Rat strain differences in peripheral and central serotonin transporter protein expression and function

Female Fischer 344 (F344) rats have been shown to display increased serotonin transporter (5-HTT) gene expression in the dorsal raphe, compared to female Lewis (LEW) rats. Herein, we explored, by means of synaptosomal preparations and in vivo microdialysis, whether central, but also peripheral, 5-HTT protein expression/function differ between strains. Midbrain and hippocampal [3H]paroxetine binding at the 5-HTT and hippocampal [3H]serotonin (5-HT) reuptake were increased in male and female F344 rats, compared to their LEW counterparts, these strain differences being observed both in rats of commercial origin and in homebred rats. Moreover, in homebred rats, it was found that these strain differences extended to blood platelet 5-HTT protein expression and function. Saturation studies of midbrain and hippocampal [3H]paroxetine binding at the 5-HTT, and hippocampal and blood platelet [3H]5-HT reuptake, also revealed significant strain differences in Bmax and Vmax values. Although F344 and LEW rats differ in the activity of the hypothalamo-pituitary-adrenal (HPA) axis, manipulations of that axis revealed that the strain differences in hippocampal [3H]paroxetine binding at 5-HTTs and [3H]5-HT reuptake were not accounted for by corticosteroids. Hippocampal extracellular 5-HT levels were reduced in F344 rats, compared to LEW rats, with the relative, but not the absolute, increase in extracellular 5-HT elicited by the local administration of citalopram being larger in F344 rats. Because the aforementioned strain differences did not lie in the coding sequences of the 5-HTT gene, our results open the promising hypothesis that F344 and LEW strains model functional polymorphisms in the promoter region of the human 5-HTT gene.     

Fox odour affects corticosterone release but not hippocampal serotonin reuptake and open field behaviour in rats

Group-housed Sprague-Dawley (SD) rats exposed for 1 h to 2,5-dihydro-2,4,5-trimethylthiazoline (TMT, a component of fox feces) did not display changes in hippocampal serotonin (5-HT) metabolism and [3H]5-HT reuptake, compared to water or butyric acid. Such an observation extended to isolated SD and Fischer 344 rats. When group-housed SD rats were tested 1 week after a 1-h exposure to TMT, hippocampal 5-HT metabolism, [3H]5-HT reuptake, and [3H]paroxetine binding at the 5-HT transporter remained unchanged. This study questions TMT as a specific predatory stimulus as both butyric acid and TMT increased plasma corticosterone levels whilst leaving intact open field behaviour (at least in group-housed SD rats).     

Behavioural, hyperthermic and neurotoxic effects of 3,4-methylenedioxymethamphetamine analogues in the Wistar rat

1. The ability of N-ethyl (MDEA) and N-butyl (MDBA) analogues of 3,4-methylenedioxymethamphetamine (MDMA, 'Ecstasy') to induce acute behavioural changes and increases in body temperature, and to cause serotonergic neurotoxicity, was assessed in young adult male Wistar rats. The in vitro ability of MDMA analogues to evoke presynaptic monoamine release from crude rat forebrain synaptosomal preparations pre-labelled with [3H]5-HT or [3H]DA was also measured. 2. In behavioural experiments, acute MDMA and MDEA (20 mg/kg, i.p.) significantly increased rat open-field locomotion scores, decreased open-field rearing, and induced stereotypy, Straub tail and head weaving. MDBA did not produce any of these behaviours. 3. After repeated dosing (8 x 20 mg/kg, i.p., twice daily for 4 days), MDMA > MDEA > MDBA > or = saline at decreasing forebrain [3H]paroxetine binding levels and concentrations of 5-HT and 5-HIAA at 14 days post-treatment. None of the analogues caused any long-term changes in dopamine or noradrenaline concentrations in the forebrain. 4. Acute MDMA and MDEA (20 mg/kg, i.p.) produced significant acute increases in rat aural temperature compared with saline-treated animals, while 20 mg/kg MDBA caused no significant effects. 5. MDA, MDMA and MDEA were equipotent at inducing [3H]5-HT release from frontal cortex/hippocampal synaptosomes, while MDBA only evoked a significant release at 100 microM concentrations. The potency order for inducing [3H]DA release from striatal synaptosomes was MDA > MDMA > MDEA = MDBA. 6. This study shows that large N-alkyl substitution decreases the ability of MDMA analogues to evoke presynaptic 5-HT and DA release, induce acute hyperthermia, hyperlocomotion and behavioural changes, and cause long-term serotonergic neurotoxicity. 7. The structure-activity relationship data presented here indicate that the neurotoxic damage caused by substituted amphetamines requires a combination of acute hyperthermia and increased neurotransmitter release. Induction of one of these effects in isolation is not sufficient to cause serotonergic nerve terminal degradation.     

Compensatory responses in the aging hippocampal serotonergic system following neurodegenerative injury with 5,7-dihydroxytryptamine

This study utilized a multidisciplinary approach to examine injury-induced compensatory responses in the aging hippocampal serotonin transporter (5-HTT), a membrane protein implicated in a variety of neurodegenerative disorders. Age-dependent cellular, anatomical, and physiological changes of the 5-HTT were evaluated in female Fischer 344 rats (2 and 17 months) following denervation of the serotonergic afferents (fimbria-fornix and cingulum bundle) to the dorsal hippocampus using the neurotoxicant 5,7-dihydroxytryptamine (5,7-DHT). Seven days following 5,7-DHT administration, a uniform loss of the hippocampal 5-HTT immunoreactivity was observed in both age groups. However, at 21 days 5-HTT immunoreactivity in young 5,7-DHT-treated animals was similar to control levels, indicative of recovery, while older animals exposed to 5,7-DHT did not show recovery of hippocampal 5-HTT expression. 5-HTT binding site density, as determined by quantitative autoradiography ([3H]citalopram), supported the immunohistochemical results by demonstrating a recovery of 5-HTT binding sites in young, but not old animals, at 21 days following the lesion (P < 0.001). Furthermore, cellular electrophysiological function of hippocampal CA1 pyramidal neurons in 3- and 18-month-old F344 rats at 21 days following 5,7-DHT or vehicle treatment were assessed using in vivo microiontophoretic application of serotonin (5-HT). Independent of changes in sensitivity to the inhibitory effects of 5-HT application, the time to recovery of cell firing following application of 5-HT was significantly increased in the 18-month 5,7-DHT group compared to the 18-month vehicle and 3-month 5,7-DHT groups (60 and 59% increases, respectively; P < 0.05). Overall, these series of studies comprise a model which can be used to identify cellular events underlying both the formation of injury-induced compensatory processes in younger animals and the lack thereof with advancing age.     

N-tert-butyl-alpha-phenylnitrone protects against 3,4-methylenedioxymethamphetamine-induced depletion of serotonin in rats

The present study examined the effect of N-tert-butyl-alpha-phenylnitrone (PBN) on 3,4-methylenedioxmathamphetamine (MDMA)-induced depletion of serotonin in the CNS. Rats were treated with two concurrent injections of MDMA (20 mg/kg, s.c.), PBN (50-400 mg/kg dissolved in ethanol, 50 mg/ml of 25% ethanol, i.p.), saline or 25% ethanol, alone or in combination, 6 h apart, and sacrificed 5 days later. Rectal temperature was measured prior to and hourly following the drug injection for 5 h. Monoamine levels in the tissue were measured by HPLC. Density of the 5-HT transporters was assayed by [3H]paroxetine binding. Rectal temperature of rats increased after MDMA, decreased after PBN, ethanol, PBN plus ethanol, and MDMA plus ethanol, and was not significantly altered after MDMA plus PBN. Levels of 5-HT and 5-HIAA in the frontal cortex, hippocampus, striatum, and brain stem of rats decreased significantly after MDMA or MDMA plus ethanol, but not after MDMA plus PBN, PBN plus ethanol (PBN dissolved in ethanol), or ethanol as compared to the saline controls. Levels of 5-HT and 5-HIAA in the brain tissues of rats treated with MDMA plus PBN were elevated as compared to those treated with MDMA plus saline. Similar results were observed in the density of 5-HT transporters in the frontal cortex and hippocampus. These results indicate that scavenging of free radicals of MDMA metabolites or reactive oxygen species by PBN and with lowering of body temperature protected against MDMA-induced depletion of serotonin transmitter.     

Reduced efficacy of fluoxetine following MDMA ("Ecstasy")-induced serotonin loss in rats

Long-term serotonin (5-HT) neuronal loss is currently a major cause of concern associated with recreational use of the substituted amphetamine 3,4 methylenedioxymethamphetamine (MDMA; "Ecstasy"). Such loss may be problematic considering that psychiatric disorders such as depression and anxiety and responses to first line treatments for these disorders are associated with 5-HT. In this study the effects of prior exposure to MDMA on behavioural and central neurochemical changes induced by the serotonin (5-HT) re-uptake inhibitor and antidepressant fluoxetine were examined in rats. Animals were administered MDMA (10 mg/kg. i.p.) four times daily for two consecutive days. One week later the animals were subjected to treatment with fluoxetine (10 mg/kg, i.p.). Fluoxetine treatment groups received either acute (saline injections for 20 days followed by 3 fluoxetine treatments over 24 h) or chronic (once daily fluoxetine for 21 days) drug administration. Prior exposure to MDMA resulted in an attenuation of fluoxetine-induced swimming behaviour in the modified forced swimming test (FST); a behavioural test of antidepressant action. In parallel MDMA treatment resulted in significant regional depletions of 5-HT and its metabolite 5-hydroxyindoleacetic acid (5-HIAA) accompanied by a reduction in cortical [3H] paroxetine binding to nerve terminal 5-HT transporters. MDMA-induced 5-HT loss was enhanced in animals following chronic fluoxetine administration. Elimination of fluoxetine and its metabolite norfluoxetine from the brain abolished this interaction between MDMA and fluoxetine treatment. Fluoxetine administration reduced both 5-HIAA and the 5-HIAA:5-HT metabolism ratio, which was attenuated in animals pre-treated with MDMA. Overall the results show that MDMA induces long-term 5-HT loss in the rodent brain and consequently diminishes behaviour and reductions in 5-HT metabolism induced by the antidepressant fluoxetine. These results have potential clinical relevance, suggesting that 5-HT re-uptake inhibitors such as fluoxetine may be less effective at treating depression in chronic abusers of MDMA.     

Corticosterone regulation of serotonin transporter and 5-HT1A receptor expression in the aging brain

Hypercortisolemia is often observed in patients suffering from major depression. As the serotonergic (5-hydroxytryptamine; 5-HT) system plays a major role in the etiology of depression, a loss of endocrine and neurotransmitter system interactions, including corticosterone regulation of 5-HT transporter (5-HTT) and 5-HT receptor expression, may underlie age-related deficits in the regulation of the hypothalamic-pituitary-adrenal (HPA) axis and correlate with an increased incidence of depression with advancing age. In this study, female Fischer 344 rats, ages 3, 13, and 18 months, were bilaterally adrenalectomized and supplemented for 3 weeks with corticosterone (0, 200, or 600 mg; LC, MC, or HC, respectively) containing 21 day sustained-release pellets implanted subcutaneously. Quantitative autoradiography of hippocampal and cortical regions using [3H]citalopram revealed a significant decrease in hippocampal 5-HTT binding in the 3-month HC treatment group compared to age-matched MC and LC groups; this loss was not present in the 13- or 18-month groups. Similarly, quantitative autoradiography using the radiolabeled 5-HT(1A) receptor agonist 8-hydroxy-2-(di-N-propylamino) tetralin demonstrated a significant decline in receptor density in 3- and 13-month MC and HC groups as compared to age-matched LC groups in the hippocampus. These hormone treatments (MC or HC), however, failed to alter hippocampal 5-HT(1A) binding site density in the 18-month groups as compared to the age-matched LC group. The 5-HT(2A) receptor was also evaluated using [3H]ketanserin and showed no age- or corticosterone-related changes in the cortex. Overall, an age-associated deficit in the regulation of the hippocampal serotonergic system by varied corticosterone treatment was revealed in the present study, which may underlie the increased incidence of depression and hypercortisolemia found with advancing age.     

Occupancy of serotonin transporters by paroxetine and citalopram during treatment of depression: a [(11)C]DASB PET imaging study.

OBJECTIVE: Selective serotonin reuptake inhibitors are commonly used to treat major depression; however, the percentage of serotonin (5-HT) transporter (5-HTT) sites occupied during clinical dosing is unknown. This study measured the proportion of 5-HTT sites blocked during paroxetine and citalopram treatment of depression and assessed the relationship between serum paroxetine levels and the proportion of 5-HTT sites blocked. METHOD: Twelve medication-free depressed patients completed a 6-week trial of either paroxetine (N=8) or citalopram (N=4). Striatal 5-HTT binding potential was measured with [(11)C]DASB and positron emission tomography, before and after 4 weeks of treatment. The binding potential is proportional to receptor density. Striatal 5-HTT binding potential was measured twice in six healthy subjects and once in 11 healthy subjects. RESULTS: A significant decrease in striatal 5-HTT binding potential was found after either treatment, compared to changes found over a 4-week period in healthy subjects. For patients treated with 20 mg/day of paroxetine (N=7), the mean proportion of 5-HTT sites occupied was 83%. For patients treated with 20 mg/day of citalopram (N=4), the mean 5-HTT occupancy was 77%. 5-HTT occupancy increased in a nonlinear relationship with serum levels of paroxetine such that a plateau of occupancy around 85% occurred for serum paroxetine levels greater than 28 microg/liter. CONCLUSIONS: During treatment with clinical doses of paroxetine or citalopram, approximately 80% of 5-HTT receptors are occupied. This change in 5-HTT binding potential is greater than the known physiological range of changes in 5-HTT binding potential but may be necessary for some therapeutic effects.     

Persistent cerebrovascular effects of MDMA and acute responses to the drug

Acutely, 3,4,-methylenedioxymethamphetamine (MDMA) induces cerebrovascular dysfunction [Quate et al., (2004)Psychopharmacol., 173, 287-295]. In the longer term the same single dose results in depletion of 5-hydroxytrptamine (5-HT) nerve terminals. In this study we examined the cerebrovascular consequences of this persistent neurodegeneration, and the acute effects of subsequent MDMA exposure, upon the relationship that normally exists between local cerebral blood flow (LCBF) and local cerebral glucose utilization (LCMRglu). Dark agouti (DA) rats were pre-treated with 15 mg/kg i.p. MDMA or saline. Three weeks later, rats from each pre-treatment group were treated with an acute dose of MDMA (15 mg/kg i.p.) or saline. Quantitative autoradiographic imaging was used to measure LCBF or LCMRglu with [(14)C]-iodoantipyrine and [(14)C]-2-deoxyglucose, respectively. Serotonergic terminal depletion was assessed using radioligand binding with [(3)H]-paroxetine and immunohistochemistry. Three weeks after MDMA pre-treatment there were significant reductions in densities of 5-HT transporter (SERT)-positive fibres (-46%) and [(3)H]-paroxetine binding (-47%). In animals pre-treated with MDMA there were widespread significant decreases in LCMRglu, but no change in LCBF indicating a persistent loss of cerebrovascular constrictor tone. In both pre-treatment groups, acute MDMA produced significant increases in LCMRglu, while LCBF was significantly decreased. In 50% of MDMA-pre-treated rats, random areas of focal hyperaemia indicated a loss of autoregulatory capacity in response to MDMA-induced hypertension. These results suggest that cerebrovascular regulatory dysfunction resulting from acute exposure to MDMA is not diminished by previous exposure, despite a significant depletion in 5-HT terminals. However, there may be a sub-population, or individual circumstances, in which this dysfunction develops into a condition that might predispose to stroke.     

3H]paroxetine binding and serotonin content of rat and rabbit cortical areas, hippocampus, neostriatum, ventral mesencephalic tegmentum, and midbrain raphe nuclei region.

The high-affinity binding of [3H]paroxetine to membranes was measured in different regions of the rat and rabbit brain: cingulate, frontal, parietal, piriform, entorhinal, and visual cortical areas; dorsal and ventral hippocampus; rostral and caudal halves of neostriatum (rat) or caudate nucleus and putamen (rabbit); ventral mesencephalic tegmentum; and midbrain raphe nuclei region. The tissue concentrations of serotonin (5-HT), 5-hydroxyindole-3-acetic acid (5-HIAA) and 5-hydroxy-l-tryptophan (5-HTP) were also determined by high-performance liquid chromatography (HPLC) in the same brain samples. The regional density of [3H]paroxetine binding varied in both species; the highest values (Bmax) were found in the midbrain raphe region and ventral mesencephalic tegmentum. The cortical values ranged from moderate to low, with a significantly higher density in the cingulate cortex of the rat compared with rabbit. In the rat, there was also a higher density in the ventral than dorsal hippocampus, and the caudal than rostral neostriatum. In the rabbit, the hippocampal and neostriatal values were generally lower and more uniform. In both species, there was an excellent correlation between regional 5-HT levels and specific [3H]paroxetine binding (r = 0.87 in the rat and 0.96 in the rabbit). Considering the available quantitative data on the number of 5-HT nerve cell bodies and axon terminals in different regions of the rat brain, it appears likely that the high amount of [3H]paroxetine binding in the midbrain raphe region and ventral mesencephalic tegmentum reflects the presence of 5-HT uptake sites on 5-HT nerve cell bodies and dendrites as well as axon terminals. In other brain regions, the heterogeneous distribution of [3H]paroxetine binding parallels that of the number of 5-HT axon terminals, emphasizing the potential usefulness of this radioligand as a marker of 5-HT innervation density.     

Platelet serotonin uptake and paroxetine binding among allelic genotypes of the serotonin transporter in alcoholics

Expression rates of long (L) and short (S) alleles of the serotonin (5-HT) transporter (5-HTT) gene have been shown to differ under various circumstances. We compared 5-HTT uptake (function) level and paroxetine binding (density) in platelets of alcoholics as indices of 5-HTT expression rate among LL, LS, and SS genotypes. Concentration curves of [3H]5-HT and [3H]paroxetine were used to quantify the equilibrium constant (Km) and maximum 5-HT uptake rate (Vmax) for 5-HTT uptake into intact platelets and the dissociation constant (Kd) and maximum specific binding density (Bmax) for paroxetine binding to platelet membranes, respectively. Genotypes were determined using electrophoresis with fluorescent markers. Vmax for 5-HTT uptake did not correlate with Bmax for paroxetine binding (r=-0.095, P=0.415). Means of Vmax and Bmax did not differ in a statistically significant manner among LL, LS, and SS genotypes in these alcoholic subjects. However, Vmax for LL and SS appeared to have a bimodal distribution, so the percentage of subjects with Vmax <200 fmol/min-10(7) platelets was statistically significantly higher in LL than in SS (51.5% vs. 22.7%, respectively), with an odds ratio of 3.6 (P<0.05). The percentage of Vmax <200 fmol/min-10(7) platelets for LS was 39.3% (not significant vs. LL or SS). Previous studies of healthy human controls have shown that 5-HTT density in raphe nuclei and 5-HTT uptake in platelets are higher in the LL genotype than in S carriers. Our findings in currently drinking alcoholics support the hypothesis that those with the LL genotype of the 5'-HTTLPR region of the 5-HTT gene have reduced 5-HTT function.     

Effects of serotonergic denervation on the density and plasticity of brain muscarinic receptors in the rat.

This study investigated whether serotonergic lesion may affect density, sensitivity, and plasticity of muscarinic receptors in hippocampus and cerebral cortex. Intracerebroventricular injection of 5,7-dihydroxytryptamine (5,7-DHT) in rats produced a 90% reduction in cortical and hippocampal 5-hydroxytryptamine (5-HT) and 5-hydroxyindoleacetic acid (5-HIAA) contents. In these brain areas, the 5,7-DHT lesion did not affect the overall density of muscarinic receptors or those of M1 and non-M1 muscarinic receptor subtypes as assayed using [3H]N-methylscopolamine ([3H]NMS), [3H]pirenzepine, and [3H]NMS in the presence of pirenzepine, respectively. In addition, the binding of the muscarinic agonist [3H]oxotremorine-M (OXO-M), taken as an indirect index of coupling efficiency of non-M1 receptors with G-proteins, did not change significantly in cortex and hippocampus of 5,7-DHT-lesioned rats. Similarly, carbachol-induced accumulation of [3H]inositol phosphates (InPs) in hippocampal miniprisms showed no significant differences between tissues from 5,7-DHT-lesioned and sham-operated rats. In sham-operated rats, an intraperitoneal (i.p.) injection of scopolamine (10 mg/kg once daily) during 21 days caused an increased density of [3H]NMS binding sites in cortex (+20%) and hippocampus (+26%). This up-regulation was restricted to non-M1 receptors subtypes. In 5,7-DHT-lesioned rats, chronic scopolamine failed to modify significantly the density of cortical or hippocampal M1 or non-M1 receptors. These results suggest 1) that 5-HT denervation did not affect the density and sensitivity of muscarinic receptors and 2) that the ability of cortical and hippocampal non-M1 receptors to up-regulate following repeated injection of scopolamine requires the integrity of 5-HT neurons terminating in these brain structures.     

Effects of antihistamines on 3, 4-methylenedioxymethamphetamine-induced depletion of serotonin in rats.

This study investigated the effects of chlorpheniramine (CPA, 10-25 mg/kg), diphenhydramine (DIPH, 20 mg/kg), tripelennamine (TRIP, 20 mg/kg), and pyrilamine (PYRI, 20 mg/kg) on 3, 4-methylenedioxymethamphetamine (MDMA, 20 mg/kg x 2)-induced hyperthermia and depletion of indoles in rat brains, on the uptake of serotonin and dopamine into rat synaptosomes, on the binding affinity of CPA for biogenic amine transporters in the synaptosomes of rat brain, and on the scavenging hydroxyl free radicals activity. Rats were treated with two injections of MDMA, CPA, DIPH, TRIP, PYRI, and saline, alone or in combination of MDMA with one of the antihistamines, 6 h apart and sacrificed 5 days later. Rectal temperature was measured prior to and hourly following the drug injections for 13 h. As compared to saline controls, MDMA increased body temperature and decreased levels of indoles, measured by HPLC, in several brain regions of rats. CPA attenuated and DIPH had no effect on MDMA-induced hyperthermia, yet both attenuated the depletion of indoles, whereas PYRI and TRIP potentiated these effects. CPA inhibited the binding of [(3)H]paroxetine and [(3)H]nisoxetine to the synaptosomes of cerebral cortex and of [(3)H]win 35,428 to the synaptosomes of striatum. CPA, DIPH, TRIP, and PYRI inhibited [(3)H]serotonin uptake. CPA, PYRI, and TRIP, but not DIPH, scavenge hydroxyl radicals. Possible mechanisms of the different effects of the antihistamines on MDMA-induced hyperthermia and depletion of serotonin are discussed. Published 1999 Wiley-Liss, Inc.     

Memantine prevents MDMA-induced neurotoxicity

MDMA (ecstasy) is an illicit drug causing long-term neurotoxicity. Previous studies demonstrated the interaction of MDMA with alpha-7 nicotinic acetylcholine receptor (nAChR) in mouse brain membranes and the involvement of alpha-7 nicotinic acetylcholine receptors (nAChR) in dopaminergic neurotoxicity induced by MDMA in mice. The aim of the present study was to investigate the utility of memantine (MEM), an alpha-7 nAChR antagonist used for treatment of Alzheimer's disease patients, to prevent neurotoxicity induced by MDMA in rats and the oxidative effect of this amphetamine derivative in mice striatal synaptosomes. In isolated mouse striatal synaptosomes (an in vitro model of MDMA neurotoxicity of dopaminergic origin), MDMA (50 microM)-induced reactive oxygen species (ROS) production that was fully inhibited by MEM (0.3 microM). This effect of MEM was fully prevented by PNU 282987 (0.5 microM), a specific agonist of alpha-7 nAChR. The preventive effect of MEM on this oxidative effect can be attributed to a direct antagonism between MDMA (acting probably as agonist) and MEM (acting as antagonist) at the alpha-7 nAChR. In Dark Agouti rats (an in vivo model of MDMA neurotoxicity of serotonergic origin), a single dose of MDMA (18 mg/kg) induced persistent hyperthermia, which was not affected by MEM pre-treatment. [(3)H]Paroxetine binding (a marker of serotonergic injury) was measured in the hippocampus of animals killed at 24h and 7 days after treatment. MDMA induced a significant reduction in [(3)H]paroxetine binding sites at both times of sacrifice that was fully prevented by pre-treatment with MEM. Since previous studies demonstrate that increased glutamate activity is not involved in the neurotoxic action of MDMA, it can be concluded that the effectiveness of MEM against MDMA-induced neurotoxicity would be the result of blockade of alpha-7 nAChR, although an indirect mechanism based on the interplay among the various neurotransmission systems leading to an increase in basal acetylcholine release should also be taken into account.     

Methylenendioxyamphetamine produces serotonin nerve terminal loss and diminished behavioural and neurochemical responses to the antidepressant fluoxetine

The effect of prior exposure to methylenedioxyamphetamine (MDA) on behavioural and neurochemical responses to fluoxetine were assessed in a rat model of antidepressant action. MDA (7.5 mg/kg, i.p.) was administered to rats twice daily for 4 consecutive days, and 4 weeks later the behavioural effect of fluoxetine (5 or 20 mg/kg; i.p. x 3) was examined in the modified rat forced-swimming test. In addition, the ability of fluoxetine to reduce serotonin (5-HT) metabolism was measured as an index of its efficacy in inhibiting 5-HT reuptake in vivo. In vehicle-treated rats, fluoxetine (5 and 20 mg/kg) produced a characteristic increase in swimming behaviour in the forced-swimming test. In contrast, fluoxetine-induced swimming was markedly attenuated in MDA-treated rats. MDA pretreatment resulted in 5-HT nerve terminal degeneration, indicated by reduced 5-HT and 5-HIAA concentrations in the frontal cortex, amygdala and hippocampus, and reduced [3H]paroxetine binding in the frontal cortex. In vehicle-treated rats, fluoxetine (5 and 20 mg/kg) decreased 5-HT metabolism (5-HIAA : 5-HT ratio) in the frontal cortex, amygdala and hippocampus. MDA pretreatment attenuated the ability of fluoxetine to reduce 5-HT metabolism in all brain regions examined. These findings are the first to demonstrate that prior exposure to the methylenedioxy-substituted amphetamine MDA results in diminished responsiveness to the antidepressant fluoxetine.     

Decreased serotonin transporter binding in unaffected relatives of manic depressive patients.

BACKGROUND: There are numerous reports of decreased binding to platelet serotonin transporter (5-HTT) in depression, suggesting that it might be considered a trait marker of depression. To further investigate whether reduced 5-HTT function could be an endophenotype in manic depressive illness, we looked for abnormalities of platelet 5-HTT among subjects who are potential carriers of genetic vulnerability to manic depressive illness (MDI). METHODS: Blood samples were obtained from 20 unaffected relatives from families with at least two individuals with bipolar disorder and from 19 control participants. Plasma 5-HIAA, platelet 5-HT, and [3H] imipramine binding were measured. RESULTS: Unaffected relatives manifested lower platelet 5-HTT function than control participants as revealed both by reduced number and diminished affinity of imipramine binding sites and diminished platelet 5-HT content. CONCLUSIONS: These preliminary results suggest that reduced 5-HTT function could be considered a trait marker or an endophenotype in MDI.     

Muscarinic receptor/G-protein coupling is reduced in the dorsomedial striatum of cognitively impaired aged rats

Behavioral flexibility, the ability to modify responses due to changing task demands, is detrimentally affected by aging with a shift towards increased cognitive rigidity. The neurobiological basis of this cognitive deficit is not clear although striatal cholinergic neurotransmission has been implicated. To investigate the possible association between striatal acetylcholine signaling with age-related changes in behavioral flexibility, young, middle-aged, and aged F344 X Brown Norway F1 rats were assessed using an attentional set-shifting task that includes two tests of behavioral flexibility: reversal learning and an extra-dimensional shift. Rats were also assessed in the Morris water maze to compare potential fronto-striatal-dependent deficits with hippocampal-dependent deficits. Behaviorally characterized rats were then assessed for acetylcholine muscarinic signaling within the striatum using oxotremorine-M-stimulated [(35)S]GTPγS binding and [(3)H]AFDX-384 receptor binding autoradiography. The results showed that by old age, cognitive deficits were pronounced across cognitive domains, suggesting deterioration of both hippocampal and fronto-striatal regions. A significant decline in oxotremorine-M-stimulated [(35)S]GTPγS binding was limited to the dorsomedial striatum of aged rats when compared to young and middle-aged rats. There was no effect of age on striatal [(3)H]AFDX-384 receptor binding. These results suggest that a decrease in M2/M4 muscarinic receptor coupling is involved in the age-associated decline in behavioral flexibility.     

Cortical serotonin transporter density and verbal memory in individuals who stopped using 3,4-methylenedioxymethamphetamine (MDMA or "ecstasy"): preliminary findings

BACKGROUND: Although the popular drug 3,4-methylenedioxymethamphetamine (MDMA or "ecstasy") has been shown to damage brain serotonin (5-HT) neurons in animals, the fate and functional consequences of 5-HT neurons after MDMA injury are not known in humans. We investigated the long-term effects of MDMA use on cortical 5-HT neurons in humans and memory function, because brain 5-HT has been implicated in memory function. METHODS: Twenty-two recent MDMA users, 16 ex-MDMA users who had stopped using MDMA for more than 1 year, and 13 control subjects. The effects of MDMA use on cortical 5-HT neurons was studied by means of single-photon emission computed tomography with iodine 123-labeled 2beta-carbomethoxy-3beta-(4-iodophenyl) tropane ([(123)I]beta-CIT) by quantification of brain 5-HT transporter densities. Verbal memory performance was assessed with the Rey Auditory Verbal Learning Test. RESULTS: Mean cortical [(123)I]beta-CIT-labeled 5-HT transporter density was significantly lower in recent MDMA users than in controls (1.17 vs. 1.28 [-9%]) but not in ex-MDMA users (1.24 vs. 1.28 [-3%]). Recent and ex-MDMA users recalled significantly fewer words than did controls on the immediate recall (47.0 and 48.0 vs 60.0, respectively; P =.001) as well as the delayed recall (9.8 and 10.1 vs. 13.1, respectively; P =.003). Greater use of MDMA was associated with greater impairment in immediate verbal memory. However, memory performance was not associated with [(123)I]beta-CIT binding to cortical 5-HT transporters or duration of abstinence from MDMA. CONCLUSION: The present study suggests that, while the neurotoxic effects of MDMA on 5-HT neurons in the human cortex may be reversible, the effects of MDMA on memory function may be long-lasting.     

In vivo effects of the putative cognitive enhancer KA-672.HCl in comparison with 8-hydroxy-2-(di-N-propylamino) tetralin and haloperidol on dopamine, 3,4-dihydroxyphenylacetic acid, serotonin and 5-hydroxyindoleacetic acid levels in striatal and cortical brain regions

1. KA-672.HCl (7-methoxy-6-[3-[4-(2-methoxyphenyl)piperazin-1-yl]propoxy]-3,4-di methyl-2H-1-benzopyran-2-one hydrochloride), designed as a cognitive enhancer, has been investigated through behavioural and binding studies. However, little is known about its biochemical effects on the dopaminergic and serotoninergic system in vivo. 2. In the present study the authors investigated the effects of KA-672.HCl (0.1 mg/kg and 1 mg/kg), 8-hydroxy-2-(di-N-propylamino)tetralin (8-OH-DPAT) (1 mg/kg), haloperidol (0.1 mg/kg) and a mixture of haloperidol and 8-OH-DPAT on dopamine (DA), 3,4-dihydroxyphenylacetic acid (DOPAC), serotonin (5-HT) and 5-hydroxyindolacetic acid (5-HIAA) levels, in striatum and cerebral cortex of rats. 3. Male Wistar rats received an intraperitoneal injection of the drugs or vehicle 1 hour before striatal and cortical brain tissues were dissected out for neurochemical analysis. 4. KA-672.HCl, 8-OH-DPAT and haloperidol significantly reduced striatal DA levels, whereas only KA-672.HCl significantly reduced cortical DA levels. 8-OH-DPAT and haloperidol induced a significant increase in cortical DOPAC levels but only haloperidol significantly elevated the striatal DOPAC content. In contrast, only the higher dose of KA-672.HCl elevated striatal DOPAC levels. Furthermore, KA-672.HCl significantly reduced striatal 5-HT levels and slightly elevated striatal 5-HIAA concentrations. 8-OH-DPAT significantly decreased striatal 5-HIAA levels. All substances were able to enhance the cortical and striatal DA turnover. 5. The cortical and striatal 5-HT turnover was significantly decreased following 8-OH-DPAT treatment and significantly increased in the striatum after haloperidol and KA-672.HCl treatment. 6. The data suggest that KA-672.HCl possesses D2 antagonistic as well as 5-HT1A agonistic properties. However, additional mechanisms of actions by interaction with other neurotransmitter systems such as acetylcholine, excitatory or inhibitory amino acids need to be determined.     

Serotonin transporter occupancy of five selective serotonin reuptake inhibitors at different doses: an [11C]DASB positron emission tomography study

OBJECTIVE: Minimum therapeutic doses of paroxetine and citalopram produce 80% occupancy for the serotonin (5-HT) transporter (5-HTT). The authors used [(11)C]DASB positron emission tomography to measure occupancies of three other selective serotonin reuptake inhibitors (SSRIs) at minimum therapeutic doses. The relationship between dose and occupancy was also investigated. METHOD: Striatal 5-HTT binding potential was measured in 77 subjects before and after 4 weeks of medication administration. Binding potential is proportional to the density of receptors not blocked by medication. Subjects received citalopram, fluoxetine, sertraline, paroxetine, or extended-release venlafaxine. Healthy subjects received subtherapeutic doses; subjects with mood and anxiety disorders received therapeutic doses. Percent reduction in 5-HTT binding potential for each medication and dose was calculated. To obtain test-retest data, binding potential was measured before and after 4 weeks in six additional healthy subjects. RESULTS: Substantial occupancy occurred at subtherapeutic doses for all SSRIs. Compared to test-retest data, each drug at the minimum therapeutic dose had a significant effect on striatal 5-HTT binding potential. Mean occupancy at this dose was 76%-85%. At higher plasma SSRI concentrations, 5-HTT occupancy tended to increase above 80%. For each drug, as the dose (or plasma level) increased, occupancy increased nonlinearly, with a plateau for higher doses. CONCLUSIONS: At tolerable doses, SSRIs have increasing occupancy with increasing plasma concentration or dose. Occupancy of 80% across five SSRIs occurs at minimum therapeutic doses. This suggests that 80% 5-HTT blockade is important for therapeutic effect. Occupancy should be measured during development of antidepressant compounds targeting the 5-HTT.