scholarly journals Dopamine D2Rs Coordinate Cue-Evoked Changes in Striatal Acetylcholine Levels

2021 ◽  
Author(s):  
Kelly M. Martyniuk ◽  
Arturo Torres-Herraez ◽  
Marcelo Rubinstein ◽  
Marie A. Labouesse ◽  
Christoph Kellendonk
Keyword(s):  
Temporal Correlation ◽  
General Mechanism ◽  
Neuron Activity ◽  
Knock Out ◽  
Cholinergic Interneurons ◽  
Motivated Behavior ◽  
Mutual Regulation ◽  
Knock Out Mice

AbstractIn the striatum, acetylcholine (ACh) neuron activity is modulated co-incident with dopamine (DA) release in response to unpredicted rewards and reward predicting cues and both neuromodulators are thought to regulate each other. While this co-regulation has been studied using stimulation studies, the existence of this mutual regulation in vivo during natural behavior is still largely unexplored. One long-standing controversy has been whether striatal DA is responsible for the induction of the cholinergic pause or whether D2R modulate a pause that is induced by other mechanisms. Here, we used genetically encoded sensors in combination with pharmacological and genetic inactivation of D2Rs from cholinergic interneurons (CINs) to simultaneously measure ACh and DA levels after CIN D2R inactivation. We found that CIN D2Rs are not necessary for the induction of cue induced dips in ACh levels but regulate dip lengths and rebound ACh levels. Importantly, D2R inactivation strongly decreased the temporal correlation between DA and Ach signals not only at cue presentation but also during the intertrial interval. This points to a general mechanism by which D2Rs coordinate both signals. At the behavioral level D2R antagonism increased the latency to lever press, which was not observed in CIN-selective D2R knock out mice. This latency correlated with the cue evoked dip length supporting a role of the ACh dip and it’s regulation by D2Rs in motivated behavior. Overall, our data indicate that striatal DA coordinate phasic ACh and DA signals via CIN D2Rs which is important for the regulation of motivated behavior.

Carboxypeptidase U (TAFIa): a metallocarboxypeptidase with a distinct role in haemostasis and a possible risk factor for thrombotic disease

2005 ◽  
Vol 94 (09) ◽  
pp. 471-487 ◽  
Author(s):  
Judith Leurs ◽  
Dirk Hendriks
Keyword(s):  
Bleeding Tendency ◽  
Knock Out ◽  
Lysine Residues ◽  
Knock Out Mice ◽  
The Moment ◽  
And Function ◽  
Coagulation And Fibrinolysis

SummarySince the discovery of Carboxypeptidase U (CPU) in 1988, considerable information has been gathered about its biochemistry and function in physiological and pathophysiological circumstances. A variety of tools such as assays to measure proCPU and CPU, antibodies raised against (pro)CPU, selective CPU inhibitors and knock-out mice have been developed and are currently being used to explore the role of this metallocarboxypeptidase in different in vivo and in vitro settings. The knowledge that proCPU can be activated by thrombin and plasmin, enzymes with a key function in coagulation and fibrinolysis, and the ability of CPU to remove C-terminal lysine residues has led to the hypothesis that the proCPU/CPU pathway plays a role in the balance between coagulation and fibrinolysis. The maintenance of the equilibrium between coagulation and fibrinolysis is crucial for normal haemostasis and disturbance of this delicate balance can lead either to bleeding tendency or thrombosis. This review provides an update on several aspects of CPU known at the moment, including an extensive overview on the clinical studies performed up till now.J. Leurs is a research assistant of the Fund for Scientific Research Flanders (FWO-Vlaanderen).

Are all granzymes cytotoxic in vivo?

2014 ◽  
Vol 395 (2) ◽  
pp. 181-202 ◽  
Author(s):  
Lars T. Joeckel ◽  
Phillip I. Bird
Keyword(s):  
Serine Proteases ◽  
Ex Vivo ◽  
Cytotoxic Lymphocytes ◽  
Cytotoxic Effects ◽  
Knock Out ◽  
Granzyme A ◽  
Knock Out Mice

Abstract Granzymes are serine proteases mainly found in cytotoxic lymphocytes. The most-studied member of this group is granzyme B, which is a potent cytotoxin that has set the paradigm that all granzymes are cyototoxic. In the last 5 years, this paradigm has become controversial. On one hand, there is a plethora of sometimes contradictory publications showing mainly caspase-independent cytotoxic effects of granzyme A and the so-called orphan granzymes in vitro. On the other hand, there are increasing numbers of reports of granzymes failing to induce cell death in vitro unless very high (potentially supra-physiological) concentrations are used. Furthermore, experiments with granzyme A or granzyme M knock-out mice reveal little or no deficit in their cytotoxic lymphocytes’ killing ability ex vivo, but indicate impairment in the inflammatory response. These findings of non-cytotoxic effects of granzymes challenge dogma, and thus require alternative or additional explanations to be developed of the role of granzymes in defeating pathogens. Here we review evidence for granzyme cytotoxicity, give an overview of their non-cytotoxic functions, and suggest technical improvements for future investigations.

Abstract W P365: Critical Role of Smooth Muscle Cell Phenotypic Modulation in Cerebral Aneurysm Formation and Rupture

Stroke ◽  
2014 ◽  
Vol 45 (suppl_1) ◽  
Author(s):  
Robert M Starke ◽  
Muhammad S Ali ◽  
Nohra Chalouhi ◽  
Pascal M Jabbour ◽  
Stavropoula I Tjoumakaris ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Carotid Arteries ◽  
Critical Role ◽  
Phenotypic Modulation ◽  
Knock Out ◽  
Aneurysm Formation ◽  
Tnf Α ◽  
Knock Out Mice

Objective: Little is known about smooth muscle cell (SMC) phenotypic modulation in the cerebral circulation or pathogenesis of intracranial aneurysms. TNF-α has been associated with aneurysms, but a direct role has not been established. Methods: Cultured cerebral SMC were treated with TNF-α for PCR, western blot, chromatin immune-precipitation (CHIP), and adenovirus promoter transfection. In vivo experiments were carried out in the following models: application of TNF-α to the surface of carotid arteries, cerebral model of hypertension and hemodynamic stress, and cerebral model of aneurysm formation and rupture. The TNF-α inhibitor 3,6’dithiothalidomide (DTH) was synthesized. Results: Cultured cerebral SMC over-expressing myocardin induced expression of key SMC contractile genes (SM-α-actin, SM-22α, SM-MHC), while dominant negative suppressed expression. TNF-α treatment inhibited this contractile phenotype and induced pro-inflammatory genes (MCP-1, MMPs, VCAM-1, IL-1β). TNF-α increased expression of KLF4 and KLF4 siRNA abrogated TNF-α induced phentotypic modulation. These mechanisms were confirmed in vivo following exposure of rat carotid arteries to TNF-α and early in a model of cerebral hypertension and hemodynamic stress prior to cerebral aneurysm formation. Treatment with DTH reversed these pathological vessel wall alterations. TNF-α knock-out mice and DTH pre-treatment decreased the incidence of aneurysm formation and rupture. As compared with sham mice, TNF-α expression was not significantly different in TNF-α knock-out mice or those pre-treated with DTH, but was elevated in unruptured and ruptured aneurysms. Initiation of DTH 7 days after aneurysm induction did not alter aneurysm incidence, but resulted in stabilization and decreased rupture. CHIP assays in vivo and in vitro demonstrated that TNF-α promotes epigenetic changes through KLF4 dependent alterations in promoter regions of myocardin, SMC’s, and inflammatory genes. Conclusion: TNF-α induces phenotypic modulation of cerebral SMC through myocardin and KLF4 regulated pathways. These results demonstrate a novel role for TNF-α in promoting a pro-inflammatory phenotype. These data suggests a critical role of TNF-α in the formation and rupture of aneurysms.

scholarly journals The role of P2Y12 in the kinetics of microglial self-renewal and maturation in the adult visual cortex in vivo

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Monique Mendes ◽  
Linh Le ◽  
Jason Atlas ◽  
Zachary Brehm ◽  
Antonio Ladron-de-Guevara ◽  
...  
Keyword(s):  
Immune Cells ◽  
Self Renewal ◽  
Knock Out ◽  
Two Photon ◽  
And Migration ◽  
Knock Out Mice ◽  
Morphological Maturation ◽  
Kinetics Of

Microglia are the brain's resident immune cells with a tremendous capacity to autonomously self-renew. Because microglial self-renewal has largely been studied using static tools, its mechanisms and kinetics are not well understood. Using chronic in vivo two-photon imaging in awake mice, we confirm that cortical microglia show limited turnover and migration under basal conditions. Following depletion, however, microglial repopulation is remarkably rapid and is sustained by the dynamic division of remaining microglia, in a manner that is largely independent of signaling through the P2Y12 receptor. Mathematical modeling of microglial division demonstrates that the observed division rates can account for the rapid repopulation observed in vivo. Additionally, newly-born microglia resemble mature microglia within days of repopulation, although morphological maturation is different in newly born microglia in P2Y12 knock out mice. Our work suggests that microglia rapidly locally and that newly-born microglia do not recapitulate the slow maturation seen in development but instead take on mature roles in the CNS.

scholarly journals Combined loss of proapoptotic genes Bak or Bax with Bim synergizes to cause defects in hematopoiesis and in thymocyte apoptosis

2005 ◽  
Vol 201 (12) ◽  
pp. 1949-1960 ◽  
Author(s):  
Jack Hutcheson ◽  
John C. Scatizzi ◽  
Emily Bickel ◽  
Nathaniel J. Brown ◽  
Philippe Bouillet ◽  
...  
Keyword(s):  
Apoptotic Pathway ◽  
Knock Out ◽  
Bh3 Domain ◽  
Thymocyte Apoptosis ◽  
B Lymphoid ◽  
Knock Out Mice ◽  
Deficient Mice ◽  
Single Knockout

The proapoptotic members of the Bcl-2 family can be subdivided into members that contain several Bcl-2 homology (BH) domains and those that contain only the BH3 domain. Although it is known that BH3-only proteins and the multi-BH domain proteins, Bak and Bax, are essential for programmed cell death, the overlapping role of these two subgroups has not been examined in vivo. To investigate this, we generated Bak/Bim and Bax/Bim double deficient mice. We found that although Bax−/−Bim−/−, but not Bak−/−Bim−/−, mice display webbed hind and front paws and malocclusion of the incisors, both groups of mice present with dysregulated hematopoiesis. Combined loss of Bak and Bim or Bax and Bim causes defects in myeloid and B-lymphoid development that are more severe than those found in the single knock-out mice. Bak−/−Bim−/− mice have a complement of thymocytes that resembles those in control mice, whereas Bax−/−Bim−/− mice are more similar to Bim−/− mice. However, thymocytes isolated from Bak−/−Bim−/− or Bax−/−Bim−/− mice are markedly more resistant to apoptotic stimuli mediated by the intrinsic pathway as compared with thymocytes from single-knockout mice. These data suggest an essential overlapping role for Bak or Bax and Bim in the intrinsic apoptotic pathway.

Transgenic mice studies demonstrate a role for platelet factor 4 in thrombosis: dissociation between anticoagulant and antithrombotic effect of heparin

Blood ◽  
2004 ◽  
Vol 104 (10) ◽  
pp. 3173-3180 ◽  
Author(s):  
Don E. Eslin ◽  
Chunyan Zhang ◽  
Kathleen J. Samuels ◽  
Lubica Rauova ◽  
Li Zhai ◽  
...  
Keyword(s):  
Transgenic Mice ◽  
Thrombus Formation ◽  
Platelet Factor 4 ◽  
Platelet Factor ◽  
Knock Out ◽  
Thrombosis Model ◽  
Knock Out Mice ◽  
Positively Charged

Abstract The platelet-specific chemokine platelet factor 4 (PF4) is released in large amounts at sites of vascular injury. PF4 binds to heparin with high affinity, but its in vivo biologic role has not been defined. We studied the role of PF4 in thrombosis using heterozygote and homozygote PF4 knock-out mice (mPF4+/– and mPF4–/–, respectively) and transgenic mice overexpressing human PF4 (hPF4+). None of these lines had an overt bleeding diathesis, but in a FeCl3 carotid artery thrombosis model, all showed impaired thrombus formation. This defect in thrombus formation in the mPF4–/– animals was corrected by infusing hPF4 over a narrow concentration range. The thrombotic defect in the mPF4+/– and mPF4–/– animals was particularly sensitive to infusions of the negatively charged anticoagulant heparin. However, the same amount of heparin paradoxically normalized thrombus formation in the hPF4+ animals, although these animals were anticoagulated systemically. Upon infusion of the positively charged protein, protamine sulfate, the reverse was observed with mPF4+/– and mPF4–/– animals having improved thrombosis, with the hPF4+ animals having worsened thrombus formation. These studies support an important role for PF4 in thrombosis, and show that neutralization of PF4 is an important component of heparin's anticoagulant effect. The mechanisms underlying these observations of PF4 biology and their clinical implications remain to be determined.

scholarly journals A New Mouse Model for Complete Congenital Stationary Night Blindness Due to Gpr179 Deficiency

2021 ◽  
Vol 22 (9) ◽  
pp. 4424
Author(s):  
Elise Orhan ◽  
Marion Neuillé ◽  
Miguel de Sousa Dias ◽  
Thomas Pugliese ◽  
Christelle Michiels ◽  
...  
Keyword(s):  
Mouse Model ◽  
Night Blindness ◽  
Signal Transmission ◽  
Bipolar Cells ◽  
Congenital Stationary Night Blindness ◽  
Knock Out ◽  
Intron 1 ◽  
Knock Out Mice

Mutations in GPR179 lead to autosomal recessive complete congenital stationary night blindness (cCSNB). This condition represents a signal transmission defect from the photoreceptors to the ON-bipolar cells. To confirm the phenotype, better understand the pathogenic mechanism in vivo, and provide a model for therapeutic approaches, a Gpr179 knock-out mouse model was genetically and functionally characterized. We confirmed that the insertion of a neo/lac Z cassette in intron 1 of Gpr179 disrupts the same gene. Spectral domain optical coherence tomography reveals no obvious retinal structure abnormalities. Gpr179 knock-out mice exhibit a so-called no-b-wave (nob) phenotype with severely reduced b-wave amplitudes in the electroretinogram. Optomotor tests reveal decreased optomotor responses under scotopic conditions. Consistent with the genetic disruption of Gpr179, GPR179 is absent at the dendritic tips of ON-bipolar cells. While proteins of the same signal transmission cascade (GRM6, LRIT3, and TRPM1) are correctly localized, other proteins (RGS7, RGS11, and GNB5) known to regulate GRM6 are absent at the dendritic tips of ON-bipolar cells. These results add a new model of cCSNB, which is important to better understand the role of GPR179, its implication in patients with cCSNB, and its use for the development of therapies.

scholarly journals Topoisomerase II is regulated by translationally controlled tumor protein for cell survival during organ growth in Drosophila

2021 ◽  
Vol 12 (9) ◽  
Author(s):  
Dae-Wook Yang ◽  
Jung-Wan Mok ◽  
Stephanie B. Telerman ◽  
Robert Amson ◽  
Adam Telerman ◽  
...  
Keyword(s):  
Cell Survival ◽  
Topoisomerase Ii ◽  
Wing Disc ◽  
Organ Development ◽  
Translationally Controlled Tumor Protein ◽  
Protein Levels ◽  
Eye Disc ◽  
Mutual Regulation

AbstractRegulation of cell survival is critical for organ development. Translationally controlled tumor protein (TCTP) is a conserved protein family implicated in the control of cell survival during normal development and tumorigenesis. Previously, we have identified a human Topoisomerase II (TOP2) as a TCTP partner, but its role in vivo has been unknown. To determine the significance of this interaction, we examined their roles in developing Drosophila organs. Top2 RNAi in the wing disc leads to tissue reduction and caspase activation, indicating the essential role of Top2 for cell survival. Top2 RNAi in the eye disc also causes loss of eye and head tissues. Tctp RNAi enhances the phenotypes of Top2 RNAi. The depletion of Tctp reduces Top2 levels in the wing disc and vice versa. Wing size is reduced by Top2 overexpression, implying that proper regulation of Top2 level is important for normal organ development. The wing phenotype of Tctp RNAi is partially suppressed by Top2 overexpression. This study suggests that mutual regulation of Tctp and Top2 protein levels is critical for cell survival during organ development.

scholarly journals Loss of microglial SIRPα promotes synaptic pruning in preclinical models of neurodegeneration

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Xin Ding ◽  
Jin Wang ◽  
Miaoxin Huang ◽  
Zhangpeng Chen ◽  
Jing Liu ◽  
...  
Keyword(s):  
Alzheimer’S Disease ◽  
Alzheimer's Disease ◽  
Knock Out ◽  
Classical Complement Pathway ◽  
Synaptic Remodeling ◽  
The Central Nervous System ◽  
System Activation ◽  
Knock Out Mice ◽  
Synaptic Pruning

AbstractMicroglia play a key role in regulating synaptic remodeling in the central nervous system. Activation of classical complement pathway promotes microglia-mediated synaptic pruning during development and disease. CD47 protects synapses from excessive pruning during development, implicating microglial SIRPα, a CD47 receptor, in synaptic remodeling. However, the role of microglial SIRPα in synaptic pruning in disease remains unclear. Here, using conditional knock-out mice, we show that microglia-specific deletion of SIRPα results in decreased synaptic density. In human tissue, we observe that microglial SIRPα expression declines alongside the progression of Alzheimer’s disease. To investigate the role of SIRPα in neurodegeneration, we modulate the expression of microglial SIRPα in mouse models of Alzheimer’s disease. Loss of microglial SIRPα results in increased synaptic loss mediated by microglia engulfment and enhanced cognitive impairment. Together, these results suggest that microglial SIRPα regulates synaptic pruning in neurodegeneration.

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