Pain is clearly the number one reason for patients to consult medical doctors, and is the first symptom they want to be relieved by therapy. Opioids are the most efficient drugs to treat pain but their chronic use at pharmacological doses results in major side effects. The observation that analgesia, free of side effects, can be mediated by opioid receptors expressed on peripheral sensory fibers has opened new therapeutic perspectives for pain treatment. Furthermore, a number of studies suggest that a large part of analgesic effects induced by systemic administration of opioids can be dependent on stimulation of opioid receptors expressed on peripheral sensory fibers. Thus, the finding that endogenous opioid peptides produced by immune cells within inflamed tissues relieve pain represents a sufficiently promising therapeutic way to better understand the physiology of this endogenous pain regulatory mechanism. Although the three classes of endogenous opioids enkephalins, endorphins and dynorphins, have been described in immune cells, their relative prevalence in each immune cell subsets involved in innate and adaptive immune responses have not been investigated so far. The mechanisms involved in the synthesis, production and release of opioids by T lymphocytes in the context of an antigen-specific immune response have not been either explored. In a preliminary study, we have quantified the expression level of mRNA encoding for all three opioid precursors in dendritic cells (DCs), CD4+ and CD8+ T lymphocytes, B lymphocytes and macrophages in mice. We found that although precursors for endorphins and dynorphins may be expressed, the precursor for enkephalins (proenkephalin, PENK) is the main endogenous opioid produced by immune cell subsets. In resting conditions, CD4+ T lymphocytes, macrophages and DCs but not B lymphocytes and CD8+ T lymphocytes express PENK mRNA. Cell activation up-regulates PENK mRNA level only in DCs and CD4+ T lymphocytes. In activated CD4+ T lymphocytes, PENK mRNA level were the highest, reaching more than 50% of that measured in brain. Thus, mature activated CD4+ T lymphocytes could constitute the main sources of opioids (enkephalins) in peripheral tissues. We propose here to establish the contribution of CD4+ T lymphocytes-derived endogenous opioids to inflammatory pain relief. From all peripheral tissues, opioid receptors are the most largely expressed in the gut, where they seem to exert a strong role on inflammatory responses. The objectives of the present study will be 1/ to define the role of effector functions of CD4+ T lymphocyte on their ability to release opioids, 2/ to define the expression and release of endogenous opioids by CD4+ in gut inflammation (inflammatory bowel disease), 3/ to investigate the contribution of CD4+ T lymphocyte-derived opioids to inflammatory pain relief, 4/ to study the control of CD4+-derived opioid synthesis by inflammatory mediators such as proteases and their receptors, and 5/ to determine the opioid receptors activated by CD4+-derived endogenous opioids. The general objective of the present program is to understand the processes and functions of endogenous opioid release, in the context of adaptive and innate immune response such as inflammatory bowel diseases. Determining the conditions and cell types involved in the activation of endogenous mechanisms for the control of pain, constitute a first necessary step towards the definition of new therapeutic options for the treatment of pain: favoring endogenous control of pain.

Infectious diseases caused by apicomplexan protozoan parasites take an enormous toll on human lives and, in some cases, represent a huge burden on domestic animal welfare. Toxoplasma gondii (Toxo) parasites are the causative agent of toxoplasmosis. Toxoplasmosis is a serious veterinarian issue with dramatic consequences, especially for the sheep-related economy. In humans, toxoplasmosis is an opportunistic disease with potential deadly consequences for immunocompromised individuals and for congenitally infected newborns. Toxo also constitutes a major emerging threat, as outbreaks caused by newly isolated, highly virulent strains (especially in South-America) have killed humans with a normal immune system. Of note, research on emerging infectious diseases has been put forward as a priority by the Minister of Research’s “National Strategy for Research and Innovation”. Beyond its impact on human and animal health, Toxo represents an attractive model organism to understand the interactions of mammalian hosts with apicomplexan parasites, among which the malaria-causing Plasmodium spp. parasite is a significant example. A large array of genetic tools is now at our disposal to create mutant Toxo in order to address the specific function of individual parasite components in relevant rodent models. Recent scientific breakthroughs (to which teams 1 and 2 in this proposal have highly contributed) have revealed novel and key features of the innate and adaptive immune responses to Toxo and have shed light on the peculiar protein trafficking strategies utilized by the parasites to create a supportive environment for their growth. As an obligate intracellular organism, Toxo forms and replicates in a specialized parasitophorous vacuole (PV). Within this compartment, Toxo secretes a number of effector proteins, some of which participate in the generation of a tubulovesicular network (TVN) of lipid membranes with a still mysterious function. Additionally, the PV is known to interact and fuse with the host endoplasmic reticulum (ER). A collaborative work between team 1 and the Savina/Amigorena team at the Curie Institute has suggested that ER-PV fusion influences presentation of Toxo-derived T cell antigens. In this context, the two main objectives of this proposal are : 1/ to identify the host cell factors that drive ER-PV fusion and characterize the consequences on antigen presentation of natural Toxo antigens by MHC I molecules. Beyond antigen presentation, we will also determine whether the ER-PV fusion process impacts global host-parasite interactions. 2/ to investigate how the peculiar trafficking of Toxo antigens affect their access to the MHC I pathway and test the hypothesis that the intravacuolar TVN restricts presentation of MHC I antigens by sequestering them in the vacuole. This project is made possible by the recent discovery of a panel of natural Toxo T cell antigens, coupled to novel insights into the cell biology of antigen presentation and parasite protein trafficking. The synergistic combination of tools and expertise possessed by the two main partners of this project (N. Blanchard in Toulouse and M.-F. Cesbron-Delauw in Grenoble), added to the previously established groundwork of collaborations with the Savina team in Paris and the Moita team in Lisbon, will be instrumental for the success of this project. In summary, the proposed research may provide novel molecular and cellular clues for the amelioration of Toxo immune responses which may, in turn, lead to new prospects for therapeutic intervention against toxoplasmosis and other parasitic diseases.

Memory impairment in predemential Alzheimer's disease has usually been related to medial temporal lobe dysfunction. However, atrophy of this region also induces remote cortical reorganization, resulting in both deleterious and compensatory effects with respect to memory performance. The interplay between deleterious and compensatory mechanisms remains improperly understood, partly because these effects have been shown using different neuroimaging methods (basal state (SPECT, PET), resting state (fMRI) or activation studies (PET, fMRI)). The general objective of this research project is to gain a better understanding of the mechanisms underlying cortical reorganization in relation to memory impairment in predemential AD. We identified a series of issues (section 1.2) to propose a research project based on five different studies (section 1.3). We expect our findings will provide new insights in the memory impairment of predemential AD, its functional reorganisation and ultimately, in how to monitor and modulate these reorganization mechanisms (section 1.4). The five studies described in the following project follow a broad logical order, although each is independent and is used to tackle specific scientific questions. Basically, Study 1 aims at showing positive and negative reorganisation mechanisms using PET at basal state, Study 2 aims at assessing the effect of the reorganisation of these mechanisms on memory and Study 3 will complement these findings using cerebral structural connectivity. Study 4 will be focused on bridging methodological issues between basal state PET and resting state fMRI, building mainly on study 1. We will be using CMRGlu in FDG-PET at rest for this study. Study 5 is somehow the result of all previous studies as we will incorporate in this one the methodological issues that will have been addressed in previous studies, notably Study 1 and 4. We will investigate in this study the issue of the baseline condition during plasticity imaging and during activation tasks. This project is lead by a young neurologist, Dr. Jérémie Pariente ('MCU-PH' Toulouse University Hospital and Inserm U 825). The present project involves three other young scientists, Dr. Emmanuel Barbeau (CR2, psychologist), CNRS UMR 5549, Dr. Florence Remy (MCU, physicist), CNRS UMR 5549 and Xavier Franceries (MCU, physicist), Inserm UMR 825 all in Toulouse, France. They come from two different labs that support their project, want to build a new expert group on AD and have significant and complementary expertise on the different fields required to complete this project. The synergy between the different participants to this project is good and balanced between clinical and methodological expertise. It relies on true (and demonstrated through publications) expertise on the different fields required to complete the project satisfactorily: neurology, neuropsychology, memory assessment, neuroimaging techniques, plasticity, methodological developments, research project management.