[Common physical health problems in psychiatry]. / Problèmes courants de santé physique en psychiatrie.

COMMON PHYSICAL HEALTH PROBLEMS IN PSYCHIATRY. Somatic psychiatric care is particularly important as life expectancy declines of 13 to 16 years among individuals with severe mental disorders, primaly due to cardiovascular and cancer-related diseases. They face a triple burden: their mental disorders, reduced life expectancy and limited healthcare access. Common comorbidities related to psychotropic medications, lifestyle and the mental condition itself, affect various organs with complex clinical expressions. The multidimensional vulnerability is attributable to the side-effects of psychotropic drugs, mental disorders, and healthcare system fragmentation, hindering access to care. The proposed solutions aim for a better integration of mental health into general healthcare, with increased awareness among caregivers for a holistic approach to health.

PROBLÈMES COURANTS DE SANTÉ PHYSIQUE EN PSYCHIATRIE. Les soins somatiques en psychiatrie revêtent une importance particulière en raison de la réduction de treize à seize ans de l'espérance de vie chez les personnes atteintes de troubles mentaux sévères, principalement à cause de pathologies cardiovasculaires et cancéreuses. Elles sont exposées à une triple peine : leurs troubles psychiques, la réduction de leur espérance de vie et un accès limité aux soins. Les comorbidités courantes, liées aux psychotropes, au mode de vie et à la pathologie mentale elle-même, touchent divers organes, avec des expressions cliniques complexes. La vulnéra bilité multidimensionnelle de ces patients est attribuable aux effets indésirables des psychotropes, aux troubles psychiques et au cloisonnement du système de santé nuisant à l'accès aux soins. Les solutions proposées visent une meilleure intégration du volet santé mentale dans les soins généraux, avec une sensibilisation accrue des soignants à une approche holistique de la santé.

Agonist-selective dynamic compartmentalization of human Mu opioid receptor as revealed by resolutive FRAP analysis.

Techniques for analyzing the membrane diffusion of molecules are the most promising methods for investigating the compartmentalization of G-protein-coupled receptors, particularly as relevant to receptor signaling processes. Here, we report fluorescence recovery after photobleaching (FRAP) measurements performed at variable spot radius for human mu opioid (hMOP) receptors on SH-SY5Y neuroblastoma cells in the presence of ligands. Although an antagonist did not affect the behavior of the receptors compared with the basal state, two different agonists, DAMGO and morphine, caused markedly different changes to receptor diffusion. Like receptors in the absence of ligand, receptors bound to morphine exhibited diffusion confined to joined semipermeable domains, but with smaller domain size and diffusion coefficient. This effect was inhibited by pertussis toxin, strongly suggesting that this dynamic behavior is associated with early steps of signaling. In the presence of DAMGO, half of the receptors displayed free long-range diffusion and the other half were confined to smaller isolated domains. Hypertonic sucrose buffer suppressed this effect, which we attribute to receptor entry into clathrin-coated pits. It is likely that the observation of distinct receptor dynamics in the presence of DAMGO and morphine involves the agonist-selective phosphorylation of the receptor.

Changes of the membrane lipid organization characterized by means of a new cholesterol-pyrene probe.

We synthesized 3beta-hydroxy-pregn-5-ene-21-(1-methylpyrenyl)-20-methylidene (Py-met-chol), consisting of cholesterol steroid rings connected to a pyrene group via a linker without polar atoms. This compound has interesting spectroscopic properties when probing membranes: 1), The pyrene has hypochromic properties resulting from probe self-association processes in membranes. Using liposomes of various lipid compositions, we determined the association constants of the probe (K): K(DOPC) >> K(POPC) >> K(DMPC) > K(DMPC/15 mol % Chol) > K(DMPC/30 mol % Chol). This indicates a better probe solvation in saturated than in unsaturated lipids, and this effect is enhanced as the cholesterol concentration increases. 2), The pyrene fluorophore is characterized by monomer (I(1)-I(5)) and excimer (I(E)) emission bands. In model membranes, I(1)/I(3) and I(E)/I(3) ratios revealed a correlation between the polarity of the lipid core of the membrane and the amount of cholesterol. 3), Using this probe, we monitored the first steps of the signaling pathway of the mouse delta-opioid receptor, a G-protein-coupled receptor. The thickness of the membrane around this receptor is known to change after agonist binding. Fluorescence spectra of living Chinese hamster ovary cells overexpressing mouse delta-opioid receptor specifically revealed the agonist binding. These results indicate that Py-met-chol may be useful for screening ligands of this family of receptors.

Functional membrane diffusion of G-protein coupled receptors.

G-protein-coupled receptor function involves interactions between the receptor, G-proteins and effectors in the cell plasma membrane. The main biochemical processes have been individually identified but the mechanisms governing the successive protein-protein interactions of this complex multi-molecular machinery have yet to be established. We discuss advances in understanding the functional dynamics of the receptor resulting from diffusion measurements, and in the context of the plasma membrane organization.

Diffusion of the mu opioid receptor at the surface of human neuroblastoma SH-SY5Y cells is restricted to permeable domains.

Previous single-molecule studies have shown a long-term diffusion superimposed to a short-term confinement of the human mu opioid (hMOP) receptors at the surface of heterologous cells. However, additional ensemble average measurements are required to reach a complete understanding of the undergoing process. Here, we analyse, by fluorescence recovery after photobleaching measurements, the lateral diffusion of fully functional T7-EGFP-hMOP receptors in neuroblastoma SH-SY5Y cells naturally expressing a low level of the wild-type receptor. Experiments carried out at variable observation radii demonstrate the restriction of the receptors diffusion to sub-micrometer sized domains. Furthermore, consistently with the long-term single-molecule data, the domains are found permeable.

Dynamic confinement of NK2 receptors in the plasma membrane. Improved FRAP analysis and biological relevance.

A functional fluorescent neurokinin NK2 receptor, EGFP-NK2, was previously used to follow, by fluorescence resonance energy transfer measurements in living cells, the binding of its fluorescently labeled agonist, bodipy-neurokinin A (NKA). Local agonist application suggested that the activation and desensitization of the NK2 receptors were compartmentalized at the level of the plasma membrane. In this study, fluorescence recovery after photobleaching experiments are carried out at variable observation radius (vrFRAP) to probe EGFP-NK2 receptor mobility and confinement. Experiments are carried out at 20 degrees C to maintain the number of receptors constant at the cell surface during recordings. In the absence of agonist, 35% EGFP-NK2 receptors diffuse within domains of 420 +/- 80 nm in radius with the remaining 65% of receptors able to diffuse with a long range lateral diffusion coefficient between the domains. When cells are incubated with a saturating concentration of NKA, 30% EGFP-NK2 receptors become immobilized in small domains characterized by a radius equal to 170 +/- 50 nm. Biochemical experiments show that the confinement of EGFP-NK2 receptor is not due to its association with rafts at any given time. Colocalization of the receptor with beta-arrestin and transferrin supports that the small domains, containing 30% of activated EGFP-NK2, correspond to clathrin-coated pre-pits. The similar amount of confined EGFP-NK2 receptors found before and after activation (30-35%) is discussed in term of putative transient interactions of the receptors with preexisting scaffolds of signaling molecules.

Confined diffusion without fences of a g-protein-coupled receptor as revealed by single particle tracking.

Single particle tracking is a powerful tool for probing the organization and dynamics of the plasma membrane constituents. We used this technique to study the micro -opioid receptor belonging to the large family of the G-protein-coupled receptors involved with other partners in a signal transduction pathway. The specific labeling of the receptor coupled to a T7-tag at its N-terminus, stably expressed in fibroblastic cells, was achieved by colloidal gold coupled to a monoclonal anti T7-tag antibody. The lateral movements of the particles were followed by nanovideomicroscopy at 40 ms time resolution during 2 min with a spatial precision of 15 nm. The receptors were found to have either a slow or directed diffusion mode (10%) or a walking confined diffusion mode (90%) composed of a long-term random diffusion and a short-term confined diffusion, and corresponding to a diffusion confined within a domain that itself diffuses. The results indicate that the confinement is due to an effective harmonic potential generated by long-range attraction between the membrane proteins. A simple model for interacting membrane proteins diffusion is proposed that explains the variations with the domain size of the short-term and long-term diffusion coefficients.

Probing functionalized gold colloids for single particle tracking experiments.

Functionalized submicroscopic particles are currently used to label proteins or lipids at the surface of living cells for single particle tracking experiments. In many cases, it can be of crucial importance for the particle to be anchored to a single molecule. We have addressed this question for the labeling at the plasma membrane of NRK cells of the mu-opioid receptor bearing a T7 epitope at the N-terminus. Using biophysical methods we were able to prepare quasi-monovalent anti-T7 antibody conjugated gold colloids (40 nm diameter) leading to stable and specific binding to the receptor. The rational method, we report here, can be extended to design customized probes for the labeling of various tagged molecules.

Monoclonal antibodies recognizing larval- and pupal-specific cuticular proteins of Tenebrio molitor (Insecta, Coleoptera).

To study the sequential expression of insect epidermal cells during metamorphosis, a library of monoclonal antibodies (MABs) was prepared against the water-soluble proteins from preecdysial pupal cuticle of Tenebrio molitor. Six selected MABs recognizing only larval and pupal cuticular proteins (CPs) in immunoblot analysis were classified into three types. Type 1 recognized a 21.5 and a 22 kDa polypeptide, type 2, a 26 kDa polypeptide, and type 3, three polypeptides of 18.5, 19.5 and 21.5 kDa. They did not immunoreact with any protein of fat bodies or haemolymph from pharate pupae, suggesting that the antigens originate from the epidermis. The stage-specificity was confirmed by electron microscopic immunogold labelling. Type 1 and 3 MABs recognized antigens characterizing larval and pupal preecdysial sclerotized cuticles, while the antigens recognized by type 2 were localized in the first few lamellae of unsclerotized postecdysial cuticle. When the expression of the adult programme was inhibited by application of a juvenile hormone analogue, the larval-/pupal-specific CPs were detected in the supernumerary pupal cuticle. These results suggest that the genes encoding these proteins are juvenile hormone dependent. These MABs should be useful tools to isolate pupal-specific genes whose regulation sems to be different from that of the adult-specific ones.