Sustained remission with lamotrigine augmentation or monotherapy in female resistant depressives with mixed cyclothymic-dysthymic temperament.

BACKGROUND: The treatment of bipolar depression remains problematic. Lamotrigine has been shown in randomized controlled studies to be efficacious in preventing bipolar depression and rapid cycling states. METHODS: Twenty-four women with cyclothymic temperament and refractory depression were recruited from four outpatient sites (three primary care and one psychiatric) and treated with lamotrigine in a naturalistic, open-label study. Temperament was determined by responses on the TEMP-A self-rating scale. Eighteen (75%) of these cyclothymic patients also scored high on the depressive temperament. Eighteen (75%) met DSM-IV criteria for bipolar II disorder. In two thirds of the cases, lamotrigine was add-on therapy to an antidepressant. Response to therapy was assessed using the DSM-IV Global Assessment of Functioning (GAF). LIMITATIONS: This study was naturalistic in design, without controls or blinds. RESULTS: Of the 23 patients who remained in the study, 16 (70%) had significant, sustained responses. Of these 16, 12 (75% of responders, 52% of the total) had remissions (GAF > 80) sustained longer than 12 months. Robust, sustained responses to lamotrigine monotherapy were seen in 4 patients (17%). Seven patients (30%) received no apparent benefit from lamotrigine. CONCLUSIONS: Lamotrigine induced prolonged illness remissions in a substantial number of female patients whose symptoms were both complex and refractory. Most manifested high scores on the cyclothymic and depressive temperaments, and prior refractoriness to multiple antidepressant and antidepressant/mood stabilizer combinations, before remitting with lamotrigine augmentation or monotherapy.

TEMPS-A: progress towards validation of a self-rated clinical version of the Temperament Evaluation of the Memphis, Pisa, Paris, and San Diego Autoquestionnaire.

BACKGROUND: Our aim was to validate the Temperament Evaluation of the Memphis, Pisa, Paris, and San Diego Autoquestionnaire (TEMPS-A) in a clinical population. METHODS: The study was conducted in two Memphis mood clinics involving 398 affectively ill patients with young to middle index age (42 years+/-13 S.D.), who were 95% white, 62% female, and 51% bipolar spectrum. A subset of 157 of the entire sample were retested in 6-12 months, and the entire sample was then subjected to factor analysis (PCA extraction method with varimax rotation). RESULTS: We obtained high test-retest reliability ranging from 0.58 for the irritable, to 0.68, 0.69 and 0.70, respectively, for the cyclothymic, dysthymic and hyperthymic. The hypothesized four-factor structure of the TEMPS-A was upheld, with the cyclothymic explaining 14% of the variance, followed by the irritable, hyperthymic, and dysthymic together accounting for another 14%. Internal consistency was excellent, with Chronbach alphas ranging from 0.76 for the dysthymic to 0.88 for the cyclothymic. Exploratory factor analysis revealed 2 super factors, Factor I loading on cyclothymic, irritable, and dysthymic temperaments, and Factor II loading heavily on the hyperthymic. The 50-item TEMPS-A-Clinical Version was constructed by using a cutoff of alpha > or =0.4 for traits loading exclusively on their original temperaments. We also proposed a longer 69-item version for future study, in which we permitted a greater number of traits based on clinical considerations (alpha cutoff 0.30). LIMITATION: The sample was preponderantly white, and may not generalize to other U.S. ethnic groups. This earlier version of TEMPS-A did not include the anxious temperament. CONCLUSIONS: We psychometrically validated the TEMPS-A in affectively ill outpatients, leading to an instrument suitable for use in psychiatric, especially affectively ill, populations. It is noteworthy that in this clinically ill population we succeeded in measuring traits which could make subjects vulnerable to affective episodes, as well as those of adaptive nature. For instance, the dysthymic emerged as bound to routine, self-blaming, shy-nonassertive, sensitive to criticism, yet self-denying, dependable, and preferring to work for someone else rather than be the boss. The hyperthymic had the highest number of "positive" traits: upbeat, fun-loving, outgoing, jocular, optimistic, confident, full of ideas, eloquent, on the go, short-sleeper, tireless, who likes to be the boss, but single-minded, risk-taker, and unlikely to admit to his/her meddlesome nature. The cyclothymic emerged as labile with rapid shifts in mood; unstable in energy, self-esteem and socialization; unevenly gifted and dilettante; yet keen in perception, intense in emotions, and romantic. The irritable emerged as skeptical and critical (which might be considered intellectual virtues), but otherwise having the "darkest" nature of all temperaments: grouchy, complaining, dissatisfied; anger- and violence-prone, and sexually jealous. The foregoing temperament attributes, observed in a moderately severe group of patients with affective disorders, nonetheless testify to the evolutionary context of these disorders-"submissive" behavior, territoriality, romantic charm, and last, but not least, sexually jealous with its associated specter of violence. We hypothesize that the putative social and limbic mechanisms underlying mood disorders appear to have archaic origins on an evolutionary scale. We finally submit that the traits underlying affective disorders are very much part of human nature.

Characterization and partial purification of the 8-9S androgen receptor from benign prostatic hyperplasia tissues.

The present study reports a 3,800-fold purification of the 8-9S androgen-receptor complex from benign prostate hyperplasia (BPH) tissues using differential chromatography. In addition, the BPH androgen receptor complexes have been characterized using sucrose density gradient (SDG) ultracentrifugation, gel permeation, and anion exchange high performance liquid chromatography (HPLC). Results indicate that a) under nontransforming conditions, BPH cytosols contained both 8-9S (40-78%) and 4S (22-60%) androgen-receptor forms, b) apparent molecular weights of these androgen-receptor apparent molecular weights of these androgen-receptor complexes, as analyzed by gel permeation HPLC, were estimated to correspond at 270 kDa, and 90 kDa respectively, c) 8-9S androgen-receptor complexes were retained on an anion exchange HPLC column and could be eluted at 0.22 M KCl at a linear gradient, whereas 4S complexes were not retained on anion exchange columns under identical experimental conditions, d) 10X dilution of BPH cytosols containing only the 4S (0.6 M KCl) form and subsequent chromatography on anion exchange HPLC system was indicative of fragmentation (these fragments were retained on anion exchange columns and could be eluted by 0.33 M KCl on a linear gradient HPLC), and e) increased temperature (22 C) was permissive of proteolytic fragmentation (fragments were estimated to correspond at 30, 15, and 5 kDa). The results are discussed in relationship with the composition of the nontransformed androgen-receptor molecules.

Toxinological studies on the nematocyst venom of Chrysaora achlyos.

Nematocyst venoms from both oral arms and lappets of Chrysaora achlyos were prepared and found to have factors producing mouse lethality, hemolysis and hepatocyte toxicity. These venoms had less potency than those of Chrysaora quinquecirrha a phylogenetic, congeneric cousin. Envenomated bathers had significant species-specific anti venom IgG and also cross-reacting antibody to Chrysaora quinquecirrha nematocyst venoms. There were similarities and contrasts in the capillary electropherograms and sodium dodecyl sulfate (SDS) gels between C. achlyos nematocyst venoms and those of their C. quinquecirrha counterparts.

Electrophysiological and hemolytic activity elicited by the venom of the jellyfish Cassiopea xamachana.

In this study, we determined hemolysis activity in human and sheep erythrocytes, and characterized the electrical responses in Xenopus oocyte membrane elicited by the venom of the jellyfish Cassiopea xamachana (Cx). The Cx venom produced hemolysis in both species, being more potent on human red cells. The electrophysiological study showed that the Cx venom elicited three different responses in the oocytes. One current was generated in all the oocytes tested and corresponded with a slow inward current (I(Cx)) associated with an increase in membrane conductance. I(Cx) was concentration-dependent and had a reversal potential of -10.3+/-0.4 mV. Ionic substitution studies indicated that the conductive pathway was mainly permeable to cations and non-selective. The oocyte membrane resistance was completely recovered after washout of the venom, this suggested that the effect was due to generation of a specific membrane conductance as opposed to a possible non-specific membrane breakdown. A comparative study with three distinct native cationic channels present in the oocyte membrane [i.e. (1) hemi-gap-junction channels, (2) mechanosensitive channels, and (3) the ouabain-sensitive channel activated by palytoxin], showed that I(Cx) might correspond to opening of mechanosensitive channels or to activation of an unknown cationic channel located in the oocyte membrane. The bioactive fraction eliciting I(Cx) were peptides and was separated from two other peptidic hemolytic fractions by chromatography.

A comparison of the toxinological characteristics of two Cassiopea and Aurelia species.

A comparison of the toxinological properties of nematocyst venoms from Old and New World Cassiopea and Aurelia species was undertaken. The cnidom of venomous Cassiopea andromeda (Ca) and Aurelia (Aa(RS)) from the Red Sea was identical to that of nonvenomous Bahamian Cassiopea xamancha (Cx) and Chesapeake Bay Aurelia aurita (Aa(CB)), respectively. A clean nematocyst preparation of Ca and both Aurelias could be obtained but algal particles could not be separated completely from the Cx nematocysts. Further purification of all four nematocyst preparations showed significant differences in the action of their protein. Only the Cassiopea had coexisting dermonecrotic and vasopermeability producing properties and Ca's hemolytic activity was associated with mouse lethality. The protein, hemolysin and phospholipase gel filtration eluant curves of Ca venom were similar. Venomous Aa(RS) actively stung lips and contained more potent mouse lethal, demonecrotic, vasopermeability plus hemolytic factors than Aa(CB). Cross reactivity of convalescent human serum obtained from patients stung by Ca and venomous Cx collected in Central America occurred. This was also observed between sera of bathers stung by Aa(RS) and stinging Aurelia which appeared in Florida during the recent El Niño year. IgG was stimulated by several nematocyst proteins since many venom subfractions tested positive at high titers against convalescent sera. T-cell proliferation of mice primed with either Aurelia venom was positive against the homologous preparation with cross reactivity to the heterologous venom. Crude venoms of both Red Sea jellyfish metabolically stimulated cultured human hepatocytes more than their New World counterparts. This data shows that considerable similarities and differences exist in the venoms of these Old and New World Cassiopea and Aurelia medusae with the Eastern species being more potent.

Characterization of androgen receptor by high performance liquid chromatography and sucrose density gradient ultracentrifugation in normal and malignant human prostatic tissues.

In the present study we have characterized androgen-receptor complexes of normal and malignant human prostate cytosols using sucrose density gradient centrifugation, gel permeation and anion exchange high performance liquid chromatography (HPLC). Our results indicated that: 1) malignant tissue cytosols differed from normal by the presence of a 4-5S androgen receptor form which accounted for 30% of total specific-binding of malignant tissue cytosols, 2) 8-9S androgen-receptor complexes in normal and malignant prostate cytosols were estimated as 270kDa by gel permeation HPLC, 3) 8-9S complexes were retained and could be eluted by 0.22M KCl on a linear gradient anion exchange HPLC, 4) 4-5S androgen-receptor complexes were estimated as 90kDa by gel permeation HPLC and were not retained on anion exchange HPLC in our experimental conditions, and 5) either 10X dilution of the 4-5S complexes and subsequent anion exchange HPLC, or anion exchange chromatography of 8-9S complexes at 22 degrees C were causing fragmentation of the androgen receptor molecule from normal and malignant tissues. These fragments had enhanced affinity for anion exchange columns. These results are discussed in relation to the composition of the nontransformed androgen receptor macromolecule.

Synthesis of two photoreactive heterobifunctional reagents derived from hexanoic acid.

The synthesis of two photoreactive heterobifunctional reagents derived from hexanoic acid is described. The compounds are succinimido 6-N-(4-azidobenzoyl)aminohexanoate (1c) and succinimido 6-mercapto-S-(4-azidothiophenyl)hexanoate (2c). Compound 1c was synthesized from benzyloxycarbonyl 4-aminobenzoic acid and 6-aminohexanoic acid benzyl ester. Compound 2c was obtained by disulfide exchange of dithiobis-4-aminobenzene with 6-mercaptohexanoic acid. The azido function was introduced by displacement of the corresponding diazonium salt and the active ester, by the mixed anhydride method. Both compounds were decomposed by ultraviolet irradiation. Phytohemagglutinin was modified by reaction with reagent 1c or 2c. Irradiation afforded polymeric lectin derivatives resulting from intermolecular cross-linking. Sodium dodecyl sulfate--polyacrylamide gel electrophoresis analyses revealed the presence of Coomasie-positive bands of molecular weight 33000, 67000, 120000, 144000, and higher. Polymeric structures resulting from irradiation of phytohemagglutinin modified with compound 2c were cleaved by reduction with 2-mercaptoethanol or with dithiothreitol. Phytohemagglutinin modified by reaction with compound 1c or 2c retained its ability to stimulate pig spleen lymphocytes in vitro. Whereas the lectin treated with reagent 2c was active as the unmodified protein, the lectin treated with compound 1c was more active than unmodified lectin at nearly all the concentrations tested.

Toxinological studies of the venom from Cassiopea xamachana nematocysts isolated by flow cytometry.

The tentacle epithelial tissue of Cassiopea xamachana contains nematocysts and symbiotic algal particles. These two structures were dissociated, analyzed and sorted by flow cytometry. A simple separating method was developed utilizing the algal chlorophyll autofluorescence and the nematocysts' fluorescence after the uptake of fluorescent stains. A five-fold increase in mouse lethality; significantly more potent hemolytic and cytosensing activities; as well as a cleanup in the capillary electropherogram and SDS gel profiles for the crude nematocyst venom preparations prepared by fluorescence activated cell sorting (FACS), was observed relative to alternative methods. Because the hemolytic potency of pre-sorting nematocyst venom was minimal and the post-sorting counterpart was significantly positive, the possibility that algae inhibited the venom's toxinological activity was considered.

Toxinological and immunological studies of capillary electrophoresis fractionated Chrysaora quinquecirrha (Desor) fishing tentacle and Chironex fleckeri Southcott nematocyst venoms.

Repeated runs of capillary electrophoresis (CE) were used to study partially-purified jellyfish nematocyst venom protein in concentrations sufficient to perform toxinological assays. Nematocyst venoms from Chironex fleckeri (Cf) and Chysaora quinquecirrha were processed. The CE eluate was divided into quadrants by scanning protein content. The fourth fraction of both jellyfish venoms, contained proteins with the smallest molecular weight components, which were responsible for the highest hemolysins and the humoral and cell-mediated immunological activity. Cytotoxic Cf lethal factor activity against human liver cells was widely dispersed throughout both venoms but more prominent in fraction 4. A V(beta) receptor human T-cell repertoire was not species-specific for either crude or fractionated jellyfish nematocyst venom.

Application of high-performance liquid chromatofocusing to the study of prostatic androgen receptors. Effect of stabilizing agents on the heterogeneity of receptor structure.

Cytosolic androgen receptors from normal rat prostate were analyzed by high-performance liquid chromatofocusing. Two ion-exchange columns, AX-300 and AX-500, and two equilibration systems, Tris-HCl and imidazole-HCl, were used. pH gradients ranged between 8.3 and 3.5 for Tris-HCl and from 7.7 to 3.5 for imidazole-HCl. In the absence of sodium molybdate and inhibitors of proteolytic enzyme, six specific radioactive fractions (pH: 7.9, 7.7, 7.0, 5.1, 4.7 and 4.4) were eluted from AX-300 equilibrated with Tris-HCl in a ratio of 28:20:52 for acidic, intermediary and basic forms, respectively; similar results were obtained with AX-500 although this column was less effective in resolving basic forms of the receptor. The buffering capacity of imidazole-HCl was lower than that of Tris-HCl, resulting in a steeper elution pH profile. The resolution between acidic and basic forms was thus diminished and only four specific radioactive fractions at pH 7.2, 7.1, 6.5 and 3.6, were observed on AX-500 in a ratio of 23:10:67 for acidic, intermediary and basic forms. In the presence of sodium molybdate, two acidic fractions were found with Tris-HCl at pH 4.3 and 4.7 (47%) on AX-300, whereas the radioactivity of fractions at pH 7.0 and 5.1 (32%) was considerably lowered and intermediary forms remained unchanged (21%). With imidazole-HCl on AX-500, the peak at pH 7.2 disappeared and the acidic form shifted from pH 3.6 to 4.3. In the presence of inhibitors of proteolytic enzyme and sodium molybdate, specifically bound radioactivity was found mostly in a broad acidic fraction (75%) at pH 4.5 on columns equilibrated with Tris-HCl; radioactivity at pH 7.6 disappeared completely but a small amount (15%) remained at pH 7.9. In imidazole-HCl, a main radioactive fraction was eluted at pH 7.1 and two other fractions were collected at pH 6.8 and 4.3 respectively. In conclusion, multiple forms of the rat prostate androgen receptor were evinced by high-performance liquid chromatofocusing. Tris-HCl proved to be a more efficient equilibration system than imidazole-HCl for the resolution of rat prostate cytosolic binding proteins. Under the experimental conditions used, sodium molybdate and inhibitors of proteolytic enzyme greatly favored the acidic form to the detriment of the intermediary and basic entities.

Studies on the characterization of rat prostate androgen receptors.

In the presence of sodium molybdate and protease inhibitors, two forms of androgen-receptor complexes were observed which sedimented in the areas of 8-9S and 5-7S by SDG centrifugation. The intermediary 5-7S form was better seen when complexes were incubated at low KCl concentrations. The sedimentation coefficient of this form fluctuated between 5 and 7S depending on the KCl concentration. At high ionic strength (0.6M KCl) in all media, one form only was observed having a sedimentation coefficient value of 4.3S. By gel exclusion chromatography, we also observed two specific entities at 75A and 68A; in the presence of 0.6M KCl, however, two entities were found at 68A and 43A. The constant presence of protease inhibitors in all buffers was necessary to separate the intermediary 68A form. We calculated molecular weights of about 270 kDa, 190 kDa, and 80 kDa respectively for these three forms. [3H]R1881-receptor complexes bound to DEAE-cellulose and were eluted in the absence of glycerol at 0.1M and 0.2M KCl. Material found at 0.1M KCl sedimented in the areas of 5-7S and 8-9S in nearly equal proportion, and that found at 0.2M KCl sedimented in the 8-9S area only. When the cytosol was chromatographed at a fast flow rate (4 ml/min), untransformed 8-9S receptors did not bind to phosphocellulose, but transformed complexes were retained, could be eluted with 0.4M KCl and sedimented in the 4S area on KCl free SDG centrifugation. When the excluded untransformed 8-9S complexes were re-chromatographed at a slow flow rate (1 ml/min), they were retained on phosphocellulose, and could be eluted with 0.3M KCl.(ABSTRACT TRUNCATED AT 250 WORDS)

Characteristics of the rat prostate androgen receptors analyzed by sucrose density gradient and high-performance liquid chromatofocusing.

Rat prostate cytosolic androgen-receptor complexes were analyzed by sucrose density gradient (SDG) centrifugation and by high-performance liquid chromatofocusing (HPCF). Without protecting agents, these complexes were resolved by HPCF at basic (8.25-7.1), intermediary (7.0-5.0) and acidic (4.6-4.2) pH. Sodium molybdate stabilized labeled complexes which migrated in the 8-9S and 3.5-6S areas on SDG. These were further stabilized by the presence of sodium molybdate and four protease inhibitors: complexes then sedimented mainly in the 8-9S area with a shoulder at 6-7S. Forms eluting at acidic pH on HPCF were favored by the presence of sodium molybdate and further enhanced by the addition of inhibitors, to the detriment of basic ones. Furthermore, when chromatographed on phosphocellulose (P-c), unretained complexes sedimented as a symmetrical peak on SDG centrifugation in the 8-9S area, but were eluted from HPCF columns as two entities at pH 4.1 and 4.6. The P-c retained complexes subsequently detached by 0.6 M KCl, were resolved into three entities by HPCF with a major component at pH 8.2, which sedimented in the 4S areas. These results demonstrate that the gradual decrease in the negative net charge of androgen receptor correlates with the gradual reduction in mass of the androgen-receptor complex. Moreover, this can be interpreted as further evidence for a heterogeneity of androgen receptor population in rat prostate, suggesting the involvement of a multistep mechanism preceding the induction of specific gene transcription by the hormone.

Purification and characterization of the untransformed androgen receptor in rat prostate.

The isolation and characterization of the untransformed form of androgen receptors has not yet been successful, owing to their inherent lability as well as to their ready proteolysis. In this study, we have stabilized rat prostate androgen receptors by sodium molybdate and by rapid filtration on phosphocellulose. Proteases were inhibited by bacitracin, aprotinin, leupeptin and PMSF. Under these conditions the untransformed complex was purified approx 3000-fold, corresponding to 18% yield, by differential chromatography on DEAE cellulose and phosphocellulose gels. The partially purified receptor has the same ionic characteristics as the original untransformed receptor of crude cytosol; in addition, it possesses a Stokes' radius of 75 A, as determined by Sephacryl S-300 gel filtration, a sedimentation coefficient of 8.8S, a calculated molecular weight of 275 kDa and a friction coefficient of 1.6. The [3H]R1881 receptor complex was specific to androgens since unlabelled R1881 and dihydrotestosterone were able to completely displace bound [3H]R1881, whereas estradiol, cortisol, and triamcinolone acetonide did not compete. The purified complex was a multimer dissociable by 0.6 M KCl, resulting in a form migrating in the 4S area on sucrose density gradient. After treatment with 0.5% formaldehyde, three forms were obtained, migrating in the areas of 8-9, 5-6 and 3-4S respectively, of a sucrose density gradient containing 0.6 M KCl. This is the first step towards the purification to homogeneity of the untransformed androgen receptor.

Characterization of androgen receptors in normal and malignant human prostatic tissues.

The aim of this work was to study the physico-chemical characteristics of androgen receptors of normal and malignant human prostatic tissues. Association (k+1) and dissociation (k-1) rate constants and sedimentation profiles on sucrose density gradients were determined on [3H]androgen receptor complexes. In the presence of 20 mM sodium molybdate, no significant difference in k+1 and k-1 values could be found between cytosolic receptor preparations from normal and malignant specimens. The values obtained for k+1 (mean +/- SD) were 5 +/- 2 X 10(6)M-1 min-1 (N = 3) and 5 +/- 2 X 10(6)M-1 (N = 5); k-1 values of 23 +/- 4 X 10(-4) min-1 (N = 4) and 25 +/- 3 X 10(-4) min-1 (N = 5) were obtained for normal and malignant tissues respectively. Similar Nmax values were also obtained for normal [(mean +/- SD) 26 +/- 10 fmol/mg cytosolic protein (N = 5)] and malignant [20 +/- 9 fmol/mg protein (N = 7)] tissues. A statistically significant difference was found however, between k-1 values measured on [3H]androgen receptor complexes of nuclear extracts; values of 15 +/- 3 X 10(-4) min-1 (N = 4) and 9 +/- 1 X 10(-4) min-1 (N = 6) were found for normal and malignant tissue preparations respectively. This was also accompanied by a higher level of androgen receptors in nuclear extracts of malignant [Nmax, 308 +/- 171 fmol/g of tissue (N = 8)] compared to normal [Nmax, 68 +/- 11 fmol/g of tissue (N = 5)] tissues. The cytosolic [3H]androgen receptor complexes prepared from normal tissues sedimented mainly in the 8-9S area on sucrose density gradient whereas those from malignant tissues sedimented 50% in the 8-9S area and 50% in the 4S area. In conclusion, in this study, we found a different sedimentation profile of androgen-receptor complexes from the cytosol of prostatic cancers compared to normal as well as a diminution of their dissociation rate constant in nuclei.

Application of octadecylsilyl-silica in purification studies related to the nontransformed rat ventral prostate androgen receptor.

We investigated the application of octadecylsilyl (ODS)-silica in studies related to characterization and purification of the nontransformed rat ventral prostate androgen receptor. The results indicated that ODS-silica successfully separates the free [3H]R1881 from the labeled [3H]R1881 transformed (4-5S) and nontransformed (8-9S) rat ventral prostate androgen receptors. Partial purification of the 8-9S receptor form was performed by the fractionation of rat cytosol using cartridges of the ODS-silica and fast-flow-rate phosphocellulose chromatography. Further purification was accomplished by differential chromatography (DEAE-cellulose and slow-flow-rate phosphocellulose chromatography). This partially purified 8-9S receptor, when analyzed on a gel permeation high-performance liquid chromatography column, resulted in a three-peak pattern of UV absorbance. One of these peaks corresponded to a 59-kD non[3H]R1881-binding protein and the remaining two corresponded to 270-kD and 190-kD[3H]R1881-binding proteins. These results demonstrate the usefulness of ODS-silica in androgen receptor studies. The association of a 59-kD nonsteroid binding protein with the nontransformed rat ventral prostate androgen receptor is discussed.