Gian Luigi Corona

Opposing Actions of D1 and D2‐Dopamine Receptors on Arachidonic Acid Release and Cyclic AMP Production in Striatal Neurons

Abstract: D1‐and D2‐dopamine receptors exert important physiological actions on striatal neurons, but the intracellular second messenger pathways activated by these receptors are still incompletely understood. Using primary cultures of rat striatal cells, we have examined the effects of activating D1 or D2 receptors on arachidonic acid (AA) release and cyclic AMP accumulation. In striatal neurons labeled by incubation with [3H]AA, D2‐receptor stimulation enhanced release of [3H]AA produced by application of the Ca2+ ionophore A23187 or of the purinergic agonist ATP. By contrast, D1‐receptor stimulation inhibited [3H]AA release. This inhibitory effect of D1 receptors was accompanied by stimulation of adenylyl cyclase activity, measured as accumulation of cyclic AMP, and was mimicked by application of the adenylyl cyclase activator forskolin. The results indicate the existence of a novel signaling pathway for D2 and D1 receptors in striatum, potentiation and inhibition, respectively, of Ca2+‐evoked AA release.

A sensitive fluorimetric method for determination of platelet-bound and plasma free serotonin

A sensitive and specific fluorimetric assay for blood platelet-bound and plasma free serotonin is described. Serotonin is concentrated and purified on Amberlite CG-50 resin, eluted by 0.15 mol/l HCl and its native fluorescence is read at 295/350 nm. The good recovery (90% about), reproducibility and sensitivity (the smallest amount detectable is 5 ng) make the assay particularly useful for the detection of free serotonin in plasma. The whole procedure is rapid enough to permit the analysis of 15--20 samples within one working day and thus the method is suitable for routine determination of serotonin in biological specimens.

Blood noradrenaline and 5-HT levels in depressed women during amitriptyline or lithium treatment.

Noradrenaline levels and platelet and free serotonin concentrations were studied in depressed women in-patients (n=78) before and during amitriptyline (n=41) or lithium treatment (n=37). Pronounced monthly differences in platelet serotonin level have been shown in these subjects before treatment. In all clinical subgroups (neurotic, involutional, manic-depressive patients) a significant fall in platelet serotonin level was observed with amitriptyline medication while an increase was noted with lithium. No significant correlations between serotonin concentrations and clinical outcome were found. Amitriptyline treatment also produced a decrease in peripheral noradrenaline concentration in all subgroups, while an increase was observed with lithium. Some correlations between noradrenaline level and degree of depression were noted in patients treated with amitriptyline or lithium. A more extended analysis of blood amine levels could supply meaningful information on the peripheral action of antidepressive drugs on noradrenaline and serotonin concentrations in depression.

Catecholaminergic, Neuroendocrine and Anxiety Responses to Acute Psychological Stress in Healthy Subjects: Influence of Alprazolam Administration

We studied the effect of alprazolam (APZ) in 12 healthy volunteers on the psychological stress-induced activation of emotion and on the pituitary-adrenal, adrenomedullary and sympathoneuronal systems. After 3 days of placebo or APZ (1 mg/day orally) administration, we examined plasma levels of adrenocorticotropic hormone, cortisol, L-3,4 dihydroxyphenylalanine, dopamine, norepinephrine (NE), epinephrine, metanephrine, normetanephrine, homovanillic acid, vanillylmandelic acid, 3-methoxy-4-hydroxy phenyglycol, urinary levels of cortisol and catecholamines, circulatory responses and state anxiety levels in subjects undergoing psychological stress based on viewing horror, violence, danger and war film clips. Film viewing produced modest rises of state anxiety levels, of plasma NE concentration and of diastolic blood pressure in both the placebo and drug groups. APZ significantly reduced anxiety levels at the beginning of the experimental session and caused a decrease of noradrenergic and dopaminergic neurotransmitter and cortisol concentrations. Our data suggest that APZ reduced anxiety related to the expectation of the event, while the circuitry between structures responsible for anxiety and peripheral sympathoneural function was still found to be partly sensitive to film viewing.

Aspects of amitriptyline and nortriptyline plasma levels monitoring in depression

Fifty-nine depressed female inpatients were treated with 100 mg amitriptiline (AMT) IM for 4 weeks. Depression ratings and determinations of the parent drug and nortriptyline (NT) were performed weekly. No direct relationship between plasma AMT + NT concentrations and therapeutic response was apparent, but beneficial therapeutic responses and significantly lower side-effect scores were more frequently noted in subjects with concentrations in the 100–200 ng/ml range. AMT + NT concentrations were significantly correlated with age. No significant difference was found in the number of responders between younger and older subjects with two clinical improvement criteria; however, a significant difference emerged when a third more restrictive clinical outcome criterion was adopted. The implications of the present findings for patient treatment and for the interpretation of previous studies are discussed. The data collected point to a possible usefulness of monitoring AMT and NT plasma levels, even if further investigations are needed.

Identification and evaluation of amitryptiline and its basic metabolites in rabbit's urine

Abstract The metabolism of amitryptiline has been studied in rabbits. 10-hydroxy amitryptiline, 10,11-hydroxy amitryptiline, nor-amitryptiline, 10-hydroxy noramitryptiline and 10,11-hydroxy nor-amitryptiline were detected in rabbits urine and studied by means of TLC. In addition, a quantitative determination of amitryptiline and some of its metabolites is reported.

Modulation of dopamine-induced cAMP production in rat striatal cultures by the calcium ionophore A23187 and by phorbol-12-myristate-13-acetate

The modulation of cAMP formation by protein kinase C (PKC), activated by phorbol-12-myristate-13-acetate, and by Ca2+ entry, using the ionophore A23187, was investigated in rat striatal neurons grown in primary dissociated cell culture. Phorbol-12-myristate-13-acetate (PMA) potentiated forskolin-induced and dopamine-induced cAMP formation in a concentration-dependent manner. In contrast, the calcium ionophore A23187 inhibited dopamine-induced cAMP formation. When PMA and A23187 were tested simultaneously, the levels of cAMP were not statistically different from those found in the presence of dopamine alone. Furthermore, the decreasing effect of A23187 on cAMP formation was enhanced when PKC was desensitized by pretreating the neurons with 1 microM PMA for 18 h. These data indicate that in striatal neurons Ca2+ entry and PKC activation exert opposing effect on cAMP production.

Assay of urinary vanilmandelic, homovanillic, and 5-hydroxyindole acetic acids by liquid chromatography with electrochemical detection

A reversed-phase liquid chromatographic method with amperometric detection has been developed for the simultaneous determination of the noradrenaline metabolite, vanilmandelic acid, the dopamine metabolite, homovanillic acid, and the serotonin metabolite, 5-hydroxyindole acetic acid, in human urine. After purification by an extraction procedure, the metabolites were rapidly separated under isocratic conditions. Detection and quantification were performed with an electrochemical detector using a carbon paste electrode. The present method is sensitive, selective and, achieves a high degree of precision by the use of isovanilmandelic acid as an internal standard. This provides a suitable tool for clinical and research applications.

Urinary 3-methoxy-4-hydroxyphenylglycol determination using reversed-phase chromatography with amperometric detection

A reversed-phase liquid chromatographic method with amperometric detection has been developed for the determination of urinary 3-methoxy-4-hydroxyphenylglycol. Before and after enzymatic deconjugation, it was purified by an extraction procedure and rapidly quantified under isocratic conditions. The 24-h excretion profile in normal human subjects (eight males and seven females) was determined; our results are consistent with those arrived at in a number of other studies. The present method is highly sensitive and selective; in addition, a good degree of precision is assured by use of 4-methoxy-3-hydroxyphenylglycol as internal standard.

Cell membrane modifications in rabbit isolated hepatocytes following a chronic amitryptiline treatment

Abstract The influence of a chronic amitryptiline pretreatment on the transport of the same drug through the cell membrane has been studied in rabbit isolated hepatocytes, comparing the amount of amitryptiline metabolized by intact hepatocytes with that by disrupted ones. The results show that the amitryptiline metabolism in intact hepatocytes from chronically pretreated rabbits is significantly lower than that in the disrupted ones at all incubation times. By contrast this difference is not present between intact and disrupted hepatocytes from untreated rabbits. These findings support evidence for the hypothesis that the amitryptiline pretreatment affects the permeability of the hepatocyte membrane, thus preventing the penetration of the drug itself into the cells and consequently decreasing its metabolism.