Region-dependent effects of acute and chronic tranylcypromine in vivo on [3H]2-BFI binding to brain imidazoline I(2) sites.

An imidazoline I(2) site has been localised to monoamine oxidase. However, in vitro studies of the effect of monoamine oxidase inhibitors on imidazoline I(2)-site radioligand binding have produced conflicting findings. Using the technique of autoradiography, we examined the effect of in vivo administration of the irreversible monoamine oxidase inhibitor tranylcypromine on binding of the imidazoline I(2) site-specific ligand [3H]2-(-2-benzofuranyl)-2-imidazoline ([3H]2-BFI) in four rat brain nuclei which are known to possess a high density of imidazoline I(2) sites, together with cerebral cortex and cerebellum which show weaker binding. A single acute pre-treatment with tranylcypromine significantly increased imidazoline I(2) site-specific binding in four regions: arcuate nucleus, interpeduncular nucleus, pineal gland and area postrema, but effects in cortical areas and cerebellum were not significant. The extent of the increase was proportional to the control binding in each region. In contrast, five daily treatments with the same dose of tranylcypromine significantly reduced [3H]2-BFI binding in these same areas. The potential role of monoamine oxidase isoforms in these changes is discussed.

Former chronic methylenedioxymethamphetamine (MDMA or ecstasy) users report mild depressive symptoms.

Previous work has indicated recreational use of methylenedioxymethamphetamine (MDMA or ecstasy) is associated with elevated scores on self-report measures of depression. We sought to examine the long-term effects of consumption on depression in a group of individuals who had consumed large quantities of the drug in the past, but were now leading relatively drug free lives. Respondents to this study (n = 29) had consumed an average of 1.5 ecstasy tablets in the last month, 8.4 in the last 6 months and 23.3 in the last 12 months. The estimated total consumed was 527 tablets, indicating that these respondents were indeed former chronic users of the drug. None of the respondents had consumed ecstasy in the last 14 days. Levels of depression (Beck's Depression Inventory) were significantly (p < 0.01) elevated compared to a matched non-drug using control group. Within the group of former chronic users, these levels of depression were not significantly affected by current use of alcohol, cannabis or amphetamine, but were positively correlated with an external locus of control (p < 0.05), infrequent but severe- (p < 0.05) and frequent but mild- (p < 0.005) self-report measures of life stress. Multiple regression indicated that levels of frequent but mild life stress (p < 0.005) and the quantity of ecstasy tablets respondents consumed over a 12-h period (p < 0.05) were the only variables that were significant predictors of self-reported levels of depression. The results of this study indicate that former chronic ecstasy users report higher levels of depression than their matched controls.

Effects of nicotine on head-shakes and tryptophan metabolites.

RATIONALE: Nicotine appears to ameliorate the tics of Tourette syndrome. There is evidence that plasma concentrations of the tryptophan metabolite kynurenine may be elevated in this condition. Rodent head-shakes have been proposed as a putative model of Tourette syndrome and are potentiated by kynurenine. OBJECTIVES: To determine the effects of acute and chronic nicotine on mouse head-shakes, and to study whether nicotine influences brain and plasma levels of kynurenine and certain of its further metabolites in this species. METHODS: Behavioural and biochemical studies. RESULTS: Acute (-10 min) administration of (-)-nicotine, or the nicotinic agonist (+)-epibatidine, dose dependently attenuated head-shakes induced by the 5-HT2A/2C receptor agonist +/-1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (DOI). This attenuation was inhibited by the nicotinic receptor antagonist mecamylamine. Acute nicotine did not affect either spontaneous head-shakes or plasma and brain kynurenine. Fifteen hours after the last of twice daily injections of nicotine (1.6 mg/kg for 7 days), the frequency of spontaneous and DOI-induced head-shakes was significantly potentiated and there was a significant elevation of both plasma and brain kynurenine, although no differences were detected in plasma concentrations of tryptophan, kynurenic acid, 3-hydroxykynurenine or 3-hydroxyanthranilic acid. Brain levels of 5-hydroxytryptamine and 5-hydroxyindole acetic acid were also unaffected. In contrast, all these measures were unchanged 15 h after a single nicotine dose (1.6 mg/kg). CONCLUSIONS: The acute studies indicate that head-shakes induced by DOI are indeed inhibited by nicotinic receptor agonists and suggest that this is not a consequence of an increase in kynurenine. While a role for kynurenine or its metabolites in increasing the head-shake rate after chronic nicotine cannot be excluded, alternative explanations included alterations in the expression or functional status of nicotinic receptor components and further work will be required to characterise this effect.

The brain decade in debate: II. Panic or anxiety? From animal models to a neurobiological basis.

This article is a transcription of an electronic symposium sponsored by the Brazilian Society of Neuroscience and Behavior (SBNeC). Invited researchers from the European Union, North America and Brazil discussed two issues on anxiety, namely whether panic is a very intense anxiety or something else, and what aspects of clinical anxiety are reproduced by animal models. Concerning the first issue, most participants agreed that generalized anxiety and panic disorder are different on the basis of clinical manifestations, drug response and animal models. Also, underlying brain structures, neurotransmitter modulation and hormonal changes seem to involve important differences. It is also common knowledge that existing animal models generate different types of fear/anxiety. A challenge for future research is to establish a good correlation between animal models and nosological classification.

The brain decade in debate: II. Panic or anxiety? From animal models to a neurobiological basis

This article is a transcription of an electronic symposium sponsored by the Brazilian Society of Neuroscience and Behavior (SBNeC). Invited researchers from the European Union, North America and Brazil discussed two issues on anxiety, namely whether panic is a very intense anxiety or something else, and what aspects of clinical anxiety are reproduced by animal models. Concerning the first issue, most participants agreed that generalized anxiety and panic disorder are different on the basis of clinical manifestations, drug response and animal models. Also, underlying brain structures, neurotransmitter modulation and hormonal changes seem to involve important differences. It is also common knowledge that existing animal models generate different types of fear/anxiety. A challenge for future research is to establish a good correlation between animal models and nosological classification

Chronic administration of the cholesterol reducing drug gemfibrozil fails to alter 5-HT1A and 5-HT2A mediated receptor behaviours in rats.

Some studies have suggested that reductions in plasma cholesterol might be associated with suicidal behaviour. Serotonergic systems are thought to be involved in suicidal ideation and behaviour and links with altered 5-HT1A and 5-HT2A receptors have been proposed. We have therefore examined the effects of cholesterol reduction using gemfibrozil, upon 5-HT2A and 5-HT1A receptor-related behaviours in rats. Rats were treated chronically (57 days) with gemfibrozil (50 mg/kg p.o.) or gum acacia vehicle and challenged sequentially with the 5-HT1A agonist 8-hydroxy-d-n-aminopropyl tetralin, to elicit 5-HT1A syndrome behaviours, and the 5-HT2 receptor agonist 1-(2,5-dimethoxy-4-iodophenyl)2-amino-propane to establish their head-shake frequency. Significant cholesterol reduction, within the clinical range, failed to induce any changes in either 5-HT1A or 5-HT2A mediated behaviours. These data suggest that cholesterol reduction fails to induce changes in 5-HT1A and 5-HT2A receptor tone suggesting that the reduction of plasma cholesterol, within the human clinical range, does not result in neuroplasticity of the 5-HT1A and 5-HT2A receptors in rats.

Studies on uptake, sub-cellular trafficking and efflux of antisense oligodeoxynucleotides in glioma cells using self-assembling cationic lipoplexes as delivery systems.

The cellular uptake of antisense oligodeoxynucleotides (ODNs) may be enhanced by the use of carriers such as cationic liposomes or lipoplexes, but little is known about the intracellular fate and subcellular trafficking of these systems in target cells. In this study, we report on the cellular uptake and biodistribution of ODNs in the presence and absence of optimised self-assembled cationic lipoplexes using the C6 glioma cell line as an in vitro model. Biotin or radiolabelled 15-mer phosphorothioate (PS) ODNs were synthesised and their cellular uptake and subcellular biodistribution characterised in the presence and absence of an optimised cationic lipoplex delivery system using studies ranging from cellular association, cellular efflux and transmission electron microscopy (TEM). Ultrastructural studies clearly showed PS ODNs in the absence of liposomal delivery to be sequestered within endosomal and lysosomal vesicular bodies indicative of endocytic uptake. ODNs were also visible, to a lesser extent, in the nucleus and cytoplasm. By employing DOSPA (2'-(1",2"-dioleoyloxypropyldimethyl-ammonium bromide)-N-ethyl-6-amidospermine tetra trifluoroacetic acid) and DOPE (dioleoylphosphatidylethanolamine) complex in a 3 : 1 ratio, as a delivery system for ODNs at a optimal lipid/DNA charge ratio of 1 : 1, the level of ODN cellular association was significantly increased by approximately 10-12 fold with a concomitant change in subcellular distribution of PS ODN. TEM studies indicated enhanced penetration of ODN within the cytosol and the cell nucleus with reduced presence in vesicular compartments. Efflux studies confirmed that cationic lipoplexes promoted entry of ODNs into 'deeper' cellular compartments, consistent with endosomal release. Optimised cationic lipoplexes improved cellular delivery of ODNs by enhancing cell association, uptake and by favourably modulating the intracellular trafficking and distribution of ODNs into non-vesicular compartments including the cytosol and nucleus.

Quinolinic acid inhibits DOI-induced head shakes in mice.

Quinolinic acid, a metabolite of tryptophan in the kynurenine pathway, inhibited the 5-HT(2A/2C) direct agonist (+/-)-1-(2,5-dimethoxy-4-iodophenyl)-2-amino propane hydrochloride (DOI) induced head shakes in mice in a dose-dependent manner. This effect was observed 10 min after intracerebroventricular (i.c.v.) administration of 0.0625-1 microg/mouse. 3-Hydroxyanthranilic acid, which is the immediate stable precursor of quinolinic acid in the kynurenine pathway, showed a similar effect 60-120 min after subcutaneous (s.c.) administration, without showing any effect 10 min after i.c.v. administration or 20-40 min after s.c. administration. These observations suggest that quinolinic acid can alter central effects mediated via 5-HT2A receptors.

The thyrotrophin-releasing hormone analogue MK771 induces tic-like behaviours: the effects of dopamine D1 and D2 receptor antagonists.

Thyrotrophin-releasing hormone (TRH) and its analogues induce tic-like behaviours in rodents such as blinking and forepaw licking. Changes in spontaneous blinking frequency are observed in several disease states with dopamine abnormalities and dopaminergic agents modulate blinking. We have therefore investigated the effects of dopamine D1 and D2 receptor antagonists on TRH analogue (1-pyro-2-aminoadipyl-L-histidyl-L-thiazolidine-4-carboxamide; MK771)-induced blinking and bouts of forepaw licking. MK771 (2.5 mg/kg)-induced blinking was not attenuated by the dopamine D1 receptor antagonists (+)-7-chloro-hydroxy-3-methyl-1-phenyl-2,3,4,5-tetrahydro(1H)-3-benzazep ine maleate (SCH23390) (0.01, 1.0 and 5.0 mg/kg) and ((-)-trans-6,7,7a,8,9,13b-hexahydro-3-chloro-2-hydroxy-N-methyl-5- H-benz[2,1b]azepine (SCH39166; 1.0 and 5.0 mg/kg) or the dopamine D2 receptor antagonists raclopride (3.0 and 5.0 mg/kg) and sulpiride (5.0 and 10.0 mg/kg). D1 but not D2 receptor antagonists attenuated MK771-induced forepaw licking. MK771-induced blinking, therefore, appears not to involve dopamine D1 or D2 receptors and contrary to previously held belief dopamine does not appear to be pivotal in the control of blinking, while MK771-induced forepaw licking is modulated by dopamine D1 but not D2 receptors.

Similarities in the pharmacology of spontaneous and DOI-induced head-shakes suggest 5HT2A receptors are active under physiological conditions.

Nine monoamine receptor antagonists have been compared for their potency to inhibit both spontaneously occurring and DOI ((1-)2,5-dimethoxy-4-iodophenyl)-2-aminopropane)-induced head-shakes (HS). Ritanserin, ketanserin, prazosin, haloperidol, pimozide, SCH 23390 and SCH 39166 potently and dose-dependently antagonised both types of HS while sulpiride and raclopride produced weak and partial antagonism. The potency of these agents to inhibit spontaneous HS and DOI-induced HS was closely correlated (r = 0.94) and was significantly related to 5HT2A receptor and to alpha 1-adrenoceptor affinities taken from published sources. Potency was independent of affinity for D2 receptors but there was a possible influence of D1 receptor affinity. HS have been proposed to model Tourette's Syndrome; thus the present findings may have implications for the mechanism of action of antipsychotic agents in this condition and possibly also in schizophrenia. Contrary to previous suggestions, 5HT2A receptors may be tonically activated under physiological conditions.

Increased plasma kynurenine and its relationship to neopterin and tryptophan in Tourette's syndrome.

Fasting plasma levels of tryptophan, kynurenine and the pteridines, neopterin and tetrahydrobiopterin were measured in seven patients with Gilles de la Tourette syndrome (GTS) and 10 healthy controls. Plasma kynurenine was significantly elevated in the GTS patients. The lowest patient value was higher than the highest control value. Values for tryptophan, neopterin and tetrahydrobiopterin were similar in TS patients and controls. However, in TS patients only, there was a significant negative correlation between tryptophan and neopterin and a significant positive correlation between kynurenine and neopterin when controlling for tryptophan. This finding indicates that activation of cellular immune processes is a possible explanation for the rise in plasma kynurenine.

5-HT systems and anxiety: multiple mechanisms in the elevated X-maze.

Although much evidence supports the proposal that 5-HT neurones promote anxiety, there is also substantial evidence that the actual situation is more complex. The recent idea that 5-HT neurones, alerted by the forebrain, also function to restrain more primitive brainstem mechanisms has additional explanatory power but neither proposal accounts for the lack of responsiveness of 5-HT neurones to aversive stimulation from the environment. Evidence from the elevated X-maze anxiety model indicates that it is sensitive to multiple anxiety mechanisms which are not fully accounted for by either hypothesis. In addition, findings presented here indicate that open arm preference, risk assessment and overall motility load on separate factors in a factor analysis and that the anxiogenic action of 8-OH-DPAT under control conditions is a selective effect on open arm preference. This anxiogenic effect can be switched to anxiolytic at will by simple changes in experimental conditions. The outcome of 5-HT system manipulations on experimental anxiety may be the result of unstable balance between effects on different brain areas. It is suggested that 5-HT neurones do not carry information about threats in the environment, instead, this information is input locally where it modulates 5-HT release; this released 5-HT can have different consequences in different brain areas.

Discriminative stimulus produced by the imidazoline I2 site ligand, 2 -BFI.

2-(2-Benzofuranyl)-2-imidazoline, RX801077 (2-BFI) which has high affinity for imidazoline I(2) binding sites and very low aflinity for α(2)-adrenoceptors, has been investigated for its ability to produce a discriminative stimulus (cue) in drug-discrimination studies in rats since the existence of such a cue could assist in determining the functionality of I(2) sites. All rats subjected to training proved able to discriminate the training dose of 2-BFI (33 µmol/kg i.p) from saline vehicle and lower (5-14 µmol/kg) doses exhibited dose-dependent substitution. The mixed α(2)-adrenoceptor/I( 2) site ligand idazoxan fully substituted at 40µmol/kg. However, ethoxy idazoxan (11 µmol/kg) and fluparoxan (13 µmol/kg), selective α( 2)-adrenoceptor antagonists, also fully substituted for 2- BFI as did the monoamine oxidase (MAO) inhibitors moclobemide (99 µmol/kg) and pargyline (153 µmol/kg). A lower dose of moclobemide (16 µmol/kg) exhibited partial substitution. The α( 2)-adrenoceptor agonists clonidine (0.1 µmol/kg) and guanabenz (1.4 µmol/kg), and the benzodiazepine diazepam (14 µmol/kg), failed to substitute for 2-BFI indicating cue specificity. However, 2-BFI (14-50 µmol/kg) substituted partially but dose-dependently for clonidine (0.1 µmol/kg) in rats trained to distinguish the latter from saline. Changes in rates of response were independent of the degree of substitution. The observed pattern of drug substitution is consistent with the previously reported ability of 2-BFI to decrease MAO activity and thus increase extracellular monoamines.

5-Hydroxytryptamine pathways in anxiety and its treatment.

The effects of manipulating 5-hydroxytryptamine (5HT) neuronal function in humans and in animals are reviewed. 5HT pathways do not have a unitary function in modulating anxiety. It is proposed that, rather than acting as input or output channels for brain aversive systems, these pathways provide information concerning waking/motor status, which is crucial to the organisation of appropriate responses to threat. Each terminal region can make use of this information in different ways. Globally, the influence of 5HT neurones on higher centres appears predominantly to facilitate information processing relevant to threat, while their major influence on brainstem centres may be a restraining one.

Discriminative stimulus properties of ethoxy idazoxan.

The technique of drug discrimination was used to examine the ability of the highly selective α(2)-adrenoceptor antagonist ethoxy idazoxan, which has negligible affinity for α( 1)-adrenoceptors or I(2) imidazoline receptors, to produce an interoceptive discriminable stimulus or cue in rats. Rats were trained to respond on one lever after receiving α-ethoxy idazoxan (2.5 mg/kg, i.p.) and on the opposite lever after saline vehicle. The ethoxy idazoxan cue appeared to be mediated by antagonists of central α(2)-adrenoceptors, on the basis that dose- related substitution was produced by the highly selective α(2)-adrenoceptor antagonists idazoxan (imidazoline), fluparoxan and 1-(2-pyrimidinyl) piperazine (1-PP) (both non-imidazoline) but not by clonidine, which acts as an agonist at this receptor, nor by the peripherally acting α(2)-adrenoceptor antagonist L659,066. However, the α(2)-adrenoceptor antagonists yohimbine and atipamezole showed partial and non-dose-dependent substitution for ethoxy idazoxan over a wide range of doses. 2-BFI [2-(2-benzfuranyl)-2-imidazoline, RX801077], an imidazoline which is highly selective for I(2) imidazoline receptors over α(2)-adrenoceptors, showed dose- dependent substitution for ethoxy idazoxan, although the maximum effect (73% responding on the ethoxy idazoxan lever) fell short of criteria adopted for full substitution. Among other agents which bind to I(2) receptors, only idazoxan and 2-phenyl-2-imidazoline exhibited significant substitution; cirazoline could only be tested at very low doses because it powerfully inhibited responding in general, probably due to its α(1)-adrenoceptor agonist properties. It is suggested that the ability of 2-BFI to substitute partially for ethoxy idazoxan might be due to the ability of both agents to increase extracellular concentrations of noradrenaline.

Effects of two stressors on behaviour in the elevated X-maze: preliminary investigation of their interaction with 8-OH-DPAT.

Effects of water deprivation and restraint were compared in the rat elevated X-maze. Water deprivation for 12-48 h increased corticosterone and had a duration-dependent "anxiolytic" effect in the elevated X-maze, increasing the ratio of open/total arm entries (OTR) and the proportion of time spent on the open arms (% time) without affecting total entries. Brain 5HIAA/5HT was increased only after 24 or 48 h deprivation. Restraint for 15 min also increased plasma corticosterone and brain 5HIAA/5HT but had no effect on behaviour in the elevated X-maze when rats were tested immediately afterwards. However, 1 h restraint was "anxiogenic" in the elevated X-maze immediately after release, reducing OTR and % time, but with a less consistent reduction in total entries; reductions in OTR and % time were still present 24 h later. The 5HT1A agonist 8-hydroxy-2-(di-n-propylamino) tetralin (8-OH-DPAT) (0.1-0.2 mg/kg), administered 10 min before testing in the elevated X-maze, had "anxiogenic" actions in non-stressed rats. The effect of 0.1 mg/kg 8-OH-DPAT was not significantly altered by 24 or 48 h water deprivation but was abolished by restraint for 1 h immediately beforehand, despite the "anxiogenic" effect of restraint alone. Similar mutual antagonism of 8-OH-DPAT and restraint occurred when the dose of 8-OH-DPAT was increased to 0.2 mg/kg. Twenty-four hours after restraint, restrained rats which had received 8-OH-DPAT (0.1-0.2 mg/kg) still did not show any significant "anxiogenic effect" compared with non-restrained vehicle treated controls. Restraint-induced deficits in elevated X-maze exploration may prove a useful model with which to study the pharmacology of depression-related anxiety. However, the effects of the stressors examined, and their interaction with 8-OH-DPAT in the elevated X-maze, appear to depend on the nature of the stressor.

Elevated plasma kynurenine in Tourette syndrome.

Fasting plasma kynurenine concentrations were significantly elevated in a group of 7 patients (4 female) conforming to DSM-III-R of the American Psychiatric Association (1987) criteria for Tourette syndrome, in comparison with 10 healthy controls (7 female). Simultaneous normal plasma biopterin and neopterin concentrations indicated that this rise was probably not a consequence of peripheral cellular immune mechanisms.

Complex of rapid movements and vocalization induced by RX336-M in mice: possible relevance to Tourette's syndrome.

RX336-M (7,8-dihydro-5',6'-dimethylcyclohex-5'-eno-1',2',8',14 codeinone) induced a complex of simultaneous rapid movements of body parts accompanied by a single squeak-vocalization, which occurred at irregular intervals, when injected i.p. in mice. The complex included whole body jerks and head-shakes and its frequency was dose dependent between 0.1 and 20 mg/kg. Frequency after a 5 mg/kg dose was reduced by haloperidol (0.5-10 mg/kg, i.p.), ritanserin (0.1-10 mg/kg, i.p.) and ICI 169,369 (2.0 mg/kg, i.p.), suggesting the possible involvement of 5-HT(2A) and/or dopamine receptors. The possible relevance of the behavioural effects of RX336-M to the tics and vocalization of Tourette's syndrome is discussed.