Determination of 5-chloro-7-iodo-8-quinolinol (clioquinol) in plasma and tissues of hamsters by high-performance liquid chromatography and electrochemical detection.

This paper describes a method of determining clioquinol levels in hamster plasma and tissue by means of HPLC and electrochemical detection. Clioquinol was separated on a Nucleosil C18 300 mm x 3.9 mm i.d. 7 microm column at 1 ml/min using a phosphate/citrate buffer 0.1M (400 ml) with 600 ml of a methanol:acetonitrile (1:1, v/v) mobile phase. The retention times of clioquinol and the IS were, respectively, 11.6 and 8.1 min; the quantitation limit (CV>8%) was 5 ng/ml in plasma and 10 ng/ml in tissues. The intra- and inter-assay accuracies of the method were more than 95%, with coefficients of variation between 3.0 and 7.7%, and plasma and tissue recovery rates of 72-77%. There was a linear response to clioquinol 5-2000 ng/ml in plasma, and 10-1000 ng/g in tissues. The method is highly sensitive and selective, makes it possible to study the pharmacokinetics of plasma clioquinol after oral administration and the distribution of clioquinol in tissues, and could be used to monitor plasma clioquinol levels in humans.

Behavioural characterization of interleukin-6 overexpressing or deficient mice during agonistic encounters.

Interleukin-6 (IL-6) is a cytokine released by activated immune cells which has been shown to affect brain function. In this experiment aggressive and affiliative behaviour exhibited during agonistic encounters by transgenic male mice either not expressing (IL-6 -/-) or overexpressing (NSE-hIL-6) IL-6 in the central nervous system was investigated. All subjects were isolated for 24 days before the aggressive encounter and were 52 days old at the time of testing. Subjects were placed for 5 consecutive days in a neutral cage for 15 min with an opponent of the Balb/c strain that had been previously isolated for the same amount of time. The first and the last test sessions were videotaped to evaluate the first approach and the establishment of the social role, respectively. A number of behavioural categories were later scored. When compared with wild-type controls, IL-6 -/- mice showed a higher degree of aggressive behaviour as indicated by a higher frequency of Offensive Upright Posture, an effect more pronounced on the fifth encounter. On the contrary, NSE-hIL-6 subjects showed a tendency to be more involved in affiliative-type social interactions, displaying a higher frequency and duration of behaviours such as Anogenital, Nose or Body Sniff. IL-6 -/- mice showed a clear tendency to exhibit less affiliative interactions compared with their controls while dopamine levels were found to be modified in a number of brain regions in these mice. Overall, these data suggest that IL-6 affects both aggressive and affiliative-type interactions, although the behaviour of the NSE-hIL-6 subjects was less affected than that of the IL-6 -/- group. The effects of the genetic background of the animal in screening the outcome of gene manipulations on behaviour are also discussed.

GM-CSF affects hypothalamic neurotransmitter levels in mice: involvement of interleukin-1.

We studied the effects of granulocyte-macrophage colony stimulating factor (GM-CSF) on the brain levels of several neurotransmitters in mice. Administration of GM-CSF (5.0 and 10 microg, i.p.) significantly reduced the hypothalamic levels of glutamine, glutamic acid, GABA and aspartic acid. GM-CSF (5.0 microg, i.p.) also induced a significant reduction of norepinephrine and serotonin levels in the hypothalamus, without affecting dopamine levels. The hippocampal levels of neurotransmitters were not modified by GM-CSF administration. The peripheral administration of a specific interleukin-1 receptor antagonist (IL-1ra, 50 microg, i.p.) blocked the effects of GM-CSF. These results confirm our previous behavioural data suggesting that GM-CSF is able to exert neuromodulatory actions.

In vitro effects on monoamine uptake and release by the reversible monoamine oxidase-B inhibitors lazabemide and N-(2-aminoethyl)-p-chlorobenzamide: a comparison with L-deprenyl.

To investigate whether the reversible monoamine oxidase-B (MAO-B) inhibitors lazabemide and Ro 16-6491 have any additional effect on monoamine uptake and release, in vitro experiments were performed on rat forebrain synaptosomes and blood platelets. The effects of the two drugs were compared with those of L-deprenyl, the well-known irreversible MAO-B inhibitor which is reported to affect amine uptake. Both lazabemide and Ro 16-6491 behaved as weak inhibitors of [3H]monoamine uptake by synaptosomes, with a similar rank order of potency for amine uptake inhibition (noradrenaline (NA) > or = 5-hydroxytryptamine (5 HT) > dopamine (DA)). The IC50 values for lazabemide and Ro 16-6491, respectively, were: 86 microM and 90 microM for NA uptake; 123 microM and 90 microM for 5HT uptake; > 500 microM and > 1000 microM for DA uptake. L-Deprenyl (rank order of inhibitory potency: NA > DA > 5 HT) was four to 10 times more potent than either compound in inhibiting [3H]catecholamine uptake (IC50 = NA 23 microM, DA 109 microM), and two to three times less potent in inhibiting 5 HT uptake (IC50 233 microM). Lazabemide and Ro 16-6491 also differed from L-deprenyl in their ability to induce release of endogenous monoamines from synaptosomes. Thus, Ro 16-6491 (500 microM) induced a greater 5 HT release than did L-deprenyl, but was less effective than L-deprenyl in releasing DA. On the contrary, lazabemide was almost completely inactive on either 5 HT and DA release. The differential effect of the three MAO-B inhibitors on synaptosome 5 HT uptake and release was confirmed by [14C]5HT uptake and liberation experiments with isolated rat platelets. The data indicate that the reversible MAO-B inhibitors lazabemide and Ro 16-6491 at relatively high concentrations possess amine uptake-inhibiting properties. With regard to the effects examined, lazabemide markedly differs from L-deprenyl since it does not interfere with DA uptake nor induce amine release from synaptosomes.

Effect of aging on lazabemide binding, monoamine oxidase activity and monoamine metabolites in human frontal cortex.

Age-related modifications of monoamine oxidase-A and -B (MAO-A and MAO-B) and amine metabolite concentrations were studied in human frontal cortex taken postmortem from 22 subjects of various ages (21-75 years). Qualitative and quantitative analysis for MAO-B was provided by kinetic studies with a specific radioligand, [3H]lazabemide. The data demonstrated a significant (P < 0.05) positive correlation between the density of [3H]lazabemide binding sites (Bmax) and age of the subject, without showing an apparent modification in the dissociation constant (KD) of the radioligand. In parallel experiments, MAO-B but not MAO-A activity was shown to correlate with age (P < 0.05). The concentrations of the amine metabolites 4-hydroxy-3-methoxyphenylacetic acid (HVA), 5-hydroxyindole-3-acetic acid (5-HIAA), 3,4-dihydroxyphenylacetic acid (DOPAC), 4-hydroxy-3-methoxyphenylglycol (MHPG) and 3,4-dihydroxyphenylglycol (DHPG) were all devoid of a correlation with age. Neither did the concentrations of these metabolites relate to the respective subject's MAO-B enzymatic activity nor to [3H]lazabemide Bmax. A correlation, though rather weak, was obtained between MAO-A activity and MHPG concentration (P = 0.045). The MAO-A and -B enzyme characteristics in subjects who had committed suicide (n = 9) did not differ from those of subjects deceased for other causes (n = 13). Among the measured monoamine metabolites the concentrations of DOPAC and HVA were higher in the suicide versus control group (P < 0.05). The present data confirm in a direct manner that the increase in MAO-B activity in aging brain is due to an enhancement of the number of active sites of the enzyme and not through modifications of its kinetic characteristics. Furthermore, that neither the characteristics nor the activity of the enzyme are changed in the frontal cortex of suicide victims compared to control subjects.

Estrogen modulation of catecholamine synthesis and monoamine oxidase A activity in the human neuroblastoma cell line SK-ER3.

In order to assess the neuronal-like properties of a human neuroblastoma cell line obtained by stable transfection of the estrogen receptor (SK-ER3) a series of quantitative measurements of the activity of two neurotransmitter-related enzymes: tyrosine hydroxylase (TH) and monamine oxidase (MAO), and of catecholamine concentrations were performed. When compared to the parental SK-N-BE cell line, the stably transfected SK-ER3 cells show a more pronounced dopaminergic phenotype. The immunoreactivity to a TH antibody is in fact increased and the ratio between dopamine and noradrenaline concentrations is elevated. Treatment with estradiol further enhances the expression of this phenotype. Interestingly, in the transfected cell line MAO-A activity is decreased and further reduced by estrogen treatment. This finding substantiated by previous reports indicates that our model system might represent an interesting tool for the study of the pharmacological treatments of estrogen-induced pathological responses of nervous cells.

Effect of acute and short-term administration of cholinomimetic drugs on corticosterone secretion in the rat.

Centrally acting cholinomimetic drugs have been proposed for the therapy of cognitive disorders in aged subjects. Among the possible adverse side effects of this class of compounds, of great relevance is the stimulatory action on the adrenocortical axis, in view of the toxicity of glucocorticoids for hippocampal neurons and the immune system. The aim of the present study was to evaluate in conscious male rats the effect of acute and short-term administration of three novel cholinomimetic drugs on the release of corticosterone. The potent agonist of muscarinic receptors RU 35963 strikingly increased corticosterone levels after acute but not after short-term (6 days) administration. Similar results were obtained after administration of the reversible inhibitor of cholinesterase, eptastigmine. In contrast to RU 35963 and eptastigmine, acute administration of a choline precursor, L-alpha-glycerylphosphorylcholine, only slightly affected plasma corticosterone concentrations after both acute and short-term administration. It is concluded that activation of adrenocortical function by cholinomimetic drugs is a short-lasting event which does not represent an important side effect of these compounds when given on a long-term basis.

Effects of phosphatidylserine therapy in geriatric patients with depressive disorders.

The effects of phosphatidylserine (BC-PS) on cognitive, affective and behavioural symptoms were studied in a group of 10 elderly women with depressive disorders. Patients were treated with placebo for 15 days, followed by BC-PS (300 mg/day) for 30 days. The Hamilton Rating Scale for Depression, Gottfries-Bråne-Steen Rating Scale, Nurse's Observation Scale for Inpatient Evaluation and Buschke Selective Reminding Test were administered before and after placebo and after BC-PS therapy, to monitor changes in depression, memory and general behaviour. At the same time, basal plasma levels of noradrenaline, MHPG, DOPAC, HVA and 5-HIAA, and GH/beta-endorphin/beta-lipotropin responses to clonidine stimulation were measured. BC-PS induced consistent improvement of depressive symptoms, memory and behaviour. No changes in amine metabolite levels or in hormonal responses to alpha 2-adrenoceptor stimulation were observed.

Adrenal response in the calf to repeated simulated transport.

The adrenal responses in calves submitted to simulated transport on three occasions for 30 min were evaluated. Plasma adrenaline, cortisol and NEFA increased significantly during simulated transport but became less marked in successive trials. Haematological stress-related parameters (Hb, PCV) increased to the same extent on repeated exposure to simulated transport. Plasma noradrenaline, glucose and cholesterol values were unchanged throughout the study.

Vasopressin (DDAVP) therapy in chronic schizophrenia: effects on negative symptoms and memory.

Ten chronic undifferentiated schizophrenics, 6 men and 4 women, aged 28-63, with 6- to 31-year histories of the disease were given DDAVP to observe the effects of this neuropeptide on the prevalent negative symptoms of their illness. Patients were maintained on neuroleptic therapy and first given a 20-day course of placebo followed by 20 days of DDAVP i.m., 4 micrograms Andreasen Scale for assessment of negative symptoms, the Brief Psychiatric Rating Scale, the NOSIE Rating Scale and the Luria-Nebraska Rating Scale were administered to monitor negative symptomatology, behavior and memory before the study began, after placebo and after DDAVP administration. Patients were also given a growth hormone-clonidine test and in addition plasma basal concentrations of 3-methoxy-4-hydroxyphenylglycol (MHPG), homovanillic acid, 3,4-dihydroxyphenylacetic acid (DOPAC) and 5-hydroxyindoleacetic acid (5-HIAA) were measured at the same intervals. DDAVP therapy induced a significant improvement of negative symptomatology and a trend toward improvement of short- to medium-term memory. No changes in homovanillic acid, MHPG, 5-HIAA and DOPAC, nor of growth hormone response to clonidine stimulation were observed.

Mechanism of the antihypertensive effect of FCE 22716, a new ergoline derivative, in the spontaneously hypertensive rat.

Both acute and chronic oral administration (1-20 mg/kg) of FCE 22716, a new ergoline derivative, resulted in a dose-related fall of arterial blood pressure lasting for more than 6 h. Tachycardia was observed only at high dosages. Yohimbine, propranolol and indometacin did not modify its antihypertensive effect; on the other hand pretreatment with prazosin, a selective alpha 1-adrenoceptor antagonist and pithing, almost completely neutralized its antihypertensive effect. Haloperidol, a dopamine antagonist that crosses the blood-brain barrier, also antagonized FCE 22716 activity. The lack of effects of domperidone (DA2-receptor antagonist selectively acting on the periphery) together with the finding that norepinephrine and epinephrine levels were unchanged after treatment with FCE 22716, seem to rule out an involvement of peripheral DA2-receptors. Both in vitro (isolated organs) and in vivo the compound antagonized responses mediated by stimulation of alpha 1-adrenoceptors and S2-receptors. Radioligand binding studies in different cerebral regions are in line with the above reported in vitro and in vivo results. These data suggest that FCE 22716 is endowed with a multitarget mechanism of action, mainly involving blockade of alpha 1-adrenoceptors and S2-receptors.

Epinephrine mediates the growth hormone-releasing effect of galanin in infant rats.

The mechanism underlying the GH-releasing effect of galanin (GAL), a novel 29-amino acid peptide, was investigated in the neonatal rat. The effect of galanin was compared to that of clonidine (CLO), a drug known to release GH via endogenous GHRF. GAL administration (5-25 micrograms/kg BW, sc) induced in 10-day-old pups a clear-cut and dose-related rise in plasma GH 15 min postinjection. CLO (50-450 micrograms/kg BW, sc) induced a marked rise in plasma GH, but no dose-related effect was evident. Inhibition of hypothalamic norepinephrine and epinephrine biosynthesis by DU-18288 (6 mg/kg BW, ip) or selective inhibition of epinephrine biosynthesis by SKF-64139 (50 mg/kg BW, ip) completely abolished the GH-releasing effect of GAL (25 micrograms/kg, sc), but left unaltered the GH rise induced by CLO (150 micrograms/kg, sc). Passive immunization with an anti-GHRF serum decreased basal GH levels and prevented the GH-releasing effect of either GAL or CLO, whereas in pups pretreated with an antisomatostatin serum, CLO, but not GAL, increased the already elevated plasma GH titers. In all these data indicate that in the infant rat 1) GAL is a potent GH secretagogue; 2) the action of GAL is not exerted directly on GHRF- or somatostatin-secreting structures, but requires the intervention of catecholaminergic neurons; 3) the GH-releasing effect of GAL is ultimately exerted via GHRF release, although a mechanism operating to inhibit hypothalamic somatostatin release cannot be ruled out; and 4) differently from GAL, CLO releases GH via postsynaptic stimulation of GHRF-secreting neurons.

Serotonin and noradrenaline concentrations and serotonin uptake in platelets from hyperphenylalaninaemic patients.

In three untreated patients with phenylketonuria (PKU), three PKU and six hyperphenylalaninaemic (HPA) patients in good metabolic control, the kinetic constants of platelet in vitro uptake of [14C]serotonin (5HT) did not significantly differ from those in 12 control subjects matched for age. The platelet concentrations of endogenous 5HT and noradrenaline (NA), taken as long-term indices of the amount of these amines circulating in plasma, were lower than normal in PKU and HPA patients, whether or not they were kept on a diet. However, a reduction in plasma NA concentrations at the moment of blood collection was seen only in untreated PKU patients. These data indicate that there may be a chronic inhibition of 5HT and possibly of NA synthesis even in PKU or HPA subjects in good metabolic control, with normal psychomotor development and only slightly raised plasma phenylalanine levels.

Selective modification of renal alpha 2-adrenergic receptors in Milan hypertensive rat strain.

Cerebral and renal alpha-adrenergic receptors play an important role in the control of blood pressure. We studied alpha-adrenergic receptors in the cerebral and renal cortex of Milan hypertensive strain (MHS) and normotensive strain (MNS) rats, a genetic model of spontaneous hypertension linked to a kidney abnormality. Binding of the selective alpha 1-adrenergic antagonist [3H]prazosin and the alpha 2-adrenergic antagonist [3H]rauwolscine was used for receptor studies in tissues of prehypertensive (24-day-old) and hypertensive (60-day-old) rats. In the cerebral cortex, no between-strain differences in alpha 1-adrenergic and alpha 2-adrenergic receptor density and affinity were observed in prehypertensive and hypertensive periods. The density of these receptors increased similarly with age in MHS and MNS rats. In the renal cortex, the differences between MHS and MNS rats concerned alpha 2-adrenergic receptors only. Compared with their age-matched normotensive controls, MHS rats showed 1) a lower affinity for the antagonist (p less than 0.05) in the prehypertensive period, 2) absence of the normal age-related increase in receptor density, and 3) a lower density of [3H]rauwolscine binding sites (p less than 0.001) in the hypertensive period. In this period, studies of competitive inhibition of [3H]rauwolscine binding showed that l-epinephrine bound to one class of sites in MHS rats (pseudo-Hill plot, 0.90) and to two classes in MNS rats (pseudo-Hill plot, 0.68). In addition, the lack of any guanylylimidodiphosphate effect on the l-epinephrine competition curve observed in MHS rats suggests the uncoupling of these receptors from the guanosine 5'-triphosphate binding protein.(ABSTRACT TRUNCATED AT 250 WORDS)

Serotonin involvement in lisuride-induced mounting and in sleep.

Mounting behavior of rats induced by several drugs, such as e.g. p-CPA, 5,7-dihydroxytryptamine, L-DOPA or lisuride, appears to result from a combined decrease of 5-hydroxytryptamine (5-HT) and increase of dopamine (DA) neurotransmission. In this paper, lisuride-induced mounting is proposed as a behavioral model for detecting pharmacologically active drugs that interact with monoaminergic mechanisms, e.g. 5-HT reuptake blockers and type A monoamine oxidase inhibitors. With regard to sleep, 5-HT appears to sustain a relevant part in controlling the sleep-wakefulness cycle. However, other transmitters or neuromodulators (catecholamines, oligopeptides etc.) may also be involved in sleep mechanisms.

Mechanisms involved in the prolactin-releasing effect of benserazide.

The mechanism(s) underlying the prolactin (PRL)-releasing effect of benserazide (Bz), a peripheral inhibitor of L-aromatic amino-acid decarboxylase, was investigated in the rat. In intact male and female rats, Bz was ineffective to increase significantly plasma PRL at 0.8 mg/kg i.p. but elicited an already maximal effect at 1.6 mg/kg. Bz added to in vitro incubated anterior pituitaries (APs) did not alter PRL secretion at the dose of 3.8 X 10(-6)M but increased PRL release at 10(-4)M. Bz, even at very high doses (up to 10(-3) M), did not displace [3H]spiroperidol binding from AP membrane preparations. In rats having had mechanical ablation of the medio basal hypothalamus (MBH), Bz (15 mg/kg i.p.) induced no rise in plasma PRL and did not counteract the striking inhibitory effect of a dopamine (DA) infusion (5 micrograms/kg per min per 120 min). Administration of Bz (15 mg/kg i.p.) into intact male rats decreased significantly the DA concentrations in the median eminence (ME) but not in the residual hypothalamus and the AP. In the same rats 1-dopa (50 mg/kg i.p.) increased significantly the DA concentrations not only in the ME but also in the hypothalamus and the AP. Bz given concurrently with 1-dopa markedly reduced the rise in DA concentrations induced by 1-dopa in the ME, and greatly potentiated the increase in DA concentrations in the hypothalamus. These data indicate that the mechanism whereby a single administration of Bz increases PRL secretion in the rat is not consistent with the postulated DA receptor antagonist action of the drug, but instead implies inhibition of the decarboxylation of 1-dopa at dopaminergic nerve terminals of the ME.

Diclofenac increases beta-endorphin plasma concentrations.

Plasma and ventricular cerebro-spinal fluid (CSF) Beta-endorphin concentrations were evaluated after chromatographic separation in patients carrying a ventricular shunt before and after the administration of diclofenac or placebo. In the same subjects the ventricular CSF concentrations of the serotonin and the catecholamine metabolites 5-hydroxyindole-acetic acid (5-HIAA), homovanillic acid (HVA) and MOPEG were also evaluated. Plasma, but not ventricular, Beta-endorphin concentrations increased significantly after diclofenac, while placebo was ineffective. No significant changes in ventricular 5-HIAA, HVA or MOPEG levels were observed. These data suggest a role for Beta-endorphin in the analgesic effect of diclofenac.

Effects of light saline loading on renal renin and catecholamine release in hypertensive humans.

In 8 essential hypertensive patients infusion of 7.5 ml kg-1 b.w. of isotonic saline over 20 min decreased plasma renin activity but did not affect plasma adrenaline or noradrenaline concentrations in suprarenal aortic, renal venous and caval blood. Mean blood pressures and heart rates were unchanged throughout the saline infusion period, whereas the hematocrit was significantly decreased. These data suggest that inhibition of renin secretion after slight volume expansion with saline in hypertensive humans occurs without parallel inhibition of either renal neurosympathetic drive or humoral adrenergic tone to the kidney through circulating catecholamines.

Plasma catecholamine concentrations in normotensive rats of different strains and in spontaneously hypertensive rats under basal conditions and during cold exposure.

Under basal conditions, the levels of circulating norepinephrine (NE) and epinephrine (E) were higher in normotensive Wistar rats of different origins than in Sprague-Dawley rats. Since the decline of 3H-NE concentration in the plasma after i.v. injection was similar in Wistar and in Sprague-Dawley rats, the higher levels of endogenous NE in the former strain probably reflect greater NE release from sympathetic nerve terminals. In normotensive Sprague-Dawley and Wistar rats, plasma NE rose to various extents during cold exposure (4 degrees C), depending on the basal plasma NE levels. Compared with normotensive Wistar Kyoto rats (WKY), spontaneously hypertensive rats (SHR) had similar basal plasma E and NE concentrations, similar rates of 3H-NE disappearance, but more rapid increases to higher values of plasma NE during cold exposure. It is concluded that the basal rate of peripheral catecholamine release does not seem to be the main determining factor for arterial blood pressure in the various rat strains and that the sympathetic neuronal system of SHR is more responsive to cold exposure than that of WKY rats.