Desipramine Research Articles

Noradrenergic facilitation of shock-probe defensive burying in lateral septum of rats, and modulation by chronic treatment with desipramine

We have previously shown that acute stress-induced release of norepinephrine (NE) facilitates anxiety-like behavioral responses to stress, such as reduction in open-arm exploration on the elevated-plus maze and in social behavior on the social interaction test. Since these responses represent inhibition of ongoing behavior, it is important to also address whether NE facilitates a response that represents an activation of behavior. Correspondingly, it is unknown how a chronic elevation in tonic steady-state noradrenergic (NA) neurotransmission induced by NE reuptake blockade might alter this acute modulatory function, a regulatory process that may be pertinent to the anxiolytic effects of NE reuptake blockers such as desipramine (DMI). Therefore, in this study, we investigated noradrenergic modulation of the shock-probe defensive burying response in the lateral septum (LS). In experiment 1, shock-probe exposure induced an acute 3-fold increase in NE levels measured in LS of male Sprague-Dawley rats by microdialysis. Shock-probe exposure also induced a modest rise in plasma ACTH, taken as an indicator of perceived stress, that returned to baseline more rapidly in rats that were allowed to bury the probe compared to rats prevented from burying by providing them with minimal bedding, indicating that the active defensive burying behavior is an effective coping strategy that reduces the impact of acute shock probe-induced stress. In experiment 2, blockade of either α 1- or β-adrenergic receptors in LS by local antagonist microinjection immediately before testing reduced defensive burying and increased immobility. In the next experiment, chronic DMI treatment increased basal extracellular NE levels in LS, and attenuated the acute shock probe-induced increase in NE release in LS relative to baseline. Chronic DMI treatment decreased shock-probe defensive burying behavior in a time-dependent manner, apparent only after 2 weeks or more of drug treatment. Moreover, rats treated chronically with DMI showed no significant rise of plasma ACTH in response to shock-probe exposure. Thus, acute stress-induced release of NE in LS facilitated defensive burying, an active, adaptive behavioral coping response. Chronic treatment with the NE reuptake blocker and antidepressant drug DMI attenuated acute noradrenergic facilitation of the active burying response, and also attenuated the level of perceived stress driving that response. These results suggest that long-term regulation of the acute modulatory function of NE by chronic treatment with reuptake blockers may contribute to the mechanisms by which such drugs exert their anxiolytic effects in the treatment of stress-related psychiatric conditions, including depression and anxiety.

Vitamin E Reduces Antidepressant-Related β-Adrenoceptor Down-Regulation in Cultured Cells. Comparable Effects on St. John’s Wort and Tricyclic Antidepressant Treatment

The mode of action of antidepressants is still a matter of debate. Acute inhibition of neurotransmitter reuptake in central neuronal synapses, followed by a down-regulation of central postsynaptic beta-adrenoceptor (beta-AR) numbers were consistently observed in vivo, while a reduction in surface beta-AR density was found in cell cultures. Effects of the tricyclic antidepressant desipramine (DMI) were abolished by vitamin E (alpha-TOC) in vitro as well as in vivo. Alpha-TOC interfered with antidepressant-induced changes of cellular plasma membrane properties and with recycling of beta-AR. St. John's wort (SJW) extract reduced beta-AR numbers in cultured cells to a similar extent as DMI or the selective serotonin re-uptake inhibitor fluoxetine. We chronically co-exposed cell cultures to SJW extract and to alpha-TOC. Receptor down-regulation following exposure to the plant extract was inhibited in the presence of alpha-TOC suggesting a mode of action of SJW extract comparable to that of synthetic antidepressants. Inhibition of cell proliferation by the plant extract was also significantly reduced by alpha-TOC.

Chronic Treatment with Desipramine Improves Cognitive Performance of Rats in an Attentional Set-Shifting Test

Alterations in central monoaminergic neurotransmission are important in the actions of many antidepressants. This study tested the hypothesis that tonic elevation of noradrenergic (NA) neurotransmission in medial prefrontal cortex (mPFC) by chronic treatment with the selective norepinephrine (NE) reuptake blocker desipramine (DMI) may contribute to the beneficial cognitive effects of this antidepressant drug (AD). Male Sprague-Dawley rats were treated with DMI acutely (15 mg/kg, i.p.) or chronically for 21 days (7.5 mg/kg/day via osmotic minipump) before assessing performance on an attentional set-shifting test. The extradimensional set-shifting component of this test reflects a process of cognitive flexibility that is dependent upon mPFC, and that we have shown previously to be facilitated by NA activity in mPFC. Microdialysis was performed to measure NE release in mPFC concurrently with behavioral testing. Acute DMI treatment produced an increase in extracellular NE levels in mPFC, and a modest improvement in overall performance across all task stages of the attentional set-shifting test, but failed to produce a significant improvement in any of the individual specific tasks comprising the test sequence. Chronic DMI treatment tonically elevated basal extracellular NE levels in mPFC, associated with a significant improvement in performance specifically on the extradimensional set-shifting component of the test. There was also a significant reduction in set loss errors in rats treated chronically with DMI. Hence, tonic elevation of NA transmission in mPFC by chronic DMI treatment was associated with a time-dependent facilitation of cognitive flexibility that may contribute to the mechanism whereby chronic treatment with ADs, specifically NE reuptake blockers, may exert a beneficial therapeutic effect on cognition in depressed patients.

PET Imaging of Norepinephrine Transporters

The involvement of the norepinephrine transporter (NET) in the pathophysiology and treatment of attention deficit hyperactivity disorder (ADHD), substance abuse, neurodegenerative disorders (e.g., Alzheimer's disease (AD) and Parkinson's disease (PD)) and depression has long been recognized. However, many of these important findings have resulted from studies in vitro using postmortem tissues; as of now, these results have never been verified via in vivo methods because brain imaging of NET in living systems has been hampered due to the lack of suitable radioligands. The fact that all three monoamine (dopamine, norepinephrine, and serotonin) transporters (DAT, NET and SERT) are involved in various neurological and psychiatric diseases further emphasizes the need to develop suitable NET ligands so that researchers will be able to probe the contributions of each monoamine transporter system to specific CNS disorders. In this review article, the design and biological evaluation of several radioligands for imaging the brain NET system with PET are discussed. Based on these characterization studies, including C-11 labeled desipramine (DMI), 2-hydroxydesipramine (HDMI), talopram, talsupram, nisoxetine (Nis), oxaprotiline (Oxap), lortalamine (Lort) and C-11 and F-18 derivatives of reboxetine (RB), methylreboxetine (MRB) and their individual (R, R) and (S, S) enantiomers, in conjunction with studies with radiolabeled 4-iodo-tomoxetine and 2-iodo-nisoxetine, we have identified the superiority of (S, S)-[(11)C]MRB and the suitability of the MRB analogs as potential NET ligands for PET. In contrast, Nis, Oxap and Lort displayed high uptake in striatum (higher than thalamus). The use of these ligands is further limited by high non-specific binding and relatively low specific signal, as is characteristic of many earlier NET ligands. Thus, to our knowledge, (S, S)-[(11)C]MRB remains by far the most promising NET ligand for PET studies.

Desipramine prevents the sustained increase in corticotropin‐releasing hormone‐like immunoreactivity induced by repeated immobilization stress in the rat central extended amygdala

Clinical and experimental studies have shown that the activation of corticotropin-releasing hormone (CRH) and noradrenergic systems mediate stress-induced anxiety. Repeated immobilization stress (RIS) has been shown to induce long-lasting anxiety behavior and changes in noradrenaline turnover. The present work was aimed at studying the effect of RIS on the in situ expression of CRH-LI in the central extended amygdala and paraventricular nucleus of the hypothalamus (PVN). Our results showed that RIS for 15 days induces a significant increase of CRH-LI expression in the central extended amygdala. The increase in CRH-LI expression in the central extended amygdala was sustained even after a 25-day stress-free period. The concomitant administration of desipramine (DMI), a specific noradrenaline uptake inhibitor, fully prevented the RIS-induced increase in CRH expression. RIS also induced an increase of CRH-LI expression in the PVN that was prevented by the concomitant DMI administration. In contrast to the sustained effect observed in the central extended amygdala, the RIS-induced increase of CRH-LI expression in the PVN was nonlasting. DMI administration also prevented the RIS-induced increase of adrenal gland weight. The present findings showing that RIS induces a sustained increase of CRH expression in the central extended amygdala suggest that the repeated activation of CRH neurons and CRH receptors in the central extended amygdala may underlie the long-lasting anxiety behavior induced by RIS. Further studies should address the mechanisms involved in the effect of DMI and its eventual relevance in the therapeutic actions of DMI.

Nigrostriatal Damage with 6‐OHDA

The 6-hydroxydopamine (6-OHDA) model of Parkinson's disease in the rat represents a fundamental tool for investigating the pathophysiology of dopamine denervation. Nevertheless, 6-OHDA can induce also noradrenergic lesions; therefore desmethylimipramine (DMI) is co-administrated as a selective inhibitor of noradrenergic reuptake to protect noradrenaline (NA) fibers neighboring DA neurons and/or axons. The neurotoxin 6-OHDA must be microinfused selectively into the substantia nigra pars compacta (SNpc) or into the medial forebrain bundle (MFB) to determine the nigrostriatal lesion. However, this experimental procedure is invasive and always produces a certain amount of mechanical damage that cannot be prevented by pharmacological approaches. For this reason, we have compared two types of experimental design in which we tested critical steps of the procedures, such as the flow rate. We microinfused rats in MFB with 8 microL of total volume of a solution containing the neurotoxin (infusion rate 2 microL/min in 4 min) according with general practice, and rats microinfused with an amount of 2 microL of total volume with a slower rate (0.2 microL/min in 10 min) of infusion. Rats infused with a higher flow rate of infusion underwent striatal NA loss in spite of the administration of DMI. On the contrary, rats infused with a slow infusion flow rate had spared NA axons following DMI. These results suggest that the flow rate and the volume of 6-OHDA infusion are critical to prevent the occurrence of nonspecific mechanical effects.

Blockade of the α‐MMT‐induced depletion of rat cortical norepinephrine levels: a method for determining the potency and duration of action of compounds with norepinephrine reuptake inhibition

Neurotransmitter transporter inhibitors can prevent the depletion of brain monoamines when the depleting agent requires active uptake into neurons via a transporter. α-Methyl-m-tyrosine (α-MMT), a monoaminergic depleting agent, is β-hydroxylated to metaraminol in vivo and actively transported into noradrenergic neurons, leading to decreased levels of norepinephrine (NE). This depletion of NE by α-MMT is prevented by inhibitors of the neuronal NE transporter, but is dependent upon the dose of α-MMT administered. A 6.25 mg/kg, sc dose of α-MMT produced a 40% to 60% depletion of rat cortical NE 2 to 4 hours after administration that persisted up to 16 hours. Pretreatment with desipramine (DMI) at 10 mg/kg, sc significantly blocked NE depletion produced by a-MMT up to 8 hours regardless of whether measured 2 or 16 hours after a-MMT administration. The α-MMT-induced decrease in brain NE at 2 hours was dose-dependently blocked by a 1 hour pretreatment with DMI and reboxetine (REB) with ED50 doses of <1 and 2.0 mg/kg, sc, respectively. When administered at various times prior to α-MMT, DMI and REB pretreatment significantly blocked the α-MMT-induced depletion of rat cortical NE levels up to 8 hours with no significant effect at 24 hours. This experimental protocol allows for a rapid comparison of the potency and duration of NE reuptake inhibition in rat cortex, which has implications for development of agents effective in the treatment of central nervous system disorders.

Antidepressants and Antiepileptic Drugs for Chronic Non-Cancer Pain

The development of newer classes of antidepressants and second-generation antiepileptic drugs has created unprecedented opportunities for the treatment of chronic pain. These drugs modulate pain transmission by interacting with specific neurotransmitters and ion channels. The actions of antidepressants and antiepileptic drugs differ in neuropathic and non-neuropathic pain, and agents within each medication class have varying degrees of efficacy. Tricyclic antidepressants (e.g., amitriptyline, nortriptyline, desipramine) and certain novel antidepressants (i.e., bupropion, venlafaxine, duloxetine) are effective in the treatment of neuropathic pain. The analgesic effect of these drugs is independent of their antidepressant effect and appears strongest in agents with mixed-receptor or predominantly noradrenergic activity, rather than serotoninergic activity. First-generation antiepileptic drugs (i.e., carbamazepine, phenytoin) and second-generation antiepileptic drugs (e.g., gabapentin, pregabalin) are effective in the treatment of neuropathic pain. The efficacy of antidepressants and antiepileptic drugs in the treatment of neuropathic pain is comparable; tolerability also is comparable, but safety and side effect profiles differ. Tricyclic antidepressants are the most cost-effective agents, but second-generation antiepileptic drugs are associated with fewer safety concerns in elderly patients. Tricyclic antidepressants have documented (although limited) efficacy in the treatment of fibromyalgia and chronic low back pain. Recent evidence suggests that duloxetine and pregabalin have modest efficacy in patients with fibromyalgia. Copyright © 2005 American Academy of Family Physicians.) Permission from the AAFP was granted to publish this article in the hardcopy only. Kindly refer to www.aafp.org or subscribe to the hardcopy of South African Family Practice. (SA Fam Pract 2005;47(3): 30-36)

Synthesis and Positron Emission Tomography Evaluation of Three Norepinephrine Transporter Radioligands: [C-11]Desipramine, [C-11]Talopram and [C-11]Talsupram

Desipramine (DMI), talopram and talsupram, three of the most potent norepinephrine transporter (NET) inhibitors reported to date, were radiolabeled in high yields and at high specific radioactivity (58-75 GBq/micromol) by the methylation of nor-precursors with [C-11]methyl triflate. The regional brain distribution of each radioligand following intravenous injection into cynomolgus monkey was examined in vivo with positron emission tomography (PET). For all three radioligands, the regional brain distribution of radioactivity was slightly heterogeneous, with higher uptake of radioactivity in the mesencephalon, thalamus and lower brainstem than in striatum. The rank order of maximal brain radioactivity (as percentage of injected dose) was [C-11]DMI (2.7%) > [C-11]talsupram (1.3%) > [C-11]talopram (0.7%). The appearance of radioactive metabolites in plasma was similar for each radioligand (75-85% of radioactivity in plasma at 45 min). These metabolites were all more polar than their parent radioligand. The data show that these radioligands are inferior to existing radioligands for the study of brain NET with PET in vivo.

Desipramine attenuates working memory impairments induced by partial loss of catecholamines in the rat medial prefrontal cortex

The density of tyrosine hydroxylase-immunoreactive (TH-IR) axons in the prefrontal cortex of schizophrenic subjects may be reduced by as much as 50% in the deep cortical layers (Am J Psychiatry 156:1580-1589, 1999). Previously, we demonstrated that approximately 60% loss of TH-IR axons in the rat medial prefrontal cortex (mPFC) decreases local basal and stress-evoked extracellular dopamine (DA) concentrations, suggesting that moderate loss of DA axons in the mPFC is sufficient to alter the neurochemical activity of the remaining DA neurons (Neuroscience 93:497-505, 1999). To further assess the functional consequences of partial mPFC DA depletion, we examined the effects of 6-hydroxydopamine lesions of the rat mPFC on behavior in a T-maze delayed-response task. We also assessed whether chronic administration of the norepinephrine (NE) uptake inhibitor, desipramine (DMI), attenuates lesion-induced deficits in T-maze performance. Previous research indicates that inhibition of NE transport in the mPFC results in a concomitant increase in extracellular DA and NE. Moderate loss of mPFC DA and NE (approximately 50 and 10% loss, respectively) was sufficient to impair delayed-response behavior, in part due to an increase in perseverative responding. Chronic DMI treatment (3 mg/kg delivered via osmotic pumps) impaired performance of control rats but attenuated the deficits in delayed-response behavior in rats previously sustaining loss of mPFC DA and NE (approximately 75 and 35% loss, respectively). These data suggest that moderate loss of DA and NE in the prefrontal cortex is sufficient to impair cognitive function, and these behavioral effects are attenuated by inhibition of the NE transporter.

Early response to desipramine among women with bulimia nervosa

Numerous trials have demonstrated the efficacy of antidepressant medications for the treatment of bulimia nervosa (BN). The current study examined whether early response to medication predicted response to medication at the end of a controlled trial. Data from two previously published studies of desipramine (DMI) were used. Seventy-seven patients with BN were included in the analysis. Receiver operating characteristic (ROC) curves were constructed to examine the relation between the percentage reduction in symptoms at each week and failure to respond to antidepressant medication at the end of the trial. Eventual nonresponders to DMI could be reliably identified in the first 2 weeks of treatment. The current study provides preliminary evidence that patients with BN who will not respond to antidepressant medication can be identified in the first 2 weeks of treatment.

Role of the increased noradrenergic neurotransmission in drug self-administration

Psychostimulants increase extracellular monoamine concentrations in the CNS. While the contributions of dopamine (DA) and serotonin (5-HT) to the reinforcing effect of psychostimulants have been examined, less is known about the involvement of norepinephrine (NE). In the present study, cocaine, desipramine (DMI) and JZ-III-84 were made available to rhesus monkeys ( n = 4) responding under a progressive-ratio (PR) schedule. These compounds vary in their in vitro selectivities for blocking NE uptake relative to DA from high (DMI) to modest (JZ-III-84) to non-selective (cocaine). Additionally, cocaine mixed with DMI in mg/kg dose-ratios of 1:1 to 1:3 was made available for self-administration. NE uptake inhibition by the mixture of cocaine and DMI at a ratio of 1:3 was evaluated in an ex vivo uptake assay. Cocaine (0.01–0.1 mg/(kg injection)) and JZ-III-84 (0.001–0.1 mg/(kg injection)) functioned as positive reinforcers with sigmoidal or biphasic dose–response functions, whereas DMI failed to do so. The addition of DMI to cocaine did not systemically alter self-administration of cocaine. In the ex vivo uptake assay, the addition of DMI to cocaine significantly increased the NE uptake inhibition compared to cocaine. These results support the conclusion that CNS NE is not involved in the reinforcing mechanism of psychostimulants.

Effects of zotepine and olanzapine on noradrenaline transporter in cultured bovine adrenal medullary cells

Previously, it was demonstrated that the inhibitory effects of atypical antipsychotic drugs such as clozapine and risperidone on noradrenaline transporter (NAT) might in part be associated with their clinical profile. The present study examined the effects of zotepine on NAT in the cells and compared them with those of olanzapine. Adrenal medullary cells were isolated by a method of collagenase digestion of slices of fresh bovine adrenal medulla and the cells were plated at a density of 4 x 10(6) cells. Cells were incubated with [3H]noradrenaline (NA) in the presence or absence of zotepine or olanzapine. The amount of radioactivity taken into the cells was counted by a liquid scintillation counter. Plasma membranes of bovine adrenal medulla were prepared, and the binding of [3H]desipramine (DMI) was determined by incubating the membrane suspension in binding buffer together with zotepine or olanzapine. Specific binding of [3H] DMI was defined as that binding which was inhibited by nisoxetine. Both zotepine (10-1000 ng/ml) and olanzapine (10-1000 ng/ml) decreased [3H]NA uptake in a concentration-dependent manner. The IC50 values of zotepine and olanzapine on [3H]NA uptake were 10 +/- 4 and 14 +/- 8 ng/ml, respectively. Eadie-Hofstee analysis of [3H]NA uptake showed that treatment with zotepine and olanzapine decreased the V(max) of uptake without changing the K(m). Both zotepine (10-1000 ng/ml) and olanzapine (30-1000 ng/ml) inhibited [3H]DMI binding in a concentration-dependent manner. The IC50 values of zotepine and olanzapine on [3H]DMI binding were 50 +/- 18, and 120 +/- 38 ng/ml, respectively. Scatchard plot analysis of [3H]DMI binding showed that zotepine and olanzapine decreased the B(max) of binding without altering the K(d). The inhibitory effects of zotepine and olanzapine might be responsible in part for their clinical profile.

Sequential changes in BDNF mRNA expression and synaptic levels of AMPA receptor subunits in rat hippocampus after chronic antidepressant treatment

Increased expression of brain-derived neurotrophic factor (BDNF) appears to be involved in the mechanism of action of antidepressant drugs. It has also been proposed that potentiation of the AMPA receptor (AMPAR) function may be useful in the treatment of depression. Here we looked for the time course of the effect of different doses of two antidepressants, desipramine (DMI) and paroxetine (PAR), which differentially affect monoamine reuptake, on BDNF mRNA expression in hippocampal subfields (CA1, CA3 and dentate gyrus) and levels of AMPAR subunits in total and membrane-enriched extracts from rat hippocampus. Acute antidepressant treatment changed neither BDNF mRNA expression nor AMPAR subunit levels. In chronic treatments, rats were treated daily with the antidepressants for 7–21 days. PAR produced a time- and dose-dependent increase of BDNF expression in the three hippocampal subfields examined. On the contrary, the effect of DMI on BDNF mRNA was neither dose- nor time-dependent. In rats receiving the same chronic antidepressant treatments, PAR produced a dose-dependent increase of GluR1 and GluR2/3 levels in the membrane fraction after a 3-week treatment, and not at earlier times. DMI increased the membrane levels of AMPAR subunits after a 3-week treatment with the lower dose tested. However, a higher dose, 15 mg/kg, did not produce any change in AMPAR subunits and reduced membrane levels of α-tubulin and PSD-95, possibly indicating a disorganization of membrane scaffolding proteins. The results suggest that paroxetine, but not desipramine, enhances synaptic plasticity in the hippocampus by increasing BDNF mRNA expression, which determines a later AMPAR subunit trafficking to synaptic membranes.

Tyrosine administration does not affect desipramine-induced dopamine levels as measured in vivo in prefrontal cortex

Using in vivo microdialysis, we examined whether tyrosine administration would potentiate the desipramine (DMI)-induced elevation of medial prefrontal cortex (MPFC) dopamine (DA) levels. DMI (10 or 20 mg/kg IP) increased MPFC DA levels but not DOPA accumulation. Tyrosine (12.5–100 μg/ml) administered by reverse microdialysis did not affect DMI-induced MPFC DA levels. The data support our hypothesis that DA synthesis must be significantly increased in order for administered tyrosine to increase extracellular DA levels.

The effects of desmethylimipramine on cyclic AMP-stimulated gene transcription in a model cell system

The present study utilised an in vitro cell model of the cAMP signalling pathway to investigate the actions of desipramine (DMI) and other psychoactive agents on cAMP-driven gene transcription. The model comprised CHOβ 2 SPAP cells; Chinese hamster ovary cells expressing human β 2 adrenoceptors and a secreted placental alkaline phosphatase (SPAP) reporter gene with multiple cAMP response elements (CREs) in its promoter region. SPAP assays showed DMI to inhibit isoprenaline or forskolin-enhanced gene transcription in a time and concentration-dependent manner (IC 50 = 16.6 ± 2.0 μM after 18 h). This effect of DMI was not dependent upon activity at the levels of the β 2 receptor, cAMP accumulation or phosphorylation of the transcription factor, cAMP response element binding protein (CREB). The inhibitory effects were maintained in the presence of DMI for at least 3 weeks and were mimicked by exposure to norfluoxetine (the major metabolite of fluoxetine; IC 50 = 7.2 ± 1.8 μM) and the neuroleptics, chlorpromazine and clozapine, all at a concentration of 10 μM. Amphetamine (10 μM, 18 h) enhanced SPAP gene transcription. Ca 2+ imaging experiments ruled out an inhibitory effect of DMI on Ca 2+ influx as concluded by previous studies. The results suggest a molecular target for DMI that lies downstream of CREB phosphorylation. Whether the inhibitory action of DMI is common to naturally expressed CRE-driven genes involved in adaptive responses to antidepressants in vivo remains to be determined.

Enhancement of antidepressant-like activity by joint administration of imipramine and magnesium in the forced swim test: Behavioral and pharmacokinetic studies in mice

The effect of joint administration of imipramine (IMI) and magnesium (Mg) on antidepressant-like activity was studied in mice using forced swim test (FST). Mg doses ineffective per se (5 and 10 mg/kg) given jointly with IMI also at ineffective doses (10 and 15 mg/kg) resulted in a potent reduction in the immobility time. Since these combined treatments did not influence locomotor activity, the antidepressant-like activity was not due to non-specific behavioral activation. Moreover, we estimated the effect of joint administration of magnesium and IMI in FST on serum and brain magnesium, IMI and its active metabolite desipramine (DMI) concentrations in mice. Swim stress (mice subjected to FST) increased the magnesium concentration in serum and decreased it in the brain compared to naive animals. Moreover administration of IMI increased (normalized) magnesium brain concentration, without influence on the serum level. Joint administration of IMI and magnesium did not influence magnesium (compared with FST) or IMI and DMI (compared with IMI treatment alone) concentrations in both examined tissues. The present data demonstrated an enhancement of the antidepressant-like effect by joint administration of IMI and magnesium in the FST, and further indicate the particular role of magnesium in the antidepressant action. Since there was no increase in IMI, DMI or magnesium concentration after joint administration of magnesium and IMI, the data suggest that pharmacodynamic rather than pharmacokinetic interaction between magnesium and IMI is accountable for behavioral effect in the FST.

Substrate Binding Stoichiometry and Kinetics of the Norepinephrine Transporter

The human norepinephrine (NE) transporter (hNET) attenuates neuronal signaling by rapid NE clearance from the synaptic cleft, and NET is a target for cocaine and amphetamines as well as therapeutics for depression, obsessive-compulsive disorder, and post-traumatic stress disorder. In spite of its central importance in the nervous system, little is known about how NET substrates, such as NE, 1-methyl-4-tetrahydropyridinium (MPP+), or amphetamine, interact with NET at the molecular level. Nor do we understand the mechanisms behind the transport rate. Previously we introduced a fluorescent substrate similar to MPP+, which allowed separate and simultaneous binding and transport measurement (Schwartz, J. W., Blakely, R. D., and DeFelice, L. J. (2003) J. Biol. Chem. 278, 9768-9777). Here we use this substrate, 4-(4-(dimethylamino)styrl)-N-methyl-pyridinium (ASP+), in combination with green fluorescent protein-tagged hNETs to measure substrate-transporter stoichiometry and substrate binding kinetics. Calibrated confocal microscopy and fluorescence correlation spectroscopy reveal that hNETs, which are homomultimers, bind one substrate molecule per transporter subunit. Substrate residence at the transporter, obtained from rapid on-off kinetics revealed in fluorescence correlation spectroscopy, is 526 micros. Substrate residence obtained by infinite dilution is 1000 times slower. This novel examination of substrate-transporter kinetics indicates that a single ASP+ molecule binds and unbinds thousands of times before being transported or ultimately dissociated from hNET. Calibrated fluorescent images combined with mass spectroscopy give a transport rate of 0.06 ASP+/hNET-protein/s, thus 36,000 on-off binding events (and 36 actual departures) occur for one transport event. Therefore binding has a low probability of resulting in transport. We interpret these data to mean that inefficient binding could contribute to slow transport rates.

Cognitive Therapy vs Medications in the Treatment of Moderate to Severe Depression

There is substantial evidence that antidepressant medications treat moderate to severe depression effectively, but there is less data on cognitive therapy's effects in this population. To compare the efficacy in moderate to severe depression of antidepressant medications with cognitive therapy in a placebo-controlled trial. Random assignment to one of the following: 16 weeks of medications (n = 120), 16 weeks of cognitive therapy (n = 60), or 8 weeks of pill placebo (n = 60). Research clinics at the University of Pennsylvania, Philadelphia, and Vanderbilt University, Nashville, Tenn. Two hundred forty outpatients, aged 18 to 70 years, with moderate to severe major depressive disorder. Some study subjects received paroxetine, up to 50 mg daily, augmented by lithium carbonate or desipramine hydrochloride if necessary; others received individual cognitive therapy. The Hamilton Depression Rating Scale provided continuous severity scores and allowed for designations of response and remission. At 8 weeks, response rates in medications (50%) and cognitive therapy (43%) groups were both superior to the placebo (25%) group. Analyses based on continuous scores at 8 weeks indicated an advantage for each of the active treatments over placebo, each with a medium effect size. The advantage was significant for medication relative to placebo, and at the level of a nonsignificant trend for cognitive therapy relative to placebo. At 16 weeks, response rates were 58% in each of the active conditions; remission rates were 46% for medication, 40% for cognitive therapy. Follow-up tests of a site x treatment interaction indicated a significant difference only at Vanderbilt University, where medications were superior to cognitive therapy. Site differences in patient characteristics and in the relative experience levels of the cognitive therapists each appear to have contributed to this interaction. Cognitive therapy can be as effective as medications for the initial treatment of moderate to severe major depression, but this degree of effectiveness may depend on a high level of therapist experience or expertise.

Clinical Response to Tricyclic Antidepressants in Functional Bowel Disorders is not Related to Dosage

As shown in the per protocol analysis of a recent randomized, controlled trial, when tolerated, Desipramine (DES) is effective over placebo (PLA) in treating moderate-to-severe functional bowel disorders (FBD). Clinical experience suggests that the benefit from tricyclic antidepressants (TCA) in FBD can be achieved at doses lower than those used to treat major depression. Within psychiatry, when using higher dosage of TCAs, plasma levels can be used to adjust daily dosage to optimize a treatment response. However, in FBD, it is not known whether plasma levels at the lower dosage are similarly related to a clinical response. To determine in treating FBD, whether DES blood levels or dose taken can predict a clinical response. As part of a study of 12 wk of antidepressant and psychological treatment in 431 patients with FBD at UNC and U of Toronto, we studied those participants who completed treatment (per protocol analysis) taking DES (N = 97, dose 50-150 mg/day) or pill placebo (PLA) (N = 55 1-3 pills/day). The primary outcome measure was defined as a composite score (Satisfaction with Treatment, McGill Pain Questionnaire, Global Well-being, and IBS-QOL). The composite score was correlated with: (i) DES plasma levels at week 6, and (ii) number of pills taken over the duration of the 12-wk treatment period. In addition, we also compared DES dose with DES plasma levels. There was a modest correlation between mean DES dose at weeks 5 and 6 and DES blood level at week 6 (R = 0.2 p < 0.07). However, there were no significant correlations between the composite score either with DES dose or with DES blood levels. Detectable blood levels of DES are associated with a clinical response in FBD. However, with dosages up to 150 mg, there is no relationship between total dose or plasma level and the clinical response.