoiesis. Here, we highlight observations supporting the view that G15 functions may extend further beyond the immune system. In addition, we describe puzzling aspects of G15 signaling that offer a novel perspective in the understanding of its physiological role.

scholarly journals Quantitative role of parenchymal and non-parenchymal liver cells in the uptake of [14C]sucrose-labelled low-density lipoprotein in vivo

1984 ◽  
Vol 224 (1) ◽  
pp. 21-27 ◽  
Author(s):  
L Harkes ◽  
J C Van Berkel
Keyword(s):  
Low Density Lipoprotein ◽  
Density Lipoprotein ◽  
Cell Types ◽  
Liver Cells ◽  
Low Density ◽  
Apo B ◽  
Parenchymal Liver ◽  
Ldl Uptake ◽  
Parenchymal Cells

In order to assess the relative importance of the receptor for low-density lipoprotein (LDL) (apo-B,E receptor) in the various liver cell types for the catabolism of lipoproteins in vivo, human LDL was labelled with [14C]sucrose. Up to 4.5h after intravenous injection, [14C]sucrose becomes associated with liver almost linearly with time. During this time the liver is responsible for 70-80% of the removal of LDL from blood. A comparison of the uptake of [14C]sucrose-labelled LDL and reductive-methylated [14C]sucrose-labelled LDL ([14C]sucrose-labelled Me-LDL) by the liver shows that methylation leads to a 65% decrease of the LDL uptake. This indicated that 65% of the LDL uptake by liver is mediated by a specific apo-B,E receptor. Parenchymal and non-parenchymal liver cells were isolated at various times after intravenous injection of [14C]sucrose-labelled LDL and [14C]sucrose-labelled Me-LDL. Non-parenchymal liver cells accumulate at least 60 times as much [14C]sucrose-labelled LDL than do parenchymal cells accumulate at least 60 times as much [14C]sucrose-labelled LDL than do parenchymal cells when expressed per mg of cell protein. This factor is independent of the time after injection of LDL. Taking into account the relative protein contribution of the various liver cell types to the total liver, it can be calculated that non-parenchymal cells are responsible for 71% of the total liver uptake of [14C]sucrose-labelled LDL. A comparison of the cellular uptake of [14C]sucrose-labelled LDL and [14C]sucrose-labelled Me-LDL after 4.5h circulation indicates that 79% of the uptake of LDL by non-parenchymal cells is receptor-dependent. With parenchymal cells no significant difference in uptake between [14C]sucrose-labelled LDL and [14C]sucrose-labelled Me-LDL was found. A further separation of the nonparenchymal cells into Kupffer and endothelial cells by centrifugal elutriation shows that within the non-parenchymal-cell preparation solely the Kupffer cells are responsible for the receptor-dependent uptake of LDL. It is concluded that in rats the Kupffer cell is the main cell type responsible for the receptor-dependent catabolism of lipoproteins containing only apolipoprotein B.

G-protein-coupled receptors, channels, and Na+–H+ exchanger in nuclear membranes of heart, hepatic, vascular endothelial, and smooth muscle cellsThis paper is one of a selection of papers published in this Special Issue, entitled The Nucleus: A Cell Within A Cell.

2006 ◽  
Vol 84 (3-4) ◽  
pp. 431-441 ◽  
Author(s):  
Ghassan Bkaily ◽  
Moni Nader ◽  
Levon Avedanian ◽  
Sana Choufani ◽  
Danielle Jacques ◽  
...  
Keyword(s):  
Angiotensin Ii ◽  
G Protein ◽  
Cell Types ◽  
G Protein Coupled Receptors ◽  
Specific Cell ◽  
Endothelin 1 ◽  
Nuclear Membranes ◽  
Ionic Transporters ◽  
A Cell ◽  
G Protein Coupled

The action of several peptides and drugs is thought to be primarily dependent on their interactions with specific cell surface G-protein-coupled receptors and ionic transporters such as channels and exchangers. Recent development of 3-D confocal microscopy allowed several laboratories, including ours, to identify and study the localization of receptors, channels, and exchangers at the transcellular level of several cell types. Using this technique, we demonstrated in the nuclei of several types of cells the presence of Ca2+ channels as well as Na+–H+ exchanger and receptors such as endothelin-1 and angiotensin II receptors. Stimulation of these nuclear membrane G-protein-coupled receptors induced an increase of nuclear Ca2+. Our results suggest that, similar to the plasma membrane, nuclear membranes possess channels, exchangers and receptors such as those for endothelin-1 and angiotensin II, and that the nucleus seems to be a cell within a cell. This article will emphasize these findings.

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