[Effect of schema-focused therapy on depression, anxiety and maladaptive cognitive schemas in the elderly]. / Thérapie des schémas du sujet âgé : impact sur la dépression, l'anxiété et les schémas cognitifs typiques.

INTRODUCTION: Depression is one of the most frequent mental disorders in older people, known to increase rates of disability and mortality. Depression in late life, commonly accompanied by multiple medical illnesses, reduces quality of life and is a strong risk factor for suicide. Despite its clinical significance, depression remains underdiagnosed and inadequately treated in older patients. Cognitive-behavioural psychotherapies have the most empirical support in treating late-life depression, and are recommended by numerous guidelines in this indication. Group interventions are also recommended for older adults because they offer peer support, mitigate social isolation, encourage shared empathy and provide a context for peer feedback help from the group. Previous studies have shown that maladaptive schemas have an important role in the development or maintenance of depression and anxiety in older people, either as risk factors or as vulnerability markers, but there are no studies that have examined the effectiveness of schema-focused therapy to improve depression in late life. OBJECTIVES: The main goals of the present study were to explore the relationship of maladaptive schemas with depression and anxiety severity in aged inpatients, and to evaluate the efficacy of a cognitive-behavioural individual and group treatment program that includes schema-focused therapy on depression, anxiety, and cognitive schemas activation. METHODS: The sample consisted of aged depressed inpatients (n=51) treated in a psychiatric unit. Participants completed measures of depression (Geriatric Depression Scale [GDS]) and anxiety (State-Trait Anxiety Inventory [STAI]) severity and maladaptive schemas (Cognitive Inventory of Subjective Distress [CISD]) at pre- and post-intervention (mean hospital stay: 4weeks). RESULTS: The maladaptive schemas Loss of Individuality, Refusal of Assistance and Vulnerability are more activated in our sample of depressed subjects with regard to the reference population. Most of specific maladaptive schemas (except Fear of Losing Control) were significantly correlated to depression and anxiety-state severity. The GDS and the STAI scores, and the activation of five of seven maladaptive schemas measured by the CISD decreased significantly after treatment. DISCUSSION: This study confirms the results of previous research and shows that specific maladaptive schemas are related to depression and anxiety severity in clinically depressed aged patients. Contrary to other previous studies, we find that the activation of maladaptive schemas can decrease during a group psychotherapeutic program that includes schema-focused therapy. These findings support the feasibility of individual and group schema-focused therapy to assist older people suffering from depression effectively.

Motion of actin filaments in the presence of myosin heads and ATP.

We measured, by fluorescence correlation spectroscopy, the motion of actin filaments in solution during hydrolysis of ATP by acto-heavy meromyosin (acto-HMM). The method relies on the fact that the intensity of fluorescence fluctuates as fluorescently labeled actin filaments enter and leave a small sample volume. The rapidity of these number fluctuations is characterized by the autocorrelation function, which decays to 0 in time that is related to the average velocity of translation of filaments. The time of decay of the autocorrelation function of bare actin filaments in solution was 10.59 +/- 0.85 s. Strongly bound (rigor) heads slowed down the diffusion. Direct observation of filaments under an optical microscope showed that addition of HMM did not change the average length or flexibility of actin filaments, suggesting that the decrease in diffusion was not due to a HMM-induced change in the shape of filaments. Rather, slowing down of translational motion was caused by an increase in the volume of the diffusing complex. Surprisingly, the addition of ATP to acto-HMM accelerated the motion of actin filaments. The acceleration was the greatest at the low molar ratios of HMM:actin. Direct observation of filaments under an optical microscope showed that in the presence of ATP the average length of filaments did not change and that the filaments became stiffer, suggesting that acceleration of diffusion was not due to an ATP-induced increase in flexibility of filaments. These results show that some of the energy of splitting of ATP is impaired to actin filaments and suggest that 0.06 +/- 0.02 of HMM interferes with the diffusion of actin filaments during hydrolysis of ATP.

Diffusion of heavy meromyosin in the presence of F-actin and ATP.

We looked for evidence that the diffusion of heavy meromyosin is modified by its interaction with actin. To be able to observe diffusion in one dimension, we electrophoresed the complex of F-actin and heavy meromyosin in agarose gels in thin capillaries. The intensity profile of the electrophoretic band of the complex showed a sharp peak, which in 1% agarose in the electric field of 17.8 V cm-1 at room temperature migrated at 3.2 cm h-1. The time evolution of the profile after the electrophoresis ended was a measure of the diffusion of heavy meromyosin. After 10 min the intensity profile of heavy meromyosin diffusing in the presence of F-actin and ATP had undergone as much change as the profile of free heavy meromyosin. Modelling of the diffusion process showed that the mean diffusion coefficient of heavy meromyosin moving over actin in the presence of ATP was 7.2 x 10(-7) cm2 s-1 and that it was not statistically different from the diffusion coefficient of free heavy meromyosin. This data is interpreted to show that the diffusion of heavy meromyosin is not modified by its interaction with actin.

Velocity of movement of actin filaments in in vitro motility assay. Measured by fluorescence correlation spectroscopy.

We have measured the velocity of actin filaments in in vitro motility assay by fluorescence correlation spectroscopy. In this method, one measures fluctuations in the number of filaments in an open sample volume. The number of filaments was calculated from measurements of fluorescence of rhodamine-phalloidin bound to F-actin. Sample volume was defined by a diaphragm placed in front of the photomultiplier. Fluctuations arise when actin filaments enter and leave the sample volume due to translations driven by mechanochemical interactions with myosin heads which are immobilized on a glass surface. The average velocity of the translation of filaments determined by the correlation method, (Vc), was equal to the diameter of the diaphragm divided by the half-time of the relaxation of fluctuations. The average number of moving filaments determined by correlation method, (Nc), was inversely proportional to the relative fluctuations. By the fluctuation method it was possible to determine the average velocity of over 800 moving filaments in less than 4 min. There was good agreement between (Vc) and (Nc) and the average velocity and the average number of moving filaments determined manually. To be able to apply correlation measurements to an experimental problem, neither (Vc) nor (Nc) must depend on the position of observation of filaments. We first confirmed that this was indeed the case. We then applied the method to investigate the dependence of motility on the ATPase activity of myosin heads. ATPase activity was varied by mixing intact heads with heads which were labeled with different thiol reagents. It was found that the motion was drastically influenced by the reagent used for modification. When the reagent was N-ethyl-maleimide, 1.5% modification was sufficient to completely inhibit the motion. When the reagent was 5-iodoacetamidofluorescein, motion declined hyperbolically with the fraction of modified heads.

Orientation of actin filaments during motion in in vitro motility assay.

Rhodamine-phalloidin was added to F-actin, and the orientation of transition dipoles of the dye was measured in single actin filaments by polarization of fluorescence. Rhodamine-phalloidin was well immobilized on the surface of actin, indicating that changes in orientation of the dye reported changes in orientation of actin monomers. In stationary filaments the dipoles were inclined at 49.3 degrees with respect to the filament axis. The disorganization of dipoles in stationary filaments was insignificant. When the filaments were made to translate, the average orientation of the dye did not change, but disorganization slightly increased. Disorganization increased significantly when filaments were free in solution. We concluded that, within the accuracy of our measurements (approximately 18%), actin monomers did not undergo major reorientations during motion, but that binding of myosin heads deformed the structure of filaments.

Distribution of actin filament lengths and their orientation measured by gel electrophoresis in capillaries.

F-actin was electrophoresed in capillary tubes filled with agarose gel. The use of capillary imparted high resistance on the gel allowing the use of high enough concentration of salts to keep F-actin polymerized, and allowed the application of high electric fields without liberating considerable amount of heat. The intensity profile of the electrophoretic band of F-actin showed a peak, which in 1% agarose in the electric field of 17.8 V cm-1 at 0 degree C, migrated at 3.4 cm hr-1. Microscopic observation of actin filaments extracted from different positions along the gel showed that during electrophoresis filaments distributed themselves in such a manner that the longest polymers migrated slowest and the shortest migrated fastest. Using this observation we calculated the weight and number distributions of filament lengths from corresponding experimental intensity profiles. Phalloidin-labelled F-actin oriented in the gel upon application of an electric field. F-actin showed unusual orientational response: it oriented rapidly when the field was applied, but relaxed very slowly when the field was removed. Orientation of F-actin varied within an electrophoretic band, longest polymers showing the best orientation and short oligomers and monomers not orienting at all. The degree of orientation increased with the size of the electric field. When F-actin was labelled with phalloidin before electrophoresis, it was no longer able to migrate in the gel, but the electric field oriented it in the same way as when it was labelled after the electrophoresis. These results show that the electrophoresis of F-actin in agarose fractionates it according to its length, that by using electrophoresis it is possible to rapidly obtain distribution of filament lengths, and that F-actin migrates in agarose by the process of reptation.

Measuring orientation of actin filaments within a cell: orientation of actin in intestinal microvilli.

Orientational distribution of actin filaments within a cell is an important determinant of cellular shape and motility. To map this distribution we developed a method of measuring local orientation of actin filaments. In this method actin filaments within cells are labeled with fluorescent phalloidin and are viewed at high magnification in a fluorescent microscope. Emitted fluorescence is split by a birefringent crystal giving rise to two images created by light rays polarized orthogonally with respect to each other. The two images are recorded by a high-sensitivity video camera, and polarization of fluorescence at any point is calculated from the relative intensity of both images at this point. From the value of polarization, the orientation of the absorption dipole of the dye, and thus orientation of F-actin, can be calculated. To illustrate the utility of the method, we measured orientation of actin cores in microvilli of chicken intestinal epithelial cells. F-actin in microvillar cores was labeled with rhodamine-phalloidin; measurements showed that the orientation was the same when microvillus formed a part of a brush border and when it was separated from it suggesting that "shaving" of brush borders did not distort microvillar structure. In the absence of nucleotide, polarization of fluorescence of actin cores in isolated microvilli was best fitted by assuming that a majority of fluorophores were arranged with a perfect helical symmetry along the axis of microvillus and that the absorption dipoles of fluorophores were inclined at 52 degrees with respect to the axis. When ATP was added, the shape of isolated microvilli did not change but polarization of fluorescence decreased, indicating statistically significant increase in disorder and a change of average angle to 54 degrees. We argue that these changes were due to mechanochemical interactions between actin and myosin-I.

Distribution of actin filament lengths measured by fluorescence microscopy.

We analyzed the distribution of actin filament lengths by optical microscopy (OM). OM avoids possible alterations in the size or structure of actin filaments occurring during sample preparation for electron microscopy (EM). Images of F-actin labeled with tetramethylrhodamine isothiocyanate (TRITC)-phalloidin were analyzed for both size distribution and flexibility. In the standard buffer [25 mM potassium acetate, 4 mM MgSO4, 25 mM tris(hydroxymethyl)aminomethane acetate, pH 7.5, 20 mM beta-mercaptoethanol] filaments did not aggregate into bundles and remained stable at nanomolar concentrations for at least 1 h. At the same concentration, actin labeled directly with rhodamine (no phalloidin) formed unstable filaments whose average length decreased with time. The number average length of TRITC-phalloidin labeled filaments (Ln) was 4.90 microns, the ratio (rho) of the weight average length to the number average length was 2.06, and the correlation length (1/lambda) was 8.33 microns. These parameters were in good agreement with the values determined by EM for filaments shorter than 8 microns. Passing G-actin through a Sephadex G-150 column before polymerization did not have a significant effect on the distribution of lengths but made filaments more stiff (1/lambda = 12.5 microns). Millimolar concentration of ATP increased the correlation length, and gelsolin had the expected fragmenting effect on filaments. These results show that OM can be used as a fast and reliable method to analyze the distribution and flexibility of actin filaments and suggest that, in spite of extensive manipulation of actin filaments during sample preparation, EM is a valid tool for determination of size parameters of actin filaments.

ATPase activity of myosin in hair bundles of the bullfrog's sacculus.

Mechanoelectrical transduction by a hair cell displays adaptation, which is thought to occur as myosin-based molecular motors within the mechanically sensitive hair bundle adjust the tension transmitted to transduction channels. To assess the enzymatic capabilities of the myosin isozymes in hair bundles, we examined the actin-dependent ATPase activity of bundles isolated from the bullfrog's sacculus. Separation of 32P-labeled inorganic phosphate from unreacted [gamma-32P]ATP by thin-layer chromatography enabled us to measure the liberation of as little as 0.1 fmol phosphate. To distinguish the Mg(2+)-ATPase activity of myosin isozymes from that of other hair-bundle enzymes, we inhibited the interaction of hair-bundle myosin with actin and determined the reduction in ATPase activity. N-ethylmaleimide (NEM) decreased neither physiologically measured adaptation nor the nucleotide-hydrolytic activity of a 120-kDa protein thought to be myosin 1 beta. The NEM-insensitive, actin-activated ATPase activity of myosin increased from 1.0 fmol x s-1 in 1 mM EGTA to 2.3 fmol x s-1 in 10 microM Ca2+. This activity was largely inhibited by calmidazolium, but was unaffected by the addition of exogenous calmodulin. These results, which indicate that hair bundles contain enzymatically active, Ca(2+)-sensitive myosin molecules, are consistent with the role of Ca2+ in adaptation and with the hypothesis that myosin forms the hair cell's adaptation motor.