Lionella Palego

Region-dependent effects of flibanserin and buspirone on adenylyl cyclase activity in the human brain

The mode of action of antidepressant drugs may be related to mechanisms of receptor adaptation, involving overall the serotonin 1A (5-HT1A) receptor subtype. However, so far, the clinical effectiveness of selective compounds acting at this level has proved disappointing. This could be explained by the heterogeneity of 5-HT1A receptors within the central nervous system. In animals, two 5-HT1A agonists, flibanserin and buspirone, have shown different pharmacological properties, depending on the brain region. Since no evidence supports this observation in humans, this study sought to investigate whether these two drugs exert different effects on 5-HT1A receptor activation in three different human brain areas: the prefrontal cortex, hippocampus and raphe nuclei. 5-HT1A-mediated inhibition of forskolin-stimulated adenylyl cyclase (AC) was taken as an index of 5-HT1A receptor activation. Flibanserin significantly reduced the activity of AC post-synaptically, i.e. in the prefrontal cortex [EC50 (mean +/- S.E.M.), 28 +/- 10.2 nM; Emax, 18 +/- 2.3%] and in the hippocampus (EC50, 3.5 +/- 3.1 nM; Emax, 20 +/- 4.0%), but had no effect in the raphe nuclei, i.e. at pre-synaptic level. Vice versa, buspirone was only slightly but significantly effective in the raphe (EC50, 3.0 +/- 2.8 nM; Emax, 12 +/- 1.9%). Agonist effects were sensitive to the 5-HT1A antagonists WAY-100135 and pindobind 5-HT1A in the cortex and raphe nuclei, whereas buspirone antagonized flibanserin in the hippocampus. These findings suggest a region-related action of flibanserin and buspirone on forskolin-stimulated AC activity in human brain.

Immune Cell Imbalance in Major Depressive and Panic Disorders

We investigated subsets of peripheral immunologic cells in 12 drug-free patients affected by major depression according to DSM-III-R criteria, and who had recent evidence of somatic diseases. They were compared with 10 drug-free depressives, with 10 patients with panic disorder, and with 12 healthy volunteers, all without somatic disease. The immune subsets were measured by flow cytometry. The results showed that both groups of depressives had the same abnormalities in immune cells compared with the healthy volunteers or the panic disorder patients; in particular they presented a lower number of CD3+, CD8+ and HLA-DR+. The patients with panic attacks did not differ from healthy controls, except for CD4+ cells which were significantly lowered, even in comparison with the depressive groups. These data, although preliminary and in a small sample, suggest that some immune parameters may be influenced by the presence of a major psychiatric disorder.

The trace element content of top-soil and wild edible mushroom samples collected in Tuscany, Italy

The amount of the trace elements As, Ba, Cd, Cr, Cu, Hg, Li, Mn, Ni, Pb, Rb, Se, Sr, and Zn was measured in top soils and edible mushrooms, Boletus edulis, Macrolepiota procera, collected at five distinct green microhabitats inside the Lucca province, North-Central Italy (years 2008–2009). Results showed a top soil element content within the Italian statutory limits. Concerning the amount of mushroom elements, we observed significant species-differences obtaining higher levels of Ni, Rb, and Se in B. edulis or As, Pb, Cu in M. procera. Bioaccumulation factors (BCFs: element in mushroom/element in soil) resulted species-dependent and element-selective: in particular, B. edulis preferentially accumulated Se (BCFs varying from 14 to 153), while M. procera mainly concentrated Cu (BCFs varying from 5 to 15). As well, both species displayed between-site BCF differences. By a multivariate principal component approach, cluster analysis (CA), we could resolve two main clusters of soil element composition, corresponding to the most ecologically divergent sites. Besides, CA showed no cluster relating to element contents of B. edulis at the different collection sites, while a separation in groups was found for M. procera composition with respect to harvesting locations, suggesting uptake systems, in this saprotrophic species, sensitive to microhabitat. Regarding consumer safety, Cd, Hg, Pb levels resulted sometime relevant in present samples, never reaching values from current literature on mushrooms collected in urban-polluted areas. Our findings encourage a deeper assessment of the molecular mechanisms of metal intake by edible mushrooms, encompassing genetic biochemical and geo-ecological variables, with particular awareness to element bioavailability in soils and fungi.

Clozapine, norclozapine plasma levels, their sum and ratio in 50 psychotic patients: influence of patient-related variables.

Steady-state plasma concentrations of clozapine and norclozapine, its major metabolite, as well as their sum and ratio (norclozapine/clozapine), were evaluated in 50 in- and outpatients taking clozapine and naturalistically recruited. Drug plasma concentrations were measured by means of a reversed-phase high-performance liquid chromatography (RPLC) method with an ultraviolet detection. Daily doses (milligrams per kilogram of body weight) of clozapine correlated positively with clozapine plasma parameters, except with the norclozapine/clozapine ratio, in all patients. When the patients were divided in subgroups with respect to gender, the corresponding plasma concentrations were no longer dose-related in men. A lack of significant correlation was observed also in patients (n=23) co-treated with typical neuroleptics. Conversely, dose-concentration correlations were significant in either smoker or nonsmoker patients. No significant relationship between body weight and clozapine plasma parameters was reported. Further, we observed (1) a trend towards higher medians of clozapine or total analytes in women than those reported in men (P=.09 and .07); (2) no significant difference in plasma levels obtained in subjects younger than 34 years and subjects 34 years old or older; (3) a trend towards higher norclozapine and clozapine plus norclozapine levels (P=.05 and .08) in nonsmoker than smoker patients; (4) no significant difference between clozapine plasma parameters measured in patients co-medicated with typical neuroleptics and in patients receiving clozapine alone.

Effect of aging and sex on the [3H]-paroxetine binding to human platelets

Since there exist conflicting results with regard to the possible effect of aging on platelet [3H]-imipramine binding, taken as a peripheral marker of the serotonin (5-HT) transporter, we reinvestigated this matter by comparing the binding of the more selective ligand [3H]-paroxetine in 20 aged and 23 young subjects. The results showed that neither the maximum binding capacity nor the dissociation constant (Kd) were significantly different in the two groups. When the subjects were compared according to sex, the young females revealed a statistically significant lower Kd than the males, while the contrary was true for the aged females. The Kd was significantly and negatively correlated to age in males. In addition, a significant age x gender interaction was also observed. Therefore, the sex of a subject would seem to provoke significant age-related changes in the Kd of [3H]-paroxetine binding to platelet membranes. This might indicate modifications in the 5-HT transporter that could form the basis of a sex-related difference in vulnerability to disorders, such as depression, where a dysfunction at this level is hypothesized.

Localization and gene expression of serotonin1A (5HT1A) receptors in human brain postmortem

We investigated the binding parameters, i.e. the maximum binding capacity (Bmax) and the dissociation constant (Kd), of [3H]8-hydroxy-2-(di-N-propylamino)tetralin ([3H]8-OH-DPAT) labeling the serotonin receptor of the 1A type (5HT1A), and the distribution of the mRNA encoding it in some human brain areas obtained from autoptic samples. The results showed that the Bmax was significantly higher in the hippocampus than in the prefrontal cortex and the striatum, while the Kd had the inverse, although not significant, pattern. The expression study revealed that 5HT1A mRNA distribution in the hippocampus and prefrontal cortex was consistent with the data of the [3H]8-OH-DPAT binding. A different result was obtained in the striatum where no 5HT1A mRNA expression was detected, despite the measurement of specific [3H]8-OH-DPAT binding. These findings underline the different nature of [3H]8-OH-DPAT binding sites in different brain areas and the need for further studies on 5HT receptor gene expression in human brain.

ATP, calcium and magnesium levels in platelets of patients with primary fibromyalgia

OBJECTIVES To evaluate the intracellular levels of the high energy adenosine triphosphate nucleotide ATP and essential divalent cations, calcium and magnesium, in platelets of patients affected by primary fibromyalgia syndrome (FMs). DESIGN AND METHOD Platelet ATP and cation concentrations were measured in 25 patients affected by FMs and 25 healthy volunteers through a chemiluminescent and a fluorimetric assay, respectively. RESULTS Significant lower ATP levels were observed inside platelets of FM patients (fmol ATP/plt: 0.0169+/-0.0012 vs. healthy controls, fmol ATP/plt: 0.0306+/-0.0023, mean+/-SEM) (*** P<0.0001). A trend towards higher calcium concentrations (P=0.06) together with significant increased magnesium levels were also reported in platelets of patients by comparison with controls (P=0.02). CONCLUSIONS This preliminary study suggests that disturbances in the homeostasis of platelet ATP metabolism-signaling and calcium-magnesium flows might have a relevance in the pathogenesis of FMs.

Role of serotonin in human aggressive behaviour

This study aimed to evaluate a peripheral serotonergic marker, 3H-imipramine (3H-IMI) binding to platelet membranes, in a group of severely aggressive subjects (A), institutionalized since childhood for mental retardation, as compared with suicide attempters (S) and healthy controls (H). The maximum binding capacity of 3H-IMI to platelet membranes was statistically lower in (A) and (S) than in (H). In addition, a statistically significant difference was observed between the Bmax values of aggressive subjects and those of suicide attempters. No changes in the dissociation constant (Kd) of IMI binding were observed. These data provide further supporting evidence for the hypothesis of an abnormality of the 5HT system in aggressive behaviour and suggest that such an abnormality, as reflected by platelet markers, is more severe in suicide attempters.

Tryptophan Biochemistry: Structural, Nutritional, Metabolic, and Medical Aspects in Humans

L-Tryptophan is the unique protein amino acid (AA) bearing an indole ring: its biotransformation in living organisms contributes either to keeping this chemical group in cells and tissues or to breaking it, by generating in both cases a variety of bioactive molecules. Investigations on the biology of Trp highlight the pleiotropic effects of its small derivatives on homeostasis processes. In addition to protein turn-over, in humans the pathways of Trp indole derivatives cover the synthesis of the neurotransmitter/hormone serotonin (5-HT), the pineal gland melatonin (MLT), and the trace amine tryptamine. The breakdown of the Trp indole ring defines instead the “kynurenine shunt” which produces cell-response adapters as L-kynurenine, kynurenic and quinolinic acids, or the coenzyme nicotinamide adenine dinucleotide (NAD+). This review aims therefore at tracing a “map” of the main molecular effectors in human tryptophan (Trp) research, starting from the chemistry of this AA, dealing then with its biosphere distribution and nutritional value for humans, also focusing on some proteins responsible for its tissue-dependent uptake and biotransformation. We will thus underscore the role of Trp biochemistry in the pathogenesis of human complex diseases/syndromes primarily involving the gut, neuroimmunoendocrine/stress responses, and the CNS, supporting the use of -Omics approaches in this field.

Simultaneous plasma level analysis of clomipramine, N-desmethylclomipramine, and fluvoxamine by reversed-phase liquid chromatography.

A simple reversed-phase liquid chromatographic method enabling the simultaneous analysis in plasma of the tricyclic antidepressant clomipramine, its demethylated metabolite, and the selective serotonin reuptake inhibitor fluvoxamine, was developed. The drugs and dibenzepine, the internal standard, were extracted from 1 mL plasma through an automated solid-phase procedure, eluted in a total chromatographic time of approximately 14 min and detected by means of an ultraviolet spectrophotometer preset at 254 nm. An assay sensitivity of 10 microg/L was observed for all analytes. Recoveries for these drugs and their metabolites ranged between 65% and 98% and their coefficient of variation (within-day and day-to-day) between 1.9% and 2.9%. In spiked plasma, within-day and day-to-day imprecision data were less than 5%. The simultaneous determination of clomipramine, N-desmethylclomipramine, and fluvoxamine with adequate sensitivity and accuracy may be useful for the monitoring of drug treatment in depression and obsessive-compulsive disorder, where combinations of such drugs are employed.