Biomedical subjects

Jan Kehr

Publications and source records attributed to Jan Kehr.

At least 19 recordsLinked to original sources

PubMed2006

Determination of the dopamine agonist rotigotine in microdialysates from the rat brain by microbore column liquid chromatography with electrochemical detection.

Rotigotine, an investigational dopamine agonist formulated as a patch, is being studied in Parkinson's disease. A microdialysis technique, in combination with microbore column liquid chromatography and electrochemical detection, was developed to monitor rotigotine levels in the brain. Microdialysis probes were inserted into the striata of anesthetized rats, and samples were collected during perfusion with Ringer's solution. Rotigotine was separated using a C18 reversed-phase column. The mobile phase consisted of 50mM Na(2)HPO(4) x 2H(2)O, 2.5 mM sodium octyl sulfonate, and pH 4.5; 35% volume to volume acetonitrile. The flow rate was 30 microl/min, and the potential of the glassy carbon electrode was set to +850 mV. The method allowed monitoring of the time course of brain extracellular rotigotine levels with a detection limit of 1 nM following either intravenous (0.5 mg/kg) or subcutaneous (5.0 mg/kg) rotigotine injection.

Animals↗

PubMed2006

Galanin receptor antagonists : a potential novel pharmacological treatment for mood disorders.

The pathophysiology of mood disorders involves several genetic and social predisposing factors, as well as a dysregulated response to chronic stress. Accumulated evidence during the last two decades has implicated disturbances in brain serotonin and/or noradrenaline (norepinephrine) neurotransmission in the aetiology of depression. In fact, current pharmacological treatment for mood disorders is based on the use of drugs that act mainly by enhancing brain serotonin and noradrenaline neurotransmission by blockade of the active reuptake mechanism for these neurotransmitters. However, current antidepressant drugs have a delayed onset of therapeutic action, and a substantial number of patients do not respond adequately to them. In addition, these drugs have a number of adverse effects that limit patient compliance. In view of this, there is an intense search to identify novel (receptor) targets for antidepressant therapy. Recent studies have indicated that several neuropeptides and their receptors are potential candidates for the development of novel antidepressant treatment. In this context, galanin is of particular interest, since it is co-localised with serotonin in the dorsal raphe nucleus and with noradrenaline in the locus coeruleus, nuclei known to play a major role in affective disorders and in the action of antidepressant drugs. The actions of galanin are mediated by three receptor subtypes (GAL1, GAL2 and GAL3), which are coupled to different intracellular effector systems. Studies in rats have shown that galanin administered intracerebroventricularly is a potent inhibitor of mesencephalic serotonergic neurotransmission, as indicated by a long-lasting reduction in the release of serotonin in the hippocampus. This inhibitory effect is related to activation of the galanin receptors located on the dorsal raphe neurons. Moreover, intracerebroventricular galanin alters the gene expression of serotonin 5-HT1A autoreceptors in the dorsal raphe and also changes their functional activity. In addition, galanin produces a functional blockade of postsynaptic 5-HT1A receptor-mediated responses. Both pharmacological and genetic studies suggest a role for galanin in depression-like behaviour in rodent models. Transgenic mice overexpressing galanin under the control of the platelet-derived growth factor-beta promoter display increased immobility in the forced swim test. Intracerebroventricular administration of galanin in the rat increases depression-like behaviour, and this is fully blocked by the nonselective peptide galanin receptor antagonist M35. Importantly, M35 alone administered intracerebroventricularly produces an antidepressant-like effect. Recently, newly developed receptor-specific nonpeptidergic galanin GAL3 receptor antagonists (SNAP-37889 and SNAP-398299), which cross the blood-brain barrier after systemic administration, have shown antidepressant-like activity in several animal models. On the other hand, stimulation of the GAL2 receptor at the raphe level by local application of the GAL2 receptor agonist galanin (2-11) has been shown to increase serotonin levels in the hippocampus and dorsal raphe. These results indicate an important (mainly inhibitory) role of galanin as a regulator of brain serotonin and 5-HT1A receptor-mediated transmission, which may be of potential importance for understanding mood disorders and for the development of antidepressant drugs. Taken together, the present evidence suggests that antidepressant efficacy may be associated with compounds acting as antagonists at the GAL3 and/or possibly GAL1 receptors, and/or agonists at the GAL2 receptor.

Animals↗

PubMed2005

Protection but maintained dysfunction of nigral dopaminergic nerve cell bodies and striatal dopaminergic terminals in MPTP-lesioned mice after acute treatment with the mGluR5 antagonist MPEP.

The mGluR5 antagonist MPEP was used to study the role of mGluR5 in MPTP-induced injury of the nigrostriatal DA neurons. The findings indicate that acute blockade of mGluR5 may result in neuroprotective actions against MPTP neurotoxicity on nigral DA cell bodies and striatal DA terminals using stereological analysis of TH immunoreactivity and microdensitometry. Biochemical analysis showed no restoration of DA levels and metabolism indicating a maintained reduction of DA transmission.

Animals↗

PubMed2004

High-sensitive liquid chromatographic method for determination of neuronal release of serotonin, noradrenaline and dopamine monitored by microdialysis in the rat prefrontal cortex.

A high-sensitive liquid chromatographic method based on precolumn derivatization and fluorescence detection allowing simultaneous determination of serotonin (5-HT), noradrenaline (NA) and dopamine (DA) in brain microdialysis samples is described. 5-HT, NA and DA were derivatized with benzylamine and 1,2-diphenylethylenediamine in the presence of potassium hexacyanoferrate(III) and glycine, which yielded to highly fluorescent and stable benzoxazoles. The derivatized samples were separated on a microbore column (150 mm x 1.0mm i.d., packed with C18 silica, 5 microm) within 60 min. The mobile phase consisted of acetonitrile-Briton-Robinson buffer (pH 7.2) (32:68, v/v) containing 5 mM Na2EDTA and 5 mM octanesulfonic acid sodium salt. The detection limits (signal-to-noise ratio of 3) for 5-HT, NA and DA were 76, 42 and 95 amol/10 microl injected on-column, respectively. Microdialysis samples were collected at 10-min intervals from the probes implanted in the prefrontal cortex of awake rats. The basal levels of 5-HT, NA and DA were 7.3 +/- 0.7, 5.3 +/- 0.31 and 8.1 +/- 0.47 fmol/5 microl (mean +/- S.E.M., n = 5). Following 90-min perfusion with tetrodotoxin (1 microM) or calcium-free Ringer solution, the DA and NA levels were reduced to about 15 and 20%, respectively and the 5-HT levels to 45 and 60% of the basal levels, respectively. Reserpine, 12h after a dose of 5mg/kg i.p., reduced the extracellular 5-HT, NA and DA concentrations to about 34, 39 and 32% of the basal levels, respectively. In conclusion, the preset microdialysis/analytical method enables simultaneous monitoring of basal and pharmacologically reduced neuronal release of 5-HT, NA and DA in the rat brain.

Adrenergic Uptake Inhibitors↗

PubMed2004

Application of triple-probe microdialysis for fast pharmacokinetic/pharmacodynamic evaluation of dopamimetic activity of drug candidates in the rat brain.

The technique of microdialysis utilizing three simultaneously implanted probes in the anaesthetized rat enables monitoring of pharmacokinetic (PK) profiles of a tested drug both in blood (1st probe) and brain (2nd probe) compartments and the pharmacodynamic (PD) response of neurotransmitters (3rd probe) released into, or accumulating within the brain extracellular fluid (ECF). In the present study, the PK/PD characteristics of cocaine (psychostimulant, strong abuse potential) and methylphenidate (dopamimetic drug without reinforcing properties) and two novel NeuroSearch (NS) drug candidates, NS-A and NS-B, were examined in blood and brain microdialysates from the anaesthetized rats. The extracellular levels of dopamine (DA) were monitored in the striatum or prefrontal cortex. The NS-A compound entered the brain ECF at a slightly slower rate then methylphenidate; however, both compounds showed about the same effect on the speed of accumulation of extracellular DA concentrations, which gradually increased to about 450% of the basal, predrug levels at the end of the sampling period (180 min). The NS-B compound showed more rapid PK profiles than those observed after methylphenidate and NS-A. The concentrations of NS-B reached the maximal values already 40 min after its administration, while at that time, the corresponding DA values were still unchanged. In fact, the increase in DA concentrations was about two times slower when compared to that of methylphenidate or NS-A-drugs. Faster kinetics of NS-B and its delayed effect on extracellular DA suggests that this compound is metabolized to an active intermediate product, which itself exerts stronger dopamimetic activity in the rat prefrontal cortex that the original NS-B substance. The present study illustrates the feasibility of triple-probe microdialysis to monitor the rate of extracellular accumulation of a drug candidate and DA levels in vivo and compare the resulting PK/PD profiles to those obtained for cocaine and methylphenidate. These measures may serve as initial neurochemical indicators of potential psychomimetic or reinforcing properties of the tested substances.

Animals↗

PubMed2004

Determination of serotonin, noradrenaline, dopamine and their metabolites in rat brain extracts and microdialysis samples by column liquid chromatography with fluorescence detection following derivatization with benzylamine and 1,2-diphenylethylenediamine.

A highly selective and sensitive column liquid chromatographic method for fluorescence determination of serotonin (5-HT), dopamine (DA), noradrenaline (NA) and their related metabolites 5-hydroxyindole-3-acetic acid (5-HIAA) and 3,4-dihydroxyphenylacetic acid (DOPAC) following derivatization with benzylamine and 1,2-diphenylethylenediamine (DPE) is described. The monoamines and the metabolites (20 microl samples) were derivatized in a two-step reaction, initiated with 20 microl of 0.3M benzylamine in 0.3M 3-cyclohexylaminopropanesulfonic acid (CAPS) buffer (pH 10.0), (for 5-HT, 5-HIAA, 2 min, 24 degrees C) and followed by 20 microl of 0.1M DPE in 0.3M glycine buffer (pH 10.0), (for DA, NA, DOPAC, 20 min, 50 degrees C). Both reagents contained 0.02 M potassium hexacyanoferrate(III) and 50% (v/v) methanol. The resulting highly fluorescent and stable benzoxazole derivatives were isocratically separated on a reversed-phase column (150 mm x 1.5 mm i.d., packed with C18 silica, 5 microm) within 45 min. Using fluorescence detection at ex. and em. wavelengths of 345 and 480 nm, respectively, the detection limit (signal-to-noise ratio of 3) for 5-HT, DA, NA, 5-HIAA, L-DOPA and DOPAC ranged between 0.08 and 5.65 fmol per 20-microl injection (12-847.5 pM in standard solution). The concentrations of monoamines (expressed in microg/g wet weight, mean +/- S.E.M., n=5) in tissue extracts from the rat striatum were: 0.45+/-0.05 (5-HT), 4.27+/-0.08 (DA), 0.27+/-0.04 (NA), 0.55+/-0.06 (5-HIAA), 1.26+/-0.16 (L-DOPA) and 1.62+/-0.11 (DOPAC). Microdialysis samples were collected in 20 min intervals from the probes implanted in the striatum of awake rats. The basal monoamine levels (in fmol/20 microl, mean +/- S.E.M., n=5) in the dialysates were: 4.0+/-0.7 (5-HT), 78.4+/-9.1 (DA), 6.4+/-0.8 (NA), 785.5+/-64.5 (5-HIAA) and 5504.5+/-136.5 (DOPAC). It is concluded that the new fluorescence derivatization protocol provides an excellent means for simultaneous determination of all three monoamines both in the complex samples (e.g. brain homogenates) and also at trace levels, such as those found in the microdialysis samples.

Animals↗

PubMed2004

Hypericum perforatum L (St John's wort) preferentially increases extracellular dopamine levels in the rat prefrontal cortex.

The effects of hydro-alcoholic extracts of Hypericum perforatum L on extracellular serotonin (5-HT), noradrenaline (NA) and dopamine (DA) levels and the acidic metabolites (3,4-dihydroxyphenylacetic acid (DOPAC), homovanillic acid (HVA) and 5-hydroxy-3-indoleacetic acid (5-HIAA)) were examined by in vivo microdialysis in the prefrontal cortex of awake rats. Thus, a single dose (60 mg kg(-1) i.p. or 300 mg kg(-1) p.o.) of H. perforatum increased DA concentrations to 165 and 140% of control values, respectively, and increased locomotor activity in nonhabituated rats. DOPAC and HVA levels were markedly reduced. 5-HT concentrations were elevated only moderately, while the NA levels were not affected by any treatment. The whole-tissue analysis revealed that hypericum increased, whereas the monoamine oxidase (MAO) A/B inhibitor phenelzine decreased DA and 5-HT turnover. The present data indicate that the mechanism of action of hypericum extract in vivo is more complex than the inhibition of monoamine reuptake or metabolism observed in vitro. The finding of preferential enhancement of DA transmission is in agreement with human studies measuring DA-mediated neuroendocrine responses.

Animals↗

PubMed2004

Differential effects of (R)-, (R, S)- and (S)-8-hydroxy-2-(di-n-propylamino)tetralin on hippocampal serotonin release and induction of hypothermia in awake rats.

The effects of (R)- and (S)-optical isomers of 8-hydroxy-2-(di-n-propylamino)-tetralin (8-OH-DPAT) and of the racemate (R,S)-8-OH-DPAT on serotonin (5-HT) release in the ventral hippocampus of awake rats and on induction of the whole-body hypothermia were studied. Extracellular 5-HT levels were determined by a newly developed high-sensitive HPLC method based on derivatization with benzylamine and fluorescence detection. The basal levels of 5-HT in 20 min microdialysates from rats perfused with Ringer solution or with Ringer solution containing 1 microM citalopram were 6.3 +/- 1.3 fmol/20 microl and 36.1 +/- 4.2 fmol/20 microl (n=20), respectively. The reduction of hippocampal 5-HT levels induced by subcutaneous (s.c.) administration of (R,S)-8-OH-DPAT (0.3 mg/kg) was significantly attenuated by the presence of 5-HT reuptake inhibitor citalopram in Ringer solution only at its peak value at 40 min (maximal reduction to 60% compared to 46% of control values in Ringer-perfused rats), whereas the overall effects were comparable at both experimental conditions. Injection of (R)-8-OH-DPAT (0.3 mg/kg s.c.) caused further reduction of 5-HT levels, to 49% and 41%, respectively, whereas (S)-8-OH-DPAT (0.3 mg/kg s.c.) caused maximal reduction of 5-HT levels only to 74% of controls in both perfusion groups. Similar pattern and time-courses were observed in rats with hypothermia induced by injection of 8-OH-DPAT enantiomers, where (R,S), (R)-forms were about two-times more potent than the (S)-isomer. It is concluded that the acute systemic dose of (R)-, (S)- and (R,S)-8-OH-DPAT enantiomers exerted enantiomer-specific effects on 5-HT(1A) receptor-mediated function both at the presynaptic and postsynaptic sites as revealed by monitoring hippocampal 5-HT levels and body temperature.

8-Hydroxy-2-(di-n-propylamino)tetralin↗

PubMed2003

Enhanced hippocampal noradrenaline and serotonin release in galanin-overexpressing mice after repeated forced swimming test.

Basal and forced swimming (FS) stress-induced release of noradrenaline (NA) and serotonin (5-HT) were determined by in vivo microdialysis in the ventral hippocampus of mice overexpressing galanin under the platelet-derived growth factor B promoter (GalOE/P) or under the dopamine beta-hydroxylase promoter (GalOE/D) (only NA). WT mice served as controls. Intraventricular infusion of galanin significantly reduced basal extracellular NA in WT mice and in GalOE/P mice (albeit less so). Microdialysis sampling during a 10-min FS showed that NA and 5-HT release were elevated to 213% and 156%, respectively, in the GalOE/P group, whereas in the WT group the increases were only 127% and 119%, respectively. The second (repeated) 10-min FS (RFS) caused a marked enhancement of NA and 5-HT release in the GalOE/P mice to 344% and 275%, respectively. However, the RFS caused only a 192% increase of extracellular NA levels in the GalOE/D mice. Pretreatment with the putative peptidergic galanin receptor antagonist M35 almost completely blocked the elevation of NA and 5-HT levels in the GalOE/P after RFS. These results suggest that the NA and 5-HT hippocampal afferents in GalOE/P mice are hypersensitive to both conditioned and unconditioned stressful stimuli, such as FS, and that this effect is mediated by galanin receptors. The present findings support a role of galanin in the regulation of release of NA and 5-HT, two neurotransmitters involved in mood control.

Animals↗

PubMed2003

Simultaneous determination of 5-hydroxyindoles and catechols by high-performance liquid chromatography with fluorescence detection following derivatization with benzylamine and 1,2-diphenylethylenediamine.

A highly selective and sensitive method for the simultaneous determination of 5-hydroxyindoles and catechols (serotonin, norepinephrine, dopamine and related compounds) by high-performance liquid chromatography with fluorescence detection is described. The method is based on the two-step precolumn derivatization of 5-hydroxyindoles and catechols with benzylamine (BA) and 1,2-diphenylethylenediamine (DPE), respectively, resulting in highly fluorescent and stable benzoxazole derivatives. The first derivatization with BA proceeds at room temperature (ca. 23 degrees C) for 2 min in a mixture of 0.3 M 3-cyclohexylamino-1-propanesulfonic acid buffer (pH 10.0) and methanol in the presence of potassium hexacyanoferrate(III). The subsequent second derivatization with DPE is carried out at 50 degrees C for 20 min in the presence of glycine. The resulting fluorescent derivatives of five 5-hydroxyindoles and seven catechols are separated on a reversed-phase column (150 x 1.5 mm I.D., packed with C18 silica, 5 microm) with isocratic elution using a mixture of acetonitrile-15 mM acetate buffer (pH 4.5) (34:66, v/v) containing 1 mM octanesulfonic acid sodium salt, and are detected spectrofluorimetrically at 480 nm with excitation at 345 nm. The detection limits (signal-to-noise ratio of 3) of the related compounds are 80 amol to 86 fmol for a 20-microl injection.

Benzylamines↗

PubMed2003

Determination of histamine in microdialysis samples from rat brain by microbore column liquid chromatography following intramolecular excimer-forming derivatization with pyrene-labeling reagent.

This paper describes a sensitive and selective liquid chromatographic method with fluorescence detection for determination of histamine in brain microdialysis samples from awake rats. Samples containing histamine (10 microl) were derivatized with 20 microl of the reagent consisting of 3 mM 4-(1-pyrene)butyric acid N-hydroxysuccinimide ester (PSE), 3 mM potassium carbonate and acetonitrile (1:1:18, v/v), thereafter 20 microl volume was injected onto the microbore column packed with C18 silica gel. The histamine derivative contained two pyrene moieties, which generated intramolecular excimer fluorescence (450-540 nm) and allowed clear discrimination from the monomer fluorescence (360-420 nm) emitted by PSE itself. The separation of histamine-pyrene derivative was achieved within 25 min, the detection limit (S/N=3) was 0.3 fmol histamine in 20 microl injected. The basal extracellular levels of histamine collected in 10-min fractions (fmol per 10 microl, mean+/-S.D., not corrected for recovery, n=10 rats) were 35.45+/-4.56 (hypothalamus), 9.05+/-1.56 (prefrontal cortex), 7.83+/-0.86 (hippocampus) and 6.54+/-0.66 (striatum). The voltage-sensitive release of histamine was evaluated by perfusing the probes with high (100 mM) concentration of potassium ions or with sodium channel blocker tetrodotoxin (1 microM), and the calcium-dependent release was tested by perfusion with calcium-free Ringer solution. These data, together with physiologically induced increase of extracellular histamine in four examined brain regions during forced swimming demonstrate that this method is suitable for high-sensitive determination of neuronally released histamine under various pharmacological and physiological conditions.

Affinity Labels↗

PubMed2003

Galanin attenuates basal and antidepressant drug-induced increase of extracellular serotonin and noradrenaline levels in the rat hippocampus.

Galanin is co-localized with classical neurotransmitters, such as acetylcholine, serotonin (5-HT) and noradrenaline (NA) in neurons or in brain regions implicated in cognitive and affective behaviour. In the present study, the effects of galanin on extracellular 5-HT and NA levels in the rat hippocampus were measured by in vivo microdialysis under basal conditions and following systemic administration of antidepressant drugs. Galanin (1.5 nmol i.c.v.) reduced basal 5-HT and NA levels to 65% and 86% of controls, respectively. Galanin (0.5 and 1.5 nmol i.c.v.) dose-dependently attenuated the elevation of 5-HT concentrations induced by imipramine and citalopram (10 mg/kg i.p., each) from 350% to 312% and from 230% to 160%, respectively. Galanin at 1.5 nmol transiently attenuated the effect of desipramine-induced (10 mg/kg i.p.) increase in extracellular NA levels from a maximal increase of 389-296% of the predrug levels. It is concluded that intraventricularly administered galanin attenuated both basal 5-HT and NA release and antidepressant drug-induced accumulation of extracellular 5-HT and NA levels most likely via a predominant inhibitory action on serotonergic and noradrenergic neurons in the raphe and locus coeruleus, respectively. These results further emphasize a possible role of galanin in regulation of 5-HT and NA neurotransmission in depressive states and during the course of antidepressant therapy.

Animals↗

PubMed2003

Simultaneous determination of norepinephrine, serotonin, and 5-hydroxyindole-3-acetic acid in microdialysis samples from rat brain by microbore column liquid chromatography with fluorescence detection following derivatization with benzylamine.

A microbore column liquid chromatographic method for the simultaneous determination of norepinephrine (NE), serotonin (5-HT), and 5-hydroxyindole-3-acetic acid (5HIAA) in microdialysis samples from rat brain is described. The method is based on precolumn derivatization of NE, 5HT, and 5HIAA with benzylamine in the presence of potassium hexacyanoferrate(III) resulting in the corresponding highly fluorescent and stable benzoxazole derivatives. A 15-microl sample was mixed with 15 microl derivatization reagent solution containing 0.3M 3-cyclohexylaminopropanesulfonic acid buffer (pH 12.0), 0.5M benzylamine, 10mM potassium hexacyanoferrate(III), and methanol (1/1/1/12, v/v/v/v). The derivatization was carried out at 50 degrees C for 20 min. The benzylamine derivatives of NE, 5HT, and 5HIAA were separated on a reversed-phase column (100 x 1.0mm i.d., packed with C18 silica, 5 microm) within 30 min. The mobile phase consisted of 15 mM acetate buffer (pH 5.0) and acetonitrile (31%, v/v); the flow rate was 50 microl/min. The detection limits (signal-to-noise ratio of 3) for NE, 5HT, and 5HIAA in the injection volume of 20 microl were 90, 210, and 260 amol, respectively. Microdialysis samples were collected in 7.5-min intervals from the probes implanted in the hippocampus and prefrontal cortex of awake rats. The basal levels of NE, 5HT, and 5HIAA in the dialysates from the hippocampus were 4.2+/-0.5, 4.9+/-0.6, and 934.1 +/- 63.4 fmol/20 microl, and those from the prefrontal cortex were 6.0+/-1.2,5.51.3, and 669.1 +/- 96.0 fmol/20 microl (mean +/- SE, n=25), respectively. The NE and 5HT levels were altered by perfusion of high-potassium or low-calcium solution and following antidepressant drugs imipramine and desipramine. It is concluded that the new fluorescence derivatization method in combination with microbore column liquid chromatography allows the simultaneous determination of NE, 5HT, and 5HIAA in the microdialysis samples at higher sensitivity, providing easier maintenance in routine use than that achieved by high-performance liquid chromatographic methods with electrochemical detection.

Animals↗

PubMed2003

A sensitive and selective determination method of histamine by HPLC with intramolecular excimer-forming derivatization and fluorescence detection.

A highly sensitive, selective and simple method is described for the determination of histamine by high-performance liquid chromatography (HPLC) with fluorescence detection. The method is based on an intramolecular excimer-forming fluorescence derivatization of histamine with 4-(1-pyrene)butyric acid N-hydroxysuccinimide ester (PSE), followed by reversed-phase HPLC. Histamine, having two amino moieties in a molecule, was converted to the dipyrene-labeled derivative by reaction with PSE. The derivative afforded intramolecular excimer fluorescence (450-540 nm), which can clearly be discriminated from the monomer fluorescence (370-420 nm) emitted from PSE. Typically, a 10 micro L sample solution was mixed with 100 micro L of derivatization reagent solution, which was a mixture of 0.5 mm PSE in acetonitrile and 0.5 mm potassium carbonate in water (8:2, v/v). The derivatization was carried out at 100 degrees C for 90 min. The PSE derivative of histamine could be separated by reversed-phase ODS column with isocratic elution using acetonitrile:water (82:18, v/v) containing 0.03% triethylamine. The detection limit (singnal-to-noise ratio = 3) of histamine was 0.5 fmol for a 30 micro L injection. The method was successfully applied to the determination of histamine in human urine, and had enough selectivity and sensitivity for urinary histamine quantification.

Adult↗

PubMed2002

Increased serotonin release in mice frontal cortex and hippocampus induced by acute physiological stressors.

The effects of acute physiological stressors (5 s tail pinch, handling and forced swimming at +25 and +5 degrees C for 3 min each) on serotonin (5-HT) release in the mouse brain were investigated using in vivo microdialysis. The extracellular 5-HT levels were determined by a newly developed highly-sensitive and selective high-performance liquid chromatography method based on derivatization with benzylamine and fluorescence detection. The basal levels of 5-HT in 3 min microdialysates from the ventral hippocampus and frontal cortex were 0.68+/-0.21 and 0.75+/-0.28 fmol/6 microl (n=24), respectively. All three stressors caused an immediate, significant and reversible increase (handling: 150%; swimming: 240%) of extracellular 5-HT levels in both brain structures, suggesting a more dynamic role played by the serotonergic system in response to acute stress.

Acute Disease↗

PubMed2002

Galanin is a potent in vivo modulator of mesencephalic serotonergic neurotransmission.

Neurochemical, molecular, immunohistochemical and behavioral methods were used to examine the in vivo effects of the neuropeptide galanin on central 5-HT neurotransmission and on 5-HT(1A) receptor-mediated responses. Intraventricularly infused galanin caused a long-lasting and dose-dependent reduction of basal extracellular 5-HT levels in the ventral hippocampus of awake rats as measured by microdialysis. Infusion of galanin into the dorsal raphe nucleus (DRN), but not intra-hippocampally, reduced 5-HT release. The effect of i.c.v. galanin on 5-HT release was blocked by the galanin receptor antagonist M35, acting most likely via galanin receptors at the level of the DRN. Galanin also reduced the levels of tryptophanhydroxylase mRNA in the DRN. Therefore, the effects of galanin on 5-HT(1A) receptor-mediated responses were further investigated. Surprisingly, galanin significantly attenuated the reduction of hippocampal 5-HT release induced by systemic injection of the 5-HT(1A) receptor agonist 8-OH-DPAT. Galanin also attenuated 8-OH-DPAT-induced hypothermia and locomotor activity in rats. These results indicate that galanin has important inhibitory actions on central 5-HT neurotransmission and on 5-HT(1A) receptor-mediated events.

8-Hydroxy-2-(di-n-propylamino)tetralin↗

PubMed2002

Quantitative dual-probe microdialysis: evaluation of [3H]mannitol diffusion in agar and rat striatum.

Dual-probe microdialysis was used to study interstitial diffusion in the rat brain. A radiolabelled tracer, (3H]mannitol, was continuously infused at different concentrations via a probe acutely implanted into the striatum of an anaesthetized male rat or into a dilute agar gel. Samples were collected by a second probe placed 1 mm away from the first, and the recovered [3H]mannitol was measured by liquid scintillation counting. In the striatum, the delivery of [3H]mannitol was counteracted by its removal from the extracellular space by passive uptake into cells and clearance into the microcirculation, causing the diffusion profile to approach quasi steady-state levels within 2 h. Diffusion data from brain and agar were analysed using a mathematical model. The apparent (effective) diffusion coefficient for [3H]mannitol was D* = 2.9 x 10(-6) cm2/s, the effective volume fraction alpha* = 0.30 and the clearance rate constant kappa= 2.3 x 10(-5)/s. A tortuosity, lambda = 1.81, and penetration distance r = 4.2 mm, were calculated. We conclude that, using dual-probe microdialysis, parameters reflecting geometric and dynamic tissue properties may be obtained using appropriate mathematical analysis. Quantitative dual-probe microdialysis will be valuable in characterizing interstitial diffusion and the clearance processes underpinning volume transmission in the brain.

Agar↗

PubMed2002

Quantitative dual-probe microdialysis: mathematical model and analysis.

Steady-state microdialysis is a widely used technique to monitor the concentration changes and distributions of substances in tissues. To obtain more information about brain tissue properties from microdialysis, a dual-probe approach was applied to infuse and sample the radiotracer, [3H]mannitol, simultaneously both in agar gel and in the rat striatum. Because the molecules released by one probe and collected by the other must diffuse through the interstitial space, the concentration profile exhibits dynamic behavior that permits the assessment of the diffusion characteristics in the brain extracellular space and the clearance characteristics. In this paper a mathematical model for dual-probe microdialysis was developed to study brain interstitial diffusion and clearance processes. Theoretical expressions for the spatial distribution of the infused tracer in the brain extracellular space and the temporal concentration at the probe outlet were derived. A fitting program was developed using the simplex algorithm, which finds local minima of the standard deviations between experiments and theory by adjusting the relevant parameters. The theoretical curves accurately fitted the experimental data and generated realistic diffusion parameters, implying that the mathematical model is capable of predicting the interstitial diffusion behavior of [3H]mannitol and that it will be a valuable quantitative tool in dual-probe microdialysis.

Agar↗