ABSTRACT
Mass spectrometry imaging (MSI) is essential for visualizing drug distribution, metabolites, and significant biomolecules in pharmacokinetic studies. This study mainly focuses on imipramine, a tricyclic antidepressant that affects endogenous metabolite concentrations. The aim was to use atmospheric pressure matrix-assisted laser desorption/ionization (AP-MALDI)-MSI combined with different dimensionality reduction methods to examine the distribution and impact of imipramine on endogenous metabolites in the brains of treated wild-type mice. Brain sections from both control and imipramine-treated mice underwent AP-MALDI-MSI. Dimensionality reduction methods, including principal component analysis, multivariate curve resolution, and sparse autoencoder (SAE), were employed to extract valuable information from the MSI data. Only the SAE method identified phosphorylcholine (ChoP) as a potential marker distinguishing between the control and treated mice brains. Additionally, a significant decrease in ChoP accumulation was observed in the cerebellum, hypothalamus, thalamus, midbrain, caudate putamen, and striatum ventral regions of the treated mice brains. The application of dimensionality reduction methods, particularly the SAE method, to the AP-MALDI-MSI data is a novel approach for peak selection in AP-MALDI-MSI data analysis. This study revealed a significant decrease in ChoP in imipramine-treated mice brains.
Subject(s)
Brain , Imipramine , Phosphorylcholine , Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization , Animals , Imipramine/metabolism , Mice , Brain/metabolism , Brain/diagnostic imaging , Brain/drug effects , Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization/methods , Phosphorylcholine/metabolism , Phosphorylcholine/analogs & derivatives , Male , Antidepressive Agents, Tricyclic/pharmacokinetics , Antidepressive Agents, Tricyclic/pharmacology , Antidepressive Agents, Tricyclic/metabolism , Mice, Inbred C57BL , Principal Component AnalysisABSTRACT
Some tricyclic antidepressants (TCAs), including amitriptyline (ATL), clomipramine (CLO), and desipramine (DES), are known to be effective for management of neuropathic pain. It was previously determined that ATL, CLO, and DES are capable of voltage-dependent blocking of NMDA receptors of glutamate (NMDAR), which play a key role in pathogenesis of neuropathic pain. Despite the similar structure of ATL, CLO, and DES, efficacy of their interaction with NMDAR varies significantly. In the study presented here, we applied molecular modeling methods to investigate the mechanism of binding of ATL, CLO, and DES to NMDAR and to identify structural features of the drugs that determine their inhibitory activity against NMDAR. Molecular docking of the studied TCAs into the NMDAR channel was performed. Conformational behavior of the obtained complexes in the lipid bilayer was simulated by the method of molecular dynamics (MD). A single binding site (upper) for the tertiary amines ATL and CLO and two binding sites (upper and lower) for the secondary amine DES were identified inside the NMDAR channel. The upper and lower binding sites are located along the channel axis at different distances from the extracellular side of the plasma membrane. MD simulation revealed that the position of DES in the lower site is stabilized only in the presence of sodium cation inside the NMDAR channel. DES binds more strongly to NMDAR compared to ATL and CLO due to simultaneous interaction of two hydrogen atoms of its cationic group with the asparagine residues of the ion pore of the receptor. This feature may be responsible for the stronger side effects of DES. It has been hypothesized that ATL binds to NMDAR less efficiently compared to DES and CLO due to its lower conformational mobility. The identified features of the structure- and cation-dependent mechanism of interaction between TCAs and NMDAR will help in the further development of effective and safe analgesic therapy.
Subject(s)
Antidepressive Agents, Tricyclic , Molecular Docking Simulation , Molecular Dynamics Simulation , Receptors, N-Methyl-D-Aspartate , Receptors, N-Methyl-D-Aspartate/metabolism , Receptors, N-Methyl-D-Aspartate/chemistry , Antidepressive Agents, Tricyclic/pharmacology , Antidepressive Agents, Tricyclic/metabolism , Antidepressive Agents, Tricyclic/chemistry , Binding Sites , Amitriptyline/chemistry , Amitriptyline/metabolism , Amitriptyline/pharmacology , Humans , Clomipramine/pharmacology , Clomipramine/chemistry , Clomipramine/metabolism , Cations/metabolism , Cations/chemistry , Desipramine/pharmacology , Protein BindingABSTRACT
Fatty acid uptake is essential for cell physiological function, but detailed mechanisms remain unclear. Here, we generated an acetyl-CoA carboxylases (ACC1/2) double-knockout cell line, which lacked fatty acid biosynthesis and survived on serum fatty acids and was used to screen for fatty acid uptake inhibitors. We identified a Food and Drug Administration-approved tricyclic antidepressant, nortriptyline, that potently blocked fatty acid uptake both in vitro and in vivo. We also characterized underlying mechanisms whereby nortriptyline provoked lysosomes to release protons and induce cell acidification to suppress macropinocytosis, which accounted for fatty acid endocytosis. Furthermore, nortriptyline alone or in combination with ND-646, a selective ACC1/2 inhibitor, significantly repressed tumor growth, lipogenesis, and hepatic steatosis in mice. Therefore, we show that cells actively take up fatty acids through macropinocytosis, and we provide a potential strategy suppressing tumor growth, lipogenesis, and hepatic steatosis through controlling the cellular level of fatty acids.
Subject(s)
Fatty Liver , Metabolic Diseases , Neoplasms , Mice , Animals , Fatty Acids/metabolism , Antidepressive Agents, Tricyclic/pharmacology , Antidepressive Agents, Tricyclic/therapeutic use , Antidepressive Agents, Tricyclic/metabolism , Nortriptyline/metabolism , Nortriptyline/therapeutic use , Drug Repositioning , Fatty Liver/pathology , Metabolic Diseases/metabolism , Neoplasms/pathology , Liver/metabolismABSTRACT
Microbial diversity is reduced in the gut microbiota of animals and humans treated with selective serotonin reuptake inhibitors (SSRIs) and tricyclic antidepressants (TCAs). The mechanisms driving the changes in microbial composition, while largely unknown, is critical to understand considering that the gut microbiota plays important roles in drug metabolism and brain function. Using Escherichia coli, we show that the SSRI fluoxetine and the TCA amitriptyline exert strong selection pressure for enhanced efflux activity of the AcrAB-TolC pump, a member of the resistance-nodulation-cell division (RND) superfamily of transporters. Sequencing spontaneous fluoxetine- and amitriptyline-resistant mutants revealed mutations in marR and lon, negative regulators of AcrAB-TolC expression. In line with the broad specificity of AcrAB-TolC pumps these mutants conferred resistance to several classes of antibiotics. We show that the converse also occurs, as spontaneous chloramphenicol-resistant mutants displayed cross-resistance to SSRIs and TCAs. Chemical-genomic screens identified deletions in marR and lon, confirming the results observed for the spontaneous resistant mutants. In addition, deletions in 35 genes with no known role in drug resistance were identified that conferred cross-resistance to antibiotics and several displayed enhanced efflux activities. These results indicate that combinations of specific antidepressants and antibiotics may have important effects when both are used simultaneously or successively as they can impose selection for common mechanisms of resistance. Our work suggests that selection for enhanced efflux activities is an important factor to consider in understanding the microbial diversity changes associated with antidepressant treatments. IMPORTANCE Antidepressants are prescribed broadly for psychiatric conditions to alter neuronal levels of synaptic neurotransmitters such as serotonin and norepinephrine. Two categories of antidepressants are selective serotonin reuptake inhibitors (SSRIs) and tricyclic antidepressants (TCAs); both are among the most prescribed drugs in the United States. While it is well-established that antidepressants inhibit reuptake of neurotransmitters there is evidence that they also impact microbial diversity in the gastrointestinal tract. However, the mechanisms and therefore biological and clinical effects remain obscure. We demonstrate antidepressants may influence microbial diversity through strong selection for mutant bacteria with increased AcrAB-TolC activity, an efflux pump that removes antibiotics from cells. Furthermore, we identify a new group of genes that contribute to cross-resistance between antidepressants and antibiotics, several act by regulating efflux activity, underscoring overlapping mechanisms. Overall, this work provides new insights into bacterial responses to antidepressants important for understanding antidepressant treatment effects.
Subject(s)
Escherichia coli Proteins , Escherichia coli , Humans , Escherichia coli/genetics , Selective Serotonin Reuptake Inhibitors , Escherichia coli Proteins/metabolism , Fluoxetine/metabolism , Fluoxetine/pharmacology , Antidepressive Agents, Tricyclic/metabolism , Antidepressive Agents, Tricyclic/pharmacology , Amitriptyline/pharmacology , Antidepressive Agents/metabolism , Antidepressive Agents/pharmacology , Anti-Bacterial Agents/pharmacology , Drug Resistance, Multiple, Bacterial/genetics , Microbial Sensitivity TestsABSTRACT
Glioblastoma (GBM) is poorly responsive to therapy and invariably lethal. One conceivable strategy to circumvent this intractability is to co-target distinctive mechanistic components of the disease, aiming to concomitantly disrupt multiple capabilities required for tumor progression and therapeutic resistance. We assessed this concept by combining vascular endothelial growth factor (VEGF) pathway inhibitors that remodel the tumor vasculature with the tricyclic antidepressant imipramine, which enhances autophagy in GBM cancer cells and unexpectedly reprograms immunosuppressive tumor-associated macrophages via inhibition of histamine receptor signaling to become immunostimulatory. While neither drug is efficacious as monotherapy, the combination of imipramine with VEGF pathway inhibitors orchestrates the infiltration and activation of CD8 and CD4 T cells, producing significant therapeutic benefit in several GBM mouse models. Inclusion up front of immune-checkpoint blockade with anti-programmed death-ligand 1 (PD-L1) in eventually relapsing tumors markedly extends survival benefit. The results illustrate the potential of mechanism-guided therapeutic co-targeting of disparate biological vulnerabilities in the tumor microenvironment.
Subject(s)
Glioblastoma , Animals , Antidepressive Agents, Tricyclic/metabolism , Antidepressive Agents, Tricyclic/therapeutic use , Autophagy , B7-H1 Antigen/metabolism , Glioblastoma/pathology , Imipramine/metabolism , Imipramine/therapeutic use , Immune Checkpoint Inhibitors , Immunotherapy , Macrophages/metabolism , Mice , Neoplasm Recurrence, Local/drug therapy , Programmed Cell Death 1 Receptor , Tumor Microenvironment , Vascular Endothelial Growth Factor A/metabolismABSTRACT
PURPOSE: The present study aimed to elucidate the transport properties of imipramine and paroxetine, which are the antidepressants, across the blood-brain barrier (BBB) in rats. METHODS: In vivo influx and efflux transport of imipramine and paroxetine across the BBB were tested using integration plot analysis and a combination of brain efflux index and brain slice uptake studies, respectively. Conditionally immortalized rat brain capillary endothelial cells, TR-BBB13 cells, were utilized to characterize imipramine and paroxetine transport at the BBB in vitro. RESULTS: The in vivo influx clearance of [3H]imipramine and [3H]paroxetine in rats was determined to be 0.322 mL/(min·g brain) and 0.313 mL/(min·g brain), respectively. The efflux clearance of [3H]imipramine and [3H]paroxetine was 0.380 mL/(min·g brain) and 0.126 mL/(min·g brain), respectively. These results suggest that the net flux of paroxetine, but not imipramine, at the BBB in vivo was dominated by transport to the brain from the circulating blood. The uptake of imipramine and paroxetine by TR-BBB13 cells exhibited time- and temperature-dependence and one-saturable kinetics with a Km of 37.6 µM and 89.2 µM, respectively. In vitro uptake analyses of extracellular ion dependency and the effect of substrates/inhibitors for organic cation transporters and transport systems revealed minor contributions to known transporters and transport systems and the difference in transport properties in the BBB between imipramine and paroxetine. CONCLUSIONS: Our study showed the comprehensive outcomes of imipramine and paroxetine transport at the BBB, implying that molecular mechanism(s) distinct from previously reported transporters and transport systems are involved in the transport.
Subject(s)
Antidepressive Agents, Second-Generation/metabolism , Antidepressive Agents, Tricyclic/metabolism , Blood-Brain Barrier/metabolism , Imipramine/metabolism , Membrane Transport Proteins/metabolism , Paroxetine/metabolism , Animals , Antidepressive Agents, Second-Generation/administration & dosage , Antidepressive Agents, Tricyclic/administration & dosage , Biological Transport , Cell Line , Imipramine/administration & dosage , Injections, Intravenous , Kinetics , Male , Models, Biological , Paroxetine/administration & dosage , Permeability , Rats, WistarABSTRACT
A poly(methacrylic acid-co-ethylene glycol dimethacrylate)-based magnetic sorbent was used for the rapid and sensitive determination of tricyclic antidepressants and their main active metabolites in human urine. This material was characterized by magnetism measurements, zeta potential, scanning electron microscopy, nitrogen adsorption-desorption isotherms, and thermogravimetric analysis. The proposed analytical method is based on stir bar sorptive-dispersive microextraction (SBSDME) followed by liquid chromatography-tandem mass spectrometry. The main parameters involved in the extraction step were optimized by using the response surface methodology as a multivariate optimization method, whereas a univariate approach was employed to study the desorption parameters. Under the optimized conditions, the proposed method was properly validated showing good linearity (at least up to 50 ng mL-1) and enrichment factors (13-22), limits of detection and quantification in the low ng L-1 range (1.4-7.0 ng L-1), and good intra- and inter-day repeatability (relative standard deviations below 15%). Matrix effects were observed for the direct analysis of urine samples, but they were negligible when a 1:1 v/v dilution with deionized water was performed. Finally, the method was successfully applied to human urine samples from three volunteers, one of them consuming a prescribed drug for depression that tested positive for clomipramine and its main active metabolite. Quantitative relative recoveries (80-113%) were obtained by external calibration. The present work expands the applicability of the SBSDME to new analytes and new types of magnetic sorbents.
Subject(s)
Antidepressive Agents, Tricyclic , Polymethacrylic Acids , Solid Phase Microextraction , Humans , Antidepressive Agents, Tricyclic/chemistry , Antidepressive Agents, Tricyclic/metabolism , Antidepressive Agents, Tricyclic/urine , Chromatography, Liquid , Cobalt/chemistry , Ferric Compounds/chemistry , Magnetic Phenomena , Polymethacrylic Acids/chemistry , Silicon Dioxide/chemistry , Solid Phase Microextraction/methods , Tandem Mass SpectrometryABSTRACT
In this study, we explore the inhibitory effects of protriptyline, a tricyclic antidepressant drug, on voltage-dependent K+ (Kv) channels of rabbit coronary arterial smooth muscle cells using a whole-cell patch clamp technique. Protriptyline inhibited the vascular Kv current in a concentration-dependent manner, with an IC50 value of 5.05 ± 0.97 µM and a Hill coefficient of 0.73 ± 0.04. Protriptyline did not affect the steady-state activation kinetics. However, the drug shifted the steady-state inactivation curve to the left, suggesting that protriptyline inhibited the Kv channels by changing their voltage sensitivity. Application of 20 repetitive train pulses (1 or 2 Hz) progressively increased the protriptyline-induced inhibition of the Kv current, suggesting that protriptyline inhibited Kv channels in a use (state)-dependent manner. The extent of Kv current inhibition by protriptyline was similar during the first, second, and third step pulses. These results suggest that protriptyline-induced inhibition of the Kv current mainly occurs principally in the closed state. The increase in the inactivation recovery time constant in the presence of protriptyline also supported use (state)-dependent inhibition of Kv channels by the drug. In the presence of the Kv1.5 inhibitor, protriptyline did not induce further inhibition of the Kv channels. However, pretreatment with a Kv2.1 or Kv7 inhibitor induced further inhibition of Kv current to a similar extent to that observed with protriptyline alone. Thus, we conclude that protriptyline inhibits the vascular Kv channels in a concentration- and use-dependent manner by changing their gating properties. Furthermore, protriptyline-induced inhibition of Kv channels mainly involves the Kv1.5.
Subject(s)
Myocytes, Smooth Muscle/metabolism , Potassium Channels, Voltage-Gated/drug effects , Protriptyline/pharmacology , Animals , Antidepressive Agents, Tricyclic/metabolism , Antidepressive Agents, Tricyclic/pharmacology , Coronary Vessels/metabolism , Dose-Response Relationship, Drug , Male , Membrane Potentials/drug effects , Muscle, Smooth, Vascular/cytology , Muscle, Smooth, Vascular/metabolism , Myocytes, Smooth Muscle/cytology , Myocytes, Smooth Muscle/drug effects , Patch-Clamp Techniques , Potassium Channels, Voltage-Gated/metabolism , Protriptyline/metabolism , RabbitsABSTRACT
Pharmacogenomics, ie, the study of how an individual's genomic profile influences his or her response to drugs, has emerged as a clinical tool to optimize drug therapy. Certain variants in some genes increase the risk of severe, life-threatening adverse effects from certain drugs. Integrating pharmacogenomics into clinical practice to assist in drug selection and dosing has the potential to improve the outcomes of treatment, reduce the risk of drug-induced morbidity and death, and be cost-effective.
Subject(s)
Codeine/metabolism , Cytochrome P-450 CYP2D6/genetics , Pharmacogenetics , Pharmacogenomic Variants , Precision Medicine , Antidepressive Agents, Tricyclic/adverse effects , Antidepressive Agents, Tricyclic/metabolism , Clopidogrel/metabolism , Codeine/adverse effects , Cytochrome P-450 CYP2C19/genetics , Direct-To-Consumer Screening and Testing , Genetic Testing/economics , Genotype , Humans , Pharmacogenetics/economics , Pharmacogenetics/education , Pharmacogenetics/organization & administration , Selective Serotonin Reuptake Inhibitors/adverse effects , Selective Serotonin Reuptake Inhibitors/metabolismABSTRACT
The inhibitory activity of (±)-citalopram on human (h) α3ß4, α4ß2, and α7 nicotinic acetylcholine receptors (AChRs) was determined by Ca2+ influx assays, whereas its effect on rat α9α10 and mouse habenular α3ß4* AChRs by electrophysiological recordings. The Ca2+ influx results clearly establish that (±)-citalopram inhibits (IC50's in µM) hα3ß4 AChRs (5.1⯱â¯1.3) with higher potency than that for hα7 (18.8⯱â¯1.1) and hα4ß2 (19.1⯱â¯4.2) AChRs. This is in agreement with the [3H]imipramine competition binding results indicating that (±)-citalopram binds to imipramine sites at desensitized hα3ß4 with >2-fold higher affinity than that for hα4ß2. The electrophysiological, molecular docking, and in silico mutation results indicate that (±)-citalopram competitively inhibits rα9α10 AChRs (7.5 ± 0.9) in a voltage-independent manner by interacting mainly with orthosteric sites, whereas it inhibits a homogeneous population of α3ß4* AChRs at MHb (VI) neurons (7.6 ± 1.0) in a voltage-dependent manner by interacting mainly with a luminal site located in the middle of the ion channel, overlapping the imipramine site, which suggests an ion channel blocking mechanism. In conclusion, (±)-citalopram inhibits α3ß4 and α9α10 AChRs with higher potency compared to other AChRs but by different mechanisms. (±)-Citalopram also inhibits habenular α3ß4*AChRs, supporting the notion that these receptors are important endogenous targets related to their anti-addictive activities.
Subject(s)
Antidepressive Agents/pharmacology , Citalopram/pharmacology , Habenula/metabolism , Receptors, Nicotinic/drug effects , Animals , Antidepressive Agents, Tricyclic/metabolism , Binding, Competitive/drug effects , Calcium/metabolism , HEK293 Cells , Habenula/drug effects , Humans , Imipramine/metabolism , Models, Molecular , Molecular Docking Simulation , Molecular Dynamics Simulation , Patch-Clamp Techniques , Receptors, Nicotinic/metabolism , XenopusABSTRACT
The wide spread use of central nervous system (CNS) drugs has caused thousands of deaths in clinical practice while there are few antidotes or effective treatments to decrease their accumulation in CNS. In this study, we used amitriptyline (AMI) and dexamethasone (DEX) as the corresponding poisoning and pre-protecting drugs, respectively, to study whether DEX has the potential to reduce AMI accumulation in brain. By measuring the pharmacokinetic data of AMI and its main metabolite nortriptyline (NOR), we found that DEX possibly accelerated the metabolism and elimination of AMI with minimal effects on the concentrations of NOR in blood. Nevertheless, the results indicated that DEX reduced the brain/plasma concentration ratio of AMI and NOR, even if the plasma concentration of NOR had an upward trend. Western blot results showed the overexpression of cyp3a2 and P-gp in rat liver and brain capillaries tissues. We propose that cyp3a2 and P-gp could be upregulated in the liver and blood-brain barrier (BBB) when using DEX. Further experiments suggest that DEX may serve as the ligand of PXR to induce P-gp expression.
Subject(s)
ATP Binding Cassette Transporter, Subfamily B, Member 1/metabolism , Amitriptyline/pharmacokinetics , Antidepressive Agents, Tricyclic/pharmacokinetics , Blood-Brain Barrier/metabolism , Brain/metabolism , Cytochrome P-450 CYP3A/metabolism , Dexamethasone/pharmacology , ATP Binding Cassette Transporter, Subfamily B, Member 1/genetics , Amitriptyline/blood , Amitriptyline/metabolism , Amitriptyline/poisoning , Animals , Antidepressive Agents, Tricyclic/blood , Antidepressive Agents, Tricyclic/metabolism , Antidepressive Agents, Tricyclic/poisoning , Brain/blood supply , Capillaries/metabolism , Cytochrome P-450 CYP3A/genetics , Gene Expression/drug effects , Liver/metabolism , Male , Rats, Sprague-Dawley , Up-RegulationABSTRACT
Active pharmaceutical ingredients such as tricyclic antidepressants (TCAs) are contaminants of emerging concern which are commonly detected in wastewater effluent and which can disrupt the behavior of non-target organisms. In aquatic snails, the righting response is a critical behavior that has been shown to be inhibited by exposure to SSRI-type antidepressants. We exposed marine and freshwater snails to three tricyclic antidepressants (clomipramine, amitriptyline, and imipramine) for 1 h and measured righting response time. In the marine mud snail (Ilyanassa obsoleta), all three TCAs significantly increased righting time at concentrations as low as 156 µg/L. Similarly, in the freshwater snail Leptoxis carinata, all three TCAs increased righting time at concentrations as low as 263 µg/L. However, exposure to imipramine from 15.8 to 316 µg/L resulted in significantly faster righting time. Such low-dose stimulation and high-dose inhibition are characteristics of a hormetic response. We discuss the possible physiological mechanism of action of TCAs and other antidepressants on snail behavior, and the occurrence of non-monotonic, hormetic dose responses to human pharmaceuticals in the aquatic environment.
Subject(s)
Antidepressive Agents, Tricyclic/toxicity , Imipramine/toxicity , Snails/physiology , Water Pollutants, Chemical/toxicity , Animals , Antidepressive Agents , Antidepressive Agents, Tricyclic/metabolism , Fresh Water , Humans , Imipramine/metabolism , Wastewater , Water Pollutants, Chemical/metabolismABSTRACT
Antidepressant drugs are recommended for the treatment of Parkinson's disease (PD)-associated depression but their role in the modulation of L-DOPA-induced behavioral and neurochemical markers is poorly explored. The aim of the present study was to examine the impact of the tricyclic antidepressant amitriptyline and L-DOPA, administered chronically alone or in combination, on rotational behavior, monoamine levels and binding of radioligands to their transporters in the dopaminergic brain structures of unilaterally 6-OHDA-lesioned rats. Binding of [3H]nisoxetine to noradrenaline transporter (NET), [3H]GBR 12,935 to dopamine transporter (DAT) and [3H]citalopram to serotonin transporter (SERT) were analyzed by autoradiography. Amitriptyline administered alone did not induce rotational behavior but in combination with L-DOPA increased the number of contralateral rotations much more strongly than L-DOPA alone. The combined treatment also significantly increased the tissue dopamine (DA) content in the ipsilateral striatum and substantia nigra (SN) vs. L-DOPA alone. 6-OHDA-mediated lesion of nigrostriatal DA neurons drastically reduced DAT and NET bindings in the ipsilateral striatum. In the ipsilateral SN, DAT binding decreased while NET binding rose. SERT binding increased significantly mainly in the SN. Amitriptyline administered alone or jointly with L-DOPA had no effect on DAT binding on the lesioned side, significantly decreased SERT binding in the striatum and SN while NET binding only in the SN. Since in the DA-denervated striatum, SERT is mainly responsible for reuptake of L-DOPA-derived DA while in the SN, SERT and NET are involved, the inhibition of these transporters by antidepressant drugs may improve dopaminergic transmission and consequently motor behavior.
Subject(s)
Amitriptyline/metabolism , Antidepressive Agents, Tricyclic/metabolism , Corpus Striatum/metabolism , Levodopa/metabolism , Parkinsonian Disorders/metabolism , Substantia Nigra/metabolism , Amitriptyline/pharmacology , Animals , Antidepressive Agents, Tricyclic/pharmacology , Corpus Striatum/drug effects , Drug Interactions/physiology , Levodopa/pharmacology , Male , Motor Activity/drug effects , Motor Activity/physiology , Oxidopamine/toxicity , Parkinsonian Disorders/physiopathology , Rats , Rats, Wistar , Rotation , Substantia Nigra/drug effectsABSTRACT
A large number of Food and Drug Administration (FDA)-approved drugs have been found to inhibit the cell entry of Ebola virus (EBOV). However, since these drugs have various primary pharmacological targets, their mechanisms of action against EBOV remain largely unknown. We have previously shown that six FDA-approved drugs inhibit EBOV infection by interacting with and destabilizing the viral glycoprotein (GP). Here we show that antidepressants imipramine and clomipramine and antipsychotic drug thioridazine also directly interact with EBOV GP and determine the mode of interaction by crystallographic analysis of the complexes. The compounds bind within the same pocket as observed for other, chemically divergent complexes but with different binding modes. These details should be of value for the development of potent EBOV inhibitors.
Subject(s)
Antidepressive Agents, Tricyclic/metabolism , Antipsychotic Agents/metabolism , Ebolavirus , Glycoproteins/metabolism , Viral Proteins/metabolism , Binding Sites , Drug Design , Glycoproteins/chemistry , Models, Molecular , Protein Binding , Protein Conformation , Viral Proteins/chemistryABSTRACT
The activity of tricyclic antidepressants (TCAs) at α7 and α9α10 nicotinic acetylcholine receptors (AChRs) as well as at hippocampal α7-containing (i.e., α7*) AChRs is determined by using Ca2+ influx and electrophysiological recordings. To determine the inhibitory mechanisms, additional functional tests and molecular docking experiments are performed. The results established that TCAs (a) inhibit Ca2+ influx in GH3-α7 cells with the following potency (IC50 in µM) rank: amitriptyline (2.7⯱â¯0.3) > doxepin (5.9⯱â¯1.1) â¼ imipramine (6.6⯱â¯1.0). Interestingly, imipramine inhibits hippocampal α7* AChRs (42.2⯱â¯8.5⯵M) in a noncompetitive and voltage-dependent manner, whereas it inhibits α9α10 AChRs (0.53⯱â¯0.05⯵M) in a competitive and voltage-independent manner, and (b) inhibit [3H]imipramine binding to resting α7 AChRs with the following affinity rank (IC50 in µM): imipramine (1.6⯱â¯0.2) > amitriptyline (2.4⯱â¯0.3) > doxepin (4.9⯱â¯0.6), whereas imipramine's affinity was no significantly different to that for the desensitized state. The molecular docking and functional results support the notion that imipramine noncompetitively inhibits α7 AChRs by interacting with two overlapping luminal sites, whereas it competitively inhibits α9α10 AChRs by interacting with the orthosteric sites. Collectively our data indicate that TCAs inhibit α7, α9α10, and hippocampal α7* AChRs at clinically relevant concentrations and by different mechanisms of action.
Subject(s)
Antidepressive Agents, Tricyclic/pharmacology , Hippocampus/drug effects , Hippocampus/metabolism , Receptors, Nicotinic/metabolism , alpha7 Nicotinic Acetylcholine Receptor/antagonists & inhibitors , Animals , Antidepressive Agents, Tricyclic/metabolism , Binding Sites , Cell Line , Drug Interactions , Imipramine/pharmacology , Molecular Docking Simulation , Molecular Dynamics Simulation , Protein Conformation , Rats , Receptors, Nicotinic/chemistry , Thermodynamics , alpha7 Nicotinic Acetylcholine Receptor/chemistry , alpha7 Nicotinic Acetylcholine Receptor/metabolismABSTRACT
Despite the fact that pharmaceuticals occur in surface water in low concentrations, they can still have adverse effect on aquatic biota, because these substances are designed to have a specific mode of action even at low concentrations. To our knowledge, only little is known about the long-term effect of tricyclic antidepressant residues on non-target animals, especially fish. Hence, the aim of this study was to assess the effect of subchronic exposure to selected tricyclic antidepressants - namely, amitriptyline, nortriptyline, and clomipramine and their mixture - on early-life stages of common carp (Cyprinus carpio). Two embryo-larval toxicity tests with common carp were performed, each taking 30 days. For the experiment, three tricyclic antidepressants as well as their mixtures were tested at three different concentrations - 10, 100 and 500 µg/L. Hatching and mortality were recorded twice a day. In addition, samples were taken at regular intervals in order to record developmental stage, morphometric and condition characteristics and morphological anomalies. At the end of the test, additional samples were taken for histopathological examination and also for the determination of antioxidant and biotransformation enzyme activity and lipid peroxidation and protein carbonylation. Long-term exposure resulted in a significant increase in mortality, developmental retardation, morphological anomalies, and pathological changes in brain, heart, and cranial and caudal kidney. In addition, changes in antioxidant enzyme activity as well as increased lipid peroxidation were observed, even at the lowest tested concentrations. Hence, environmentally relevant concentrations of TCAs have the potential to cause harmful effects on early-life stages of fish.
Subject(s)
Antidepressive Agents, Tricyclic/toxicity , Carps/physiology , Water Pollutants, Chemical/toxicity , Animals , Antidepressive Agents, Tricyclic/metabolism , Antioxidants/metabolism , Embryo, Nonmammalian/drug effects , Larva/drug effects , Lipid Peroxidation/drug effects , Water Pollutants, Chemical/metabolismABSTRACT
Cationic amphiphilic drugs (CADs) comprise a wide variety of different substance classes such as antidepressants, antipsychotics, and antiarrhythmics. It is well recognized that CADs accumulate in certain intracellular compartments leading to specific morphological changes of cells. So far, no adequate technique exists allowing for ultrastructural analysis of CAD in intact cells. Azidobupramine, a recently described multifunctional antidepressant analogue, allows for the first time to perform high-resolution studies of CADs on distribution pattern and morphological changes in intact cells. We showed here that the intracellular distribution pattern of azidobupramine strongly depends on drug concentration and exposure time. The mitochondrial compartment (mDsRed) and the late endo-lysosomal compartment (CD63-GFP) were the preferred localization sites at low to intermediate concentrations (i.e. 1 µM, 5 µM). In contrast, the autophagosomal compartment (LC3-GFP) can only be reached at high concentrations (10 µM) and long exposure times (72 hrs). At the morphological level, LC3-clustering became only prominent at high concentrations (10 µM), while changes in CD63 pattern already occurred at intermediate concentrations (5 µM). To our knowledge, this is the first study that establishes a link between intracellular CAD distribution pattern and morphological changes. Therewith, our results allow for gaining deeper understanding of intracellular effects of CADs.
Subject(s)
Antidepressive Agents, Tricyclic/metabolism , Cations/metabolism , Intracellular Space/metabolism , Pharmaceutical Preparations/metabolism , Antidepressive Agents, Tricyclic/chemistry , Antidepressive Agents, Tricyclic/pharmacokinetics , Autophagosomes/metabolism , Cations/chemistry , Cations/pharmacokinetics , HeLa Cells , Humans , Hydrophobic and Hydrophilic Interactions , Lysosomes/metabolism , Mitochondria/metabolism , Pharmaceutical Preparations/chemistryABSTRACT
Depression is a debilitating chronic illness that affects around 350 million people worldwide. Current treatments, such as selective serotonin reuptake inhibitors, are not ideal because only a fraction of patients achieve remission. Tianeptine is an effective antidepressant with a previously unknown mechanism of action. We recently reported that tianeptine is a full agonist at the mu opioid receptor (MOR). Here we demonstrate that the acute and chronic antidepressant-like behavioral effects of tianeptine in mice require MOR. Interestingly, while tianeptine also produces many opiate-like behavioral effects such as analgesia and reward, it does not lead to tolerance or withdrawal. Furthermore, the primary metabolite of tianeptine (MC5), which has a longer half-life, mimics the behavioral effects of tianeptine in a MOR-dependent fashion. These results point to the possibility that MOR and its downstream signaling cascades may be novel targets for antidepressant drug development.
Subject(s)
Antidepressive Agents, Tricyclic/pharmacology , Receptors, Opioid, mu/metabolism , Thiazepines/pharmacology , Analgesics, Opioid/pharmacology , Animals , Antidepressive Agents, Tricyclic/metabolism , Antidepressive Agents, Tricyclic/pharmacokinetics , Depressive Disorder/drug therapy , Depressive Disorder/metabolism , Dose-Response Relationship, Drug , Drug Tolerance , HEK293 Cells , Humans , Male , Mice, Inbred C57BL , Mice, Knockout , Molecular Structure , Morphine/pharmacology , Receptors, Opioid, mu/agonists , Receptors, Opioid, mu/genetics , Thiazepines/metabolism , Thiazepines/pharmacokineticsABSTRACT
AIM: Amitriptyline is a widely used tricyclic antidepressant, but the metabolic studies were conducted almost 20 years ago using high-performance liquid chromatography coupled with ultraviolet detector or radiolabeled methods. RESULTS: First, multiple ion monitoring (MIM)- enhanced product ion (EPI) scan was used to obtain the diagnostic ions or neutral losses in human liver microsome incubations with amitriptyline. Subsequently, predicted multiple reaction monitoring (MRM)-EPI scan was used to identify the metabolites in human urine with the diagnostic ions or neutral losses. Finally, product ion filtering and neutral loss filtering were used as the data mining tools to screen metabolites. Consequently, a total of 28 metabolites were identified in human urine after an oral administration using LC-MS/MS. CONCLUSION: An integrated workflow using LC-MS/MS was developed to comprehensively profile the metabolites of amitriptyline in human urine, in which five N-acetyl-l-cysteine conjugates were characterized as tentative biomarkers for idiosyncratic toxicity.
Subject(s)
Amitriptyline/metabolism , Amitriptyline/urine , Antidepressive Agents, Tricyclic/metabolism , Antidepressive Agents, Tricyclic/urine , Microsomes, Liver/metabolism , Acetylcysteine/analysis , Acetylcysteine/metabolism , Acetylcysteine/urine , Amitriptyline/analysis , Antidepressive Agents, Tricyclic/analysis , Chromatography, High Pressure Liquid/methods , Humans , Metabolic Networks and Pathways , Tandem Mass Spectrometry/methodsABSTRACT
CONTEXT: Amitriptyline (AT), one of the tricyclic antidepressants, is still widely used for the treatment of the depression and control of anxiety states and panic disorders in the developing countries. OBJECTIVE: This study evaluates the catalytic activities of CYP2D6*1, CYP2D6*2, CYP2D6*10 and 22 novel alleles in Han Chinese population and their effects on the N-demethylation of AT in vitro. MATERIALS AND METHODS: CYP2D6*1 and 24 CYP2D6 allelic variants were highly expressed in insect cells, and all variants were characterized using AT as a substrate. Reactions were performed at 37 °C with 10-1000 µM substrate for 30 min. We established a HPLC method to quantify the levels of nortriptyline (NT). The kinetic parameters Km, Vmax and intrinsic clearance (Vmax/Km) of NT were calculated. RESULTS: Among the 24 CYP2D6 variants, all variants exhibited decreased intrinsic clearance values compared with wild-type CYP2D6.1. Kinetic parameters of two CYP2D6 variants (CYP2D6*92, *96) could not be determined because of absent enzyme activities. CONCLUSIONS: The comprehensive in vitro assessment of CYP2D6 variants provides significant insight into allele-specific activity towards AT in vivo.