Environmentally relevant concentrations of the tricyclic antidepressant, amitriptyline, affect feeding and reproduction in a freshwater mollusc.

Antidepressant drugs (ADDs) are one of the most extensively used pharmaceuticals globally. They act at particularly low therapeutic concentrations to modulate monoamine neurotransmission, which is one of the most evolutionary conserved pathways in both humans and animal species including invertebrates. As ADDs are widely detected in the aquatic environment at low concentrations (ng/L to low µg/L), their potential to exert drug-target mediated effects in aquatic species has raised serious concerns. Amitriptyline (AMI) is the most widely used tricyclic ADD, while monoamines, the target of ADDs, are major bioregulators of multiple key physiological processes including feeding, reproduction and behaviour in molluscs. However, the effects of AMI on feeding, reproduction and mating behaviour are unknown in molluscs despite their ecological importance, diversity and reported sensitivity to ADDs. To address this knowledge gap, we investigated the effects of environmentally relevant concentrations of AMI (0, 10, 100, 500 and 1000 ng/L) on feeding, reproduction and key locomotor behaviours, including mating, in the freshwater gastropod, Biomphalaria glabrata over a period of 28 days. To further provide insight into the sensitivity of molluscs to ADDs, AMI concentrations (exposure water and hemolymph) were determined using a novel extraction method. The Fish Plasma Model (FPM), a critical tool for prioritization assessment of pharmaceuticals with potential to cause drug target-mediated effects in fish, was then evaluated for its applicability to molluscs for the first time. Disruption of food intake (1000 ng/L) and reproductive output (500 and 1000 ng/L) were observed at particularly low hemolymph levels of AMI, whereas locomotor behaviours were unaffected. Importantly, the predicted hemolymph levels of AMI using the FPM agreed closely with the measured levels. The findings suggest that hemolymph levels of AMI may be a useful indicator of feeding and reproductive disruptions in wild population of freshwater gastropods, and confirm the applicability of the FPM to molluscs for comparative pharmaceutical hazard identification.

Amitriptyline ecotoxicity in Danio rerio (Hamilton, 1822) embryos - similar toxicity profile in the presence of nanoplastics.

Interaction of nanoplastics (NPls) with other environmental contaminants could affect their uptake by the organisms and their toxicity. Thus, the present study aims to investigate the polystyrene NPls (44 nm) interaction with the antidepressant amitriptyline (AMI) and its toxicity to Danio rerio embryos. A similar toxicological profile for NPls + AMI exposure was found for most of the evaluated endpoints, comparing with AMI single exposure, showing that the presence of NPls did not modulate the AMI toxicity. However, the behavioral assessment showed a different pattern with hypoactivity for the NPls + AMI exposure (NPls - hyperactivity; AMI - no effect). Interaction effects between NPls and AMI were also found in the protein contents (antagonism) and in the total glutathione content (synergism). This study highlights the complexity and unpredictability of NPls interaction with pharmaceuticals, important for an accurate environmental risk assessment and for the developing of effective strategies and interventions against plastic pollution.

Effects of parental exposure to amitriptyline on the survival, development, behavior, and gene expression in zebrafish offspring.

In mammals, parental exposure to amitriptyline (AMI) has been proven to contribute to congenital disabilities in their offspring. However, no studies have paid attention to the adverse effects of parental exposure to amitriptyline on fish offspring. In this study, we exposed adult zebrafish (F0) to AMI (0.8 µg/L) for 21 days. Subsequently, these zebrafish (F0) were allowed to mate, and their offspring (F1) were collected to culture in clean water for 5 days. The mortality rate, average hatching time, and heart rate at 48 h post-fertilization (hpf) of F1 were investigated. Our results showed that parental exposure to AMI induced tachycardia and increased mortality in F1 zebrafish. Under a light/dark transition test, F1 larvae born from AMI-exposed parents exhibited lower locomotor activity in the dark period and decreased thigmotaxis in the light period. The transcriptome analysis showed that parental AMI exposure dysregulated some key pathways in their offspring. Through the prediction of key driver analysis, six differentially expressed genes (DEGs) were revealed as key driver genes involved in protein processing in endoplasmic reticulum (hspa5, hsp70.1, hsp90a), ribosome (rps27a) and PPAR signaling pathway (pparab and fabp2). Considering that the concentration of AMI residual components in natural water bodies may be over our test concentration (0.8 µg/L), our findings suggested that toxicity of parental exposure to the offspring of fish should receive greater attention.

Combined exposure to microplastics and amitriptyline induced abnormal behavioral responses and oxidative stress in the eyes of zebrafish (Danio rerio).

Many studies have demonstrated that microplastics (MPs) can combine with various coexisting chemical pollutants, increasing their bioavailability and changing the combined toxicity to organisms. However, information on the combined effects of MPs and amitriptyline (AMI, a widely used tricyclic antidepressant) on aquatic species is still limited. In this study, we exposed zebrafish to MPs (2-µm polystyrene beads, 0.44 mg/L), AMI (2.5 µg/L), and their mixture for 7 days and investigated the alternation in their behaviors and ocular oxidative stress. As a result, combined exposure to MPs and AMI could significantly elevate locomotor activity, increase the frequency and duration of shoaling behavior in zebrafish, and alter their post-stimulation behaviors. Although combined exposure to MPs and AMI exhibited stronger behavioral toxicity than individual exposure, no significant interactive effects on the behavioral traits were detected, suggesting that the combined behavioral toxicity appeared to be an additive effect. However, their combined exposure to MPs or AMI significantly decreased the ocular levels of SOD, CAT, and GSH in zebrafish, with significant interaction effects on the CAT activity and GSH content. Significant correlations between some post-stimulation behavioral traits and ocular levels of SOD, CAT, and GSH in zebrafish were detected, suggesting that ocular oxidative stress induced by combined exposure to MPs and AMI may play an important role in their behavioral toxicity.

Combined exposure to microplastics and amitriptyline caused intestinal damage, oxidative stress and gut microbiota dysbiosis in zebrafish (Danio rerio).

The potential toxicity of microplastics (MPs) and hydrophilic pharmaceuticals to aquatic organisms has recently raised great public concern, yet their combined effects on aquatic organisms remain largely unknown. Herein, the combined effects of MPs and the commonly prescribed amitriptyline hydrochloride (AMI) on the intestinal tissue and gut microbiota of zebrafish (Danio rerio) were investigated. Adult zebrafish were exposed to microplastics (polystyrene, PS, 440 µg/L), AMI (2.5 µg/L), PS+AMI (440 µg/L PS + 2.5 µg/L AMI), and dechlorinated tap water (control) for 21 days, respectively. Our results showed that zebrafish rapidly ingested PS beads and accumulated them in the gut. Exposure to PS+AMI significantly enhanced the SOD and CAT activities compared to the control group, suggesting that combined exposure might increase ROS production in the zebrafish gut. Exposure to PS+AMI led to severe gut injuries, including cilia defects, partial absence and cracking of intestinal villi. Exposure to PS+AMI caused shifts in the gut bacterial communities, increasing the abundance of Proteobacteria and Actinobacteriota, and decreasing the abundance of Firmicutes, Bacteroidota and beneficial bacteria Cetobacterium, which caused dysbiosis in the gut microbiota, and subsequently may induce intestinal inflammation. Furthermore, exposure to PS+AMI disordered the predicted metabolic functions of gut microbiota, but functional changes in the PS+AMI group at KEGG level 1 and level 2 were not significantly different from those in the PS group. The results of this study extend our knowledge of the combined effects of MPs and AMI on the health of aquatic organisms, and will be helpful in assessing the combined effects of MPs and tricyclic antidepressants on aquatic organisms.

Anticancer effect of paroxetine and amitriptyline on HT29 and A549 cell lines.

INTRODUCTION: Paroxetine is functionally classified as a selective serotonin reuptake inhibitor. Paroxetine can induce mitochondria-dependent apoptosis through the ROS-MAPK pathway.Amitriptyline is a tricyclic antidepressant. This drug induces the expression of p53, thereby activating caspase-3. Amitriptyline has also been studied as a potential candidate for inducing oxidative stress and cytotoxicity in cancer cells, which may be more effective than other chemotherapy drugs. This study aims to to investigate the anticancer effects of paroxetine and amitriptyline and their combination treatment on HT29 and A549 cell lines for the first time. METHODS: In order to investigate the anticancer effect of two drugs, paroxetine and amitriptyline, on inhibiting the growth of A549 and HT29 cancer cells, oxidative stress factors and LDH enzyme and apoptosis tests were performed. RESULTS: Two drugs, amitriptyline and paroxetine alone, inhibited the growth of cancer cells in such a way that the inhibitory effect of the cells increased with the increase in the dose of the drug. In the simultaneous exposure of these two drugs, the inhibitory effect was much greater than the effect of single drug exposure. Also, these two drugs have caused LDH leakage and induction of apoptosis. CONCLUSION: According to the results of the study, it was found that these two drugs have the necessary ability to inhibit the growth of cancer cells by inducing apoptosis and LDH leakage and inducing oxidative stress.

Exposure to amitriptyline induces persistent gut damages and dysbiosis of the gut microbiota in zebrafish (Danio rerio).

Amitriptyline (AMI), the most commonly prescribed tricyclic antidepressant, is widely detected in water environments. Exposure to AMI may lead to diverse adverse effects on aquatic organisms, but little is known about the effect of short-term exposure to AMI on the gut microbiota of aquatic organisms and their recovery characteristics. In the present study, adult zebrafish (Danio rerio) were exposed to AMI (0, 2.5, 10, and 40 µg/L) for seven days, and then allowed to recover in AMI-free culture water for 21 days. The exposure caused gut damages in all the AMI treated groups of zebrafish, which became more severe after recovery compared to the control group. AMI exposure also disturbed the microbiota of zebrafish guts and rearing water even after the 21-day recovery period. Furthermore, AMI exposure affected microbes involved in the substance and energy metabolic functions in zebrafish guts and tended to increase the abundance of microbial genera associated with opportunistic pathogens. In addition, the microbial predicted metabolic functions in AMI-exposed guts of zebrafish were significantly altered after the 21-day recovery period, explaining the persistent effects of short-term exposure to AMI. The results of this study suggest that acute exposure to AMI may have persistent impacts on the gut histomorphology and the gut microbiota in aquatic organisms.

Persistent impact of amitriptyline on the behavior, brain neurotransmitter, and transcriptional profile of zebrafish (Danio rerio).

Discontinuation of amitriptyline (AMI) has been demonstrated to induce long-term withdrawal syndromes in mammals. However, no studies have focused on the persistent impacts of short-term AMI exposure on teleosts. Here, following exposure to AMI (2.5 and 40 µg/L) for 7 days (E7), zebrafish were transferred into AMI-free water to recover for 21 days (R21). The behavior, brain neurotransmitters, and brain transcriptional profiles were investigated on E7 and R21. AMI exposure induced persistent hypoactivity (2.5 and 40 µg/L) and abnormal schooling behavior (40 µg/L). AMI also induced long-term impacts on the brain serotonin (5-HT), 5-hydroxyindoleacetic acid, norepinephrine, and acetylcholine levels, several of which showed significant correlations with the locomotor activity or schooling behavior. Transcriptional analysis revealed persistent dysregulation in the pathways involved in the circadian rhythm, glycan biosynthesis and metabolism, and axon guidance in brain samples. Twelve genes were predicted as key driver genes in response to AMI exposure, and their significantly differential expression may direct changes across the related molecular networks. Moreover, upregulated brain 5-HT may serve as the central modulator of the persistent AMI pathogenesis in zebrafish. Considering AMI residues in natural waters may temporarily exceed µg/L, corresponding persistent adverse effects on teleosts should not be ignored.

Molecular Consequences of Depression Treatment: A Potential In Vitro Mechanism for Antidepressants-Induced Reprotoxic Side Effects.

The incidence of depression among humans is growing worldwide, and so is the use of antidepressants. However, our fundamental understanding regarding the mechanisms by which these drugs function and their off-target effects against human sexuality remains poorly defined. The present study aimed to determine their differential toxicity on mouse spermatogenic cells and provide mechanistic data of cell-specific response to antidepressant and neuroleptic drug treatment. To directly test reprotoxicity, the spermatogenic cells (GC-1 spg and GC-2 spd cells) were incubated for 48 and 96 h with amitriptyline (hydrochloride) (AMI), escitalopram (ESC), fluoxetine (hydrochloride) (FLU), imipramine (hydrochloride) (IMI), mirtazapine (MIR), olanzapine (OLZ), reboxetine (mesylate) (REB), and venlafaxine (hydrochloride) (VEN), and several cellular and biochemical features were assessed. Obtained results reveal that all investigated substances showed considerable reprotoxic potency leading to micronuclei formation, which, in turn, resulted in upregulation of telomeric binding factor (TRF1/TRF2) protein expression. The TRF-based response was strictly dependent on p53/p21 signaling and was followed by irreversible G2/M cell cycle arrest and finally initiation of apoptotic cell death. In conclusion, our findings suggest that antidepressants promote a telomere-focused DNA damage response in germ cell lines, which broadens the established view of antidepressants' and neuroleptic drugs' toxicity and points to the need for further research in this topic with the use of in vivo models and human samples.

Evaluation of the testicular protection conferred by damiana (Turnera diffusa Willd.) against amitriptyline-induced testicular toxicity, DNA damage and apoptosis in rats.

Psychiatric drugs, such as antidepressants, are used to treat depression based on their ability to modify chemical imbalances of the key neurotransmitters in the brain, including dopamine, serotonin, and norepinephrine. Amitriptyline, a first-reference tricyclic antidepressant derived from dibenzocycloheptadine, is frequently used, especially in neuropsychiatry, to address general depression, major depressive disorders, and fibromyalgia. Therefore, this study attempted to examine the sexual dysfunction attendant on the use of Amitriptyline by investigating the protective role that can be played by damiana. To this end, this study used damiana (Turnera diffusa Willd.) as adjuvant therapy in male albino rats receiving Amitriptyline. Sixty male albino rats were randomly allocated to six groups, with 10 rats being assigned to each group; the first group was a control, the second was treated with damiana only, the third group was given Amitriptyline, the fourth group received Amitriptyline and damiana simultaneously, the fifth group was given Amitriptyline and post-treated with damiana, and the sixth group was given Amitriptyline and then allowed time for self-healing. The findings of this study suggest that oxidative stress occurs in testicular tissue in rat groups treated with Amitriptyline, as manifested by inappropriate activity of TBARS, SOD, GSH, GR, GST, and GPx. Amitriptyline also repressed reproductive hormonal activity, as confirmed by histopathological lesions, DNA damage, and p53 protein expression. The addition of damiana, however, showed aprotective role in all testicular activity indices.

Mass spectrometry imaging reveals lipid upregulation and bile acid changes indicating amitriptyline induced steatosis in a rat model.

As a consequence of the detoxification process, drugs and drug related metabolites can accumulate in the liver, resulting in drug induced liver injury (DILI), which is the major cause for dose limitation. Amitriptyline, a commonly used tricyclic anti-depressant, is known to cause DILI. The mechanism of Amitriptyline induced liver injury is not yet completely understood. However, as it undergoes extensive hepatic metabolism, unraveling the molecular changes in the liver upon Amitriptyline treatment can help understand Amitriptyline's mode of toxicity. In this study, Amitriptyline treated male rat liver tissue was analyzed using Matrix Assisted Laser Desorption/Ionization-Mass Spectrometry Imaging (MALDI-MSI) to investigate the spatial abundances of Amitriptyline, lipids, and bile acids. The metabolism of Amitriptyline in liver tissue was successfully demonstrated, as the spatial distribution of Amitriptyline and its metabolites localize throughout treatment group liver samples. Several lipids appear upregulated, from which nine were identified as distinct phosphatidylcholine (PC) species. The detected bile acids were found to be lower in Amitriptyline treatment group. The combined results from histological findings, Oil Red O staining, and lipid zonation by MSI revealed lipid upregulation in the periportal area indicating drug induced macrovesicular steatosis (DIS).

Beneficial effects of edaravone in experimental model of amitriptyline-induced cardiotoxicity in rats.

Amitriptyline (AMT) cardiotoxicity is commonly seen with high morbidity and mortality rates in emergency departments. Nevertheless, there are still no effective treatment options for amitriptyline-induced cardiotoxicity. The aim of the present study was to evaluate the effects of edaravone, a potent antioxidant and free radical scavenger, in rats by electrocardiographic (ECG), biochemical, and scintigraphic methods. Twenty-eight male Wistar rats were randomly divided into four groups as untreated control (CON), amitriptyline-induced cardiotoxicity (AMT), edaravone treatment (EDO), and amitriptyline + edaravone treatment (AMT+EDO). Cardiotoxicity was induced by intraperitoneal (i.p.) injection of a single-dose amitriptyline (100 mg/kg). Edaravone was administered at a dose of 30 mg/kg (i.p.) after amitriptyline injection. ECG, biochemical, and scintigraphic changes due to edaravone were analyzed. AMT cardiotoxicity was characterized with conduction abnormalities (increased QRS complex, T wave, and duration of QT interval and elevation of ST segment amplitude), elevated 99mTechnetium Pyrophosphate (99mTc-PYP) uptake level, and increased cardiac troponin T level (cTnT). Edaravone treatment significantly decreased all amitriptyline-associated conduction abnormalities in ECG (p < 0.001), 99mTc-PYP uptake (p < 0.001), and serum cTnT level (p < 0.001). 99mTc-PYP scintigraphy can show amitriptyline cardiotoxicity as well as ECG abnormalities and increased values of cTnT. According to the results of the present study, edaravone has strong beneficial effects on amitriptyline-induced cardiotoxicity.

Neuronal life or death linked to depression treatment: the interplay between drugs and their stress-related outcomes relate to single or combined drug therapies.

Depression is a serious medical condition, typically treated by antidepressants. Conventional monotherapy can be effective only in 60-80% of patients, thus modern psychiatry deals with the challenge of new methods development. At the same moment, interactions between antidepressants and the occurrence of potential side effects raise serious concerns, which are even more exacerbated by the lack of relevant data on exact molecular mechanisms. Therefore, the aims of the study were to provide up-to-date information on the relative mechanisms of action of single antidepressants and their combinations. In this study, we evaluated the effect of single and combined antidepressants administration on mouse hippocampal neurons after 48 and 96 h in terms of cellular and biochemical features in vitro. We show for the first time that co-treatment with amitriptyline/imipramine + fluoxetine initiates in cells adaptation mechanisms which allow cells to adjust to stress and finally exerts less toxic events than in cells treated with single antidepressants. Antidepressants treatment induces in neuronal cells oxidative and nitrosative stress, which leads to micronuclei and double-strand DNA brakes formation. At this point, two different mechanistic events are initiated in cells treated with single and combined antidepressants. Single antidepressants (amitriptyline, imipramine or fluoxetine) activate cell cycle arrest resulting in proliferation inhibition. On the other hand, treatment with combined antidepressants (amitriptyline/imipramine + fluoxetine) initiates p16-dependent cell cycle arrest, overexpression of telomere maintenance proteins and finally restoration of proliferation. In conclusion, our findings may pave the way to better understanding of the stress-related effects on neurons associated with mono- and combined therapy with antidepressants.

Klotho-Mediated Changes in Shelterin Complex Promote Cytotoxic Autophagy and Apoptosis in Amitriptyline-Treated Hippocampal Neuronal Cells.

Amitriptyline, antidepressant frequently prescribed for treatment of depressive disorders and several neuropathic and inflammatory diseases, has been shown to cause neurotoxic effects. This effect has been partially linked with increased oxidative stress and apoptosis initiation; however, the exact mechanism is still unknown. Klotho protein due to its neuroprotective characteristics seems to be involved in the amitriptyline-mediated neurotoxicity. In this study, we have evaluated the effect of klotho silencing on mouse hippocampal cells exposed to amitriptyline. We show, for the first time, that klotho silencing intensified in hippocampal neurons amitriptyline-induced imbalance in oxido-nitrosative and mineral homeostasis, genomic instability associated with telomere dysfunction what resulted in p16- and p53/p21-mediated cell cycle arrest and activation of autophagy and apoptotic cell death in consequence. Therefore, these results indicate that klotho serves as a part of the cellular defense mechanism engaged in the protection of neurons against amitriptyline-mediated toxicity.

Amitriptyline at an Environmentally Relevant Concentration Alters the Profile of Metabolites Beyond Monoamines in Gilt-Head Bream.

The antidepressant amitriptyline is a widely used selective serotonin reuptake inhibitor that is found in the aquatic environment. The present study investigates alterations in the brain and the liver metabolome of gilt-head bream (Sparus aurata) after exposure at an environmentally relevant concentration (0.2 µg/L) of amitriptyline for 7 d. Analysis of variance-simultaneous component analysis is used to identify metabolites that distinguish exposed from control animals. Overall, alterations in lipid metabolism suggest the occurrence of oxidative stress in both the brain and the liver-a common adverse effect of xenobiotics. However, alterations in the amino acid arginine are also observed. These are likely related to the nitric oxide system that is known to be associated with the mechanism of action of antidepressants. In addition, changes in asparagine and methionine levels in the brain and pantothenate, uric acid, and formylisoglutamine/N-formimino-L-glutamate levels in the liver could indicate variation of amino acid metabolism in both tissues; and the perturbation of glutamate in the liver implies that the energy metabolism is also affected. These results reveal that environmentally relevant concentrations of amitriptyline perturb a fraction of the metabolome that is not typically associated with antidepressant exposure in fish. Environ Toxicol Chem 2019;00:1-13. © 2019 SETAC.

Tricyclic antidepressant amitriptyline inhibits autophagic flux and prevents tube formation in vascular endothelial cells.

Amitriptyline is a tricyclic antidepressant and an inhibitor of lysosomal acid sphingomyelinase (ASM). Amitriptyline is well known for its cardiovascular side effects and toxicity in psychiatric patients. However, the mechanisms underlying the cardiovascular side effects of amitriptyline remain largely undefined. This study aimed to determine the effects of amitriptyline on angiogenic capability of vascular endothelial cells in physiological settings and identify its mechanism of action. The ex vivo aortic ring angiogenesis and in vitro-cultured endothelial cell tube formation assay were used to assess the effects of amitriptyline on endothelial angiogenic capability. It was demonstrated that amitriptyline impaired the angiogenesis of aortic rings, which was similar to that found in aortic rings with haploinsufficiency of the ASM gene. In cultured mouse microvascular endothelial cells (MVECs), amitriptyline impaired the proliferation and tube formation under basal condition, which were accompanied by attenuated angiogenic signalling pathways such as endothelial nitric oxide synthase, Akt and Erk1/2 pathways. Mechanistically, amitriptyline inhibited autophagic flux without affecting autophagosome biogenesis at basal condition. ASM gene silencing or autophagy inhibition mimics the inhibitory effects of amitriptyline on endothelial cell proliferation and tube formation. Collectively, our data suggest that amitriptyline inhibits endothelial cell proliferation and angiogenesis via blockade of ASM-autophagic flux axis. It is implicated that the cardiovascular side effects of amitriptyline may be associated with its inhibitory action on physiological angiogenesis.

The Effects of Lipid Emulsion, Magnesium Sulphate and Metoprolol in Amitriptyline-Induced Cardiovascular Toxicity in Rats.

The aim of this study was to evaluate the effects of metoprolol, lipid emulsion and MgSO4 which can be recommended for prevention of long QT that is one of the lethal consequences of amitriptyline intoxication. Thirty Sprague-Dawley male rats were included. Five groups respectively received the following: saline intraperitoneally (i.p.); amitriptyline (AMT) 100 mg/kg per os (p.o.) and saline i.p.; AMT 100 mg/kg p.o. and 5 mg/kg metoprolol i.p.; AMT 100 mg/kg p.o. and 20 ml/kg lipid emulsion i.p.; AMT 100 mg/kg p.o. and 75 mg/kg MgSO4 i.p. After 1 h, all groups were analysed by ECG recordings in DII lead; their blood was taken for biochemical examination and euthanasia was performed. For histological examination, cardiac tissues were removed and sections were prepared. QTc was significantly reduced in treatment groups compared to the AMT+saline group. When compared with the AMT+saline, lipid emulsion did not affect pro-BNP and troponin levels in biochemical analysis, but it significantly reduced Caspase 3 expression in histological examination. In the group treated with AMT and metoprolol, there was no significant effect on Caspase 3 expression. In MgSO4-treated group, there was a significant decrease in troponin, pro-BNP and urea levels biochemically and significant decrease in Caspase 3 expression histologically when compared with the control group. With further studies including clinical studies, MgSO4, lipid emulsion or metoprolol may be used to improve AMT-induced cardiotoxicity. They can possibly become alternative approaches in the future for suicidal or accidental intoxication of tricyclic antidepressant in emergency departments.

Amitriptyline modulated Ca2+ signaling and induced Ca2+-independent cell viability in human osteosarcoma cells.

Amitriptyline is a widely used tricyclic antidepressant, which acts primarily as a serotonin-norepinephrine reuptake inhibitor. This study examined the effect of amitriptyline on Ca2+ homeostasis and its related mechanism in MG63 human osteosarcoma cells. Amitriptyline evoked cytosolic-free Ca2+ concentrations ([Ca2+]i) rises concentration dependently. Amitriptyline-evoked Ca2+ entry was confirmed by Mn2+-induced quench of fura-2 fluorescence. This entry was inhibited by Ca2+ entry modulators nifedipine, econazole, SKF96365, the protein kinase C (PKC) activator phorbol 12-myristate 13 acetate but was not affected by the PKC inhibitor GF109203X. In Ca2+-free medium, treatment with the endoplasmic reticulum Ca2+ pump inhibitor thapsigargin (TG) inhibited amitriptyline-evoked [Ca2+]i rises by 95%. Conversely, treatment with amitriptyline abolished TG-evoked [Ca2+]i rises. Inhibition of phospholipase C (PLC) with U73122 inhibited amitriptyline-evoked [Ca2+]i rises by 70%. Amitriptyline killed cells at 200-500 µM in a concentration-dependent fashion. Chelating cytosolic Ca2+ with 1,2-bis(2-aminophenoxy)ethane- N, N, N', N'-tetraacetic acid/AM did not reverse amitriptyline-induced cytotoxicity. Collectively, our data suggest that in MG63 cells, amitriptyline induced [Ca2+]i rises by evoking PLC-dependent Ca2+ release from the endoplasmic reticulum and Ca2+ entry via PKC-regulated store-operated Ca2+ entry. Amitriptyline also induced Ca2+-disassociated cell death.

Amitriptyline May Have Possibility to Induce Brugada Syndrome Rather than Long QT Syndrome.

Amitriptyline has been reported to induce long QT syndrome in addition to Brugada syndrome. We qualitatively and quantitatively analyzed the potential of amitriptyline to induce these lethal syndromes by using the halothane-anesthetized dogs (n = 6). Amitriptyline was intravenously administered in doses of 0.1, 1 and 10 mg/kg over 10 min every 20 min, which would provide approximately 1, 10 and 100 times higher plasma concentrations than a therapeutic one, respectively. The low dose hardly altered any of the cardiovascular variables. The middle dose increased the heart rate, cardiac output and left ventricular contractility, but decreased the total peripheral vascular resistance and left ventricular end-diastolic pressure, whereas it did not alter any of the electrocardiographic variables. The high dose decreased the mean blood pressure and left ventricular contractility; suppressed atrioventricular nodal and intraventricular conduction; shortened the repolarization period without altering the J-T peak c and T peak-T end; and prolonged the effective refractory period, providing post-repolarization refractoriness in addition to the enhancement of the middle dose-induced cardiovascular effects. Thus, amitriptyline at up to 100 times its therapeutic concentration may not be associated with the onset of long QT syndrome, but may induce Brugada syndrome.