L'aide contrainte dans le domaine de la protection de l'enfance

L’aide contrainte concerne plusieurs champs du travail social, dont celui de la protection de l’enfance. Le travailleur social de l’Office pour la protection de l’enfant (OPE) va intervenir auprès de personnes réticentes à son aide, mais avec qui pourtant il va créer une relation. Comment est-il possible d’entrer en relation avec des personnes opposées à cela ? Est-il possible d’intervenir sans relation ? Nous verrons que le pouvoir que détient le professionnel a un impact sur la relation avec le bénéficiaire. D’ailleurs, qui est le bénéficiaire ? L’enfant ou le parent ? Intervenir en contexte d’aide contrainte est-il vécu de manière négative par les professionnels ? Existerait-il également du positif ? La thématique de l’aide contrainte se trouve ainsi au coeur de ce travail de Bachelor

Extracellular and Intracellular Regulation of G Protein-Coupled Receptor Signaling

Signaling through G protein-coupled receptors (GPCRs) is an essential part of cellular communication, and nearly half of modern pharmaceuticals target GPCRs in some way. GPCRs are also subject to extensive biological regulation, which is incompletely understood. Here, I examined both extracellular metabolic control of GPCR signaling, and intracellular feedback mechanisms which regulate downstream signal transduction and receptor desensitization. I first found that the nucleotide adenosine 5'-monophosphate (AMP) is an agonist of the A1 adenosine receptor. Previously, AMP was thought to signal exclusively via hydrolysis to adenosine, and no AMP receptor was known to exist. Using a novel real-time assay of adenosine receptor activity, I showed that AMP directly activates A1R independent of hydrolysis to adenosine, but that activation of the adenosine A2B receptor required hydrolysis to adenosine. I also identified a histidine residue in the A1R binding pocket critical for receptor activation by AMP, but not adenosine. These results suggest that some of the A1R-mediated physiological effects attributed to adenosine may in fact be directly caused by AMP. Furthermore, I found that extracellular loop 2 partially determines A1R sensitivity to AMP, and that AMP stimulation elicits differential activation of signaling cascades downstream of A1R. In subsequent work, I found that the novel lipid kinase diacylglycerol kinase eta (DGKh) positively modulates signaling downstream of muscarinic and purinergic GPCRs. This effect required DGKh catalytic activity, but only a minimal level of DGKh expression. DGKh expression also suppressed extracellular signal-related kinase (ERK) phosphorylation downstream of protein kinase C (PKC) and both activation and depletion of PKC abolished the DGKh effect on GPCR signaling, indicating that DGKh enhances GPCR signaling by suppressing PKC activation, likely reducing receptor desensitization. Enhanced expression of DGKh is linked to bipolar disorder (BPD), suggesting that increased DGKh activity contributes to the dysregulation of GPCR and PKC signaling in BPD and that DGKh inhibitors may have therapeutic potential for the treatment of BPD. Together, my work expands the current understanding of adenosine receptor signaling, implies an enhanced role for multiple endogenous ligands and functional selectivity in physiological GPCR signaling, and highlights GPCR-modulating enzymes as targets for drug discovery.Doctor of Philosoph

Overexpression of Diacylglycerol Kinase Enhances G q-Coupled G Protein-Coupled Receptor Signaling

Multiple genome-wide association studies have linked diacylglycerol kinas

Incidencia de la Rugosidad de un Rodillo y otros Factores en el Cálculo de Incertidumbre en la Calibración de un Frenómetro

El calibrado de un frenómetro consiste en comparar la fuerza de frenado con un patrón de fuerza conocido y evaluar su incertidumbre como medida de la calidad de la calibración. En una auditoria del sistema de gestión de la calidad del laboratorio Calibra Uno, bajo la dirección del Organismo Argentino de Acreditación (OAA), se determinó que la rugosidad de los rodillos del frenómetro no había sido tenida en cuenta entre los factores que influyen en la calibración del mismo. Por ende, así se investigaron todas las variables predominantes en el equipo de medición, en el mensurando, en las condiciones ambientales y en la pericia del operario. El resultado fue un modelo matemático asistido por computadora, basado en la serie de Taylor, que permite calcular la incertidumbre de medición de la fuerza de frenado, y a la cual se le puede cargar las mediciones realizadas durante la calibración de un frenómetro en el Taller de Revisión Técnica Vehicular (TRTV

Biological constraints limit the use of rapamycin-inducible FKBP12-Inp54p for depleting PIP2 in dorsal root ganglia neurons

Background: Rapamycin-induced translocation systems can be used to manipulate biological processes with precise temporal control. These systems are based on rapamycin-induced dimerization of FK506 Binding Protein 12 (FKBP12) with the FKBP Rapamycin Binding (FRB) domain of mammalian target of rapamycin (mTOR). Here, we sought to adapt a rapamycin-inducible phosphatidylinositol 4,5-bisphosphate (PIP2)-specific phosphatase (Inp54p) system to deplete PIP2 in nociceptive dorsal root ganglia (DRG) neurons. Results: We genetically targeted membrane-tethered CFP-FRBPLF (a destabilized FRB mutant) to the ubiquitously expressed Rosa26 locus, generating a Rosa26-FRBPLF knockin mouse. In a second knockin mouse line, we targeted Venus-FKBP12-Inp54p to the Calcitonin gene-related peptide-alpha (CGRPα) locus. We hypothesized that after intercrossing these mice, rapamycin treatment would induce translocation of Venus-FKBP12-Inp54p to the plasma membrane in CGRP+ DRG neurons. In control experiments with cell lines, rapamycin induced translocation of Venus-FKBP12-Inp54p to the plasma membrane, and subsequent depletion of PIP2, as measured with a PIP2 biosensor. However, rapamycin did not induce translocation of Venus-FKBP12-Inp54p to the plasma membrane in FRBPLF-expressing DRG neurons (in vitro or in vivo). Moreover, rapamycin treatment did not alter PIP2-dependent thermosensation in vivo. Instead, rapamycin treatment stabilized FRBPLF in cultured DRG neurons, suggesting that rapamycin promoted dimerization of FRBPLF with endogenous FKBP12. Conclusions: Taken together, our data indicate that these knockin mice cannot be used to inducibly deplete PIP2 in DRG neurons. Moreover, our data suggest that high levels of endogenous FKBP12 could compete for binding to FRBPLF, hence limiting the use of rapamycin-inducible systems to cells with low levels of endogenous FKBP12

AMP Is an Adenosine A 1 Receptor Agonist

Numerous receptors for ATP, ADP, and adenosine exist; however, it is currently unknown whether a receptor for the related nucleotide adenosine 5′-monophosphate (AMP) exists. Using a novel cell-based assay to visualize adenosine receptor activation in real time, we found that AMP and a non-hydrolyzable AMP analog (deoxyadenosine 5′-monophosphonate, ACP) directly activated the adenosine A1 receptor (A1R). In contrast, AMP only activated the adenosine A2B receptor (A2BR) after hydrolysis to adenosine by ecto-5′-nucleotidase (NT5E, CD73) or prostatic acid phosphatase (PAP, ACPP). Adenosine and AMP were equipotent human A1R agonists in our real-time assay and in a cAMP accumulation assay. ACP also depressed cAMP levels in mouse cortical neurons through activation of endogenous A1R. Non-selective purinergic receptor antagonists (pyridoxalphosphate-6-azophenyl-2′,4′-disulfonic acid and suramin) did not block adenosine- or AMP-evoked activation. Moreover, mutation of His-251 in the human A1R ligand binding pocket reduced AMP potency without affecting adenosine potency. In contrast, mutation of a different binding pocket residue (His-278) eliminated responses to AMP and to adenosine. Taken together, our study indicates that the physiologically relevant nucleotide AMP is a full agonist of A1R. In addition, our study suggests that some of the physiological effects of AMP may be direct, and not indirect through ectonucleotidases that hydrolyze this nucleotide to adenosine

Role of kappa-opioid receptors in the effects of salvinorin A and ketamine on attention in rats

Disruptions in perception and cognition are characteristic of psychiatric conditions such as schizophrenia. Studies of pharmacological agents that alter perception and cognition in humans might provide a better understanding of the brain substrates of these complex processes. One way to study these states in rodents is with tests that require attention and visual perception for correct performance

Role of kappa-opioid receptors in the effects of salvinorin A and ketamine on attention behavior in rats

Background: Disruptions in perception and cognition are characteristic of psychiatric conditions such as schizophrenia. Studies of pharmacological agents that alter perception and cognition in humans might provide a better understanding of the brain substrates of these complex processes. One way to study these states in rodents is with tests that require attention and visual perception for correct performance. Methods: We examined the effects of two drugs that cause disruptions in perception and cognition in humans—the kappa-opioid receptor (KOR) agonist salvinorin A (salvA; 0.125–4.0 mg/kg) and the non-competitive NMDA receptor antagonist ketamine (0.63–20 mg/kg)—on behavior in rats using the 5-choice serial reaction time task (5CSRTT), a food-motivated test that quantifies attention. We also compared the binding profiles of salvA and ketamine at KORs and NMDA receptors. Results: SalvA and ketamine produced the same pattern of disruptive effects in the 5CSRTT, characterized by increases in signs often associated with reduced motivation (omission errors) and deficits in processing (elevated latencies to respond correctly). Sessions in which rats were fed before testing suggest that reduced motivation produces a subtly different pattern of behavior. Pretreatment with the KOR antagonist JDTic (10 mg/kg) blocked all salvA effects and some ketamine effects. Binding and function studies revealed that ketamine is a full agonist at KORs, although not as potent or selective as salvA. Conclusions: SalvA and ketamine have previously under-appreciated similarities in their behavioral effects and pharmacological profiles. By implication, KORs might be involved in some of the cognitive abnormalities observed in psychiatric disorders such as schizophrenia

Orally Active Adenosine A 1 Receptor Agonists with Antinociceptive Effects in Mice

Adenosine A1 receptor (A1AR) agonists have antinociceptive effects in multiple preclinical models of acute and chronic pain. Although numerous A1AR agonists have been developed, clinical applications of these agents have been hampered by their cardiovascular side effects. Herein we report a series of novel A1AR agonists, some of which are structurally related to adenosine 5′-monophosphate (5′-AMP), a naturally occurring nucleotide that itself activates A1AR. These novel compounds potently activate A1AR in several orthogonal in vitro assays and are subtype selective for A1AR over A2AAR, A2BAR, and A3AR. Among them, UNC32A (3a) is orally active and has dose-dependent antinociceptive effects in wild-type mice. The antinociceptive effects of 3a were completely abolished in A1AR knockout mice, revealing a strict dependence on A1AR for activity. The apparent lack of cardiovascular side effects when administered orally and high affinity (Ki of 36 nM for the human A1AR) make this compound potentially suitable as a therapeutic