Reboxetine

Manufactured by Bio-Techne

Sourced in United Kingdom, Spain

Reboxetine is a lab equipment product manufactured by Bio-Techne. It functions as a selective norepinephrine reuptake inhibitor (NRI).

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Close spelling variants of this product

Reboxetine mesylate

5 protocols using reboxetine

Primary Cultures of Midbrain Dopaminergic Neurons

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Primary cultures and their survival were established and investigated as described previously^{15 (link)}. E13.5 embryos of C57Bl/6J wild type mice (6 months old) were dissected according to Planket et al.^{16 (link)}. 30 000 live cells were plated on 96-well plate pre-coated with poly-L-ornithine in culture medium (Neurobasal Medium + B27 supplement 1x + 2 mM L-Glutamine) and placed in the incubator. After 45 minutes required for cells attached, 100 µl of medium containing vehicle or investigated drugs were added. Drugs being the subject of investigation: GDNF growth factor – a positive control (100 ng/µl; PeproTech), reboxetine (10 µM; Tocris, #1982) and phenylephrine (100 µM; Sigma, P6126). The optimal dosage of investigated pharmaceuticals was chosen upon own experience and available literature¹⁷. 48 hours after plating half of the medium was exchanged with fresh one containing the respective drug concentrations. 5 days after plating cells were fixed with 4% PFA, immunostained with anti-TH antibody (Millipore, AB1542, 1:2000) and counterstained with DAPI. The plates where imaged by an inverted fluorescent microscope (AxioObserver, Carl Zeiss, Germany) at 5x magnification and the number of surviving TH+ cells in each separate well were counted automatically in an unbiased way with use of the ImageJ software^{18 (link)}.

Kreiner G., Rafa-Zabłocka K., Barut J., Chmielarz P., Kot M., Bagińska M., Parlato R., Daniel W.A, & Nalepa I. (2019). Stimulation of noradrenergic transmission by reboxetine is beneficial for a mouse model of progressive parkinsonism. Scientific Reports, 9, 5262.

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Pharmacological Agents for Behavioral Study

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S,R(±)-MDMA (MDMA), M100907 (M100), RTI-336 were supplied by the National Institute on Drug Abuse (Research Technology Branch, Research Triangle Park, NC, USA). Citalopram was obtained from Eli Lilly and Co. (Indianapolis, IN, USA). Reboxetine was obtained from Tocris Bioscience (Avonmouth, Bristol, UK). (-)-1-(2,5-Dimethoxy-4-iodophenyl)-2-aminopropane hydrochloride (DOI) was obtained from Sigma-Aldrich (St. Louis, MO, USA). MDMA, citalopram, Reboxetine and DOI were dissolved in 0.9% saline immediately before experimentation. RTI-336 was initially dissolved in 100% ethanol and diluted to less than a 2% concentration with saline. M100 was dissolved in saline and 0.1N HCl. Drug vehicle served as control for each respective experiment with a given drug. All solutions were injected intraperitoneally (i.p.) at a volume of 10 μL/g body weight. MDMA was administered 30 min prior to the behavioral procedure based on a previous study, and transporter inhibitors were administered 30 min before MDMA because all three drugs reach peak brain concentrations within 30 min of administration and remain elevated for at least 90 min (Kimmel et al. 2008 ; Kreilgaard et al. 2008 (link); Strolin Benedetti et al. 1995 (link)).

Young M.B., Norrholm S.D., Khoury L.M., Jovanovic T., Rauch S.A., Reiff C.M., Dunlop B.W., Rothbaum B.O, & Howell L.L. (2017). Inhibition of serotonin transporters disrupts the enhancement of fear memory extinction by 3,4-methylenedioxymethamphetamine (MDMA). Psychopharmacology, 234(19), 2883-2895.

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Evaluation of Antidepressant Compounds

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Fluoxetine (#H6995, Bosche Scientific), citalopram (#C505000, Toronto Research Chemicals), paroxetine (#2141, Tocris), fluvoxamine (#1033), venlafaxine (#2917, Tocris), reboxetine (#1982, Tocris), imipramine (#I7379-5G, Sigma–Aldrich), clomipramine (#C7291, Sigma–Aldrich), desipramine (#3067, Tocris), phenelzine (#P6777, Sigma–Aldrich), rolipram (#R6520, Sigma–Aldrich), ketamine (#3131, Tocris), 2R, 6R-Hydroxynorketamine and 2S, 6S-Hydroxynorketamine (#6094 and #6095, respectively, Tocris), carbamazepine (#4098, Tocris), chlorpromazine (#C8138, Sigma–Aldrich), flupenthixol (#4057, Tocris), and pimozide (#0937, Tocris) were investigated in this study. The compounds were dissolved in dimethyl sulfoxide (DMSO) that was also used as a vehicle in all the experiments. Concentrations tested (Table 1) were selected based on earlier in vitro studies from our research group (Fred et al., 2019 (link); Casarotto et al., 2021 (link)) and other laboratories (Johansen et al., 2015 (link)).

Fred S.M., Kuivanen S., Ugurlu H., Casarotto P.C., Levanov L., Saksela K., Vapalahti O, & Castrén E. (2022). Antidepressant and Antipsychotic Drugs Reduce Viral Infection by SARS-CoV-2 and Fluoxetine Shows Antiviral Activity Against the Novel Variants in vitro. Frontiers in Pharmacology, 12, 755600.

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Neurotransmitter Release during Differentiation

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To detect neurotransmitter release into the media during neuronal differentiation, we collected supernatant at indicated timepoints along differentiation. The cells were treated with drugs (maprotiline 25 µM (Tocris, 0935), tomoxetine 10 µM (Tocris, 2011), nisoxetine 10 µM (Tocris, 1025) and reboxetine 10 µM (Tocris, 1982)) for 4 h or KCl (40 mM) for 30 min before collecting the medium. ELISA was performed by using the Dopamine & Noradrenaline Sensitive ELISA Assay Kit (Eagle Biosciences, BCU39-K02) following the manufacturerʼs instructions.

Tao Y., Li X., Dong Q., Kong L., Petersen A.J., Yan Y., Xu K., Zima S., Li Y., Schmidt D.K., Ayala M., Mathivanan S., Sousa A.M.M., Chang Q, & Zhang S.C. (2023). Generation of locus coeruleus norepinephrine neurons from human pluripotent stem cells. Nature biotechnology.

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Microdialysis Procedure for Neurotransmitter Assay

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All reagents used were of analytical grade and were obtained from Merck (Germany). 5-HT oxalate, NE bitartrate, (±)-8-hydroxi-2(dipropylamino)tetralin hydrobromide (8-OH-DPAT), and clonidine were from Sigma-Aldrich-RBI (Madrid, Spain). Fluoxetine, reboxetine, citalopram hydrobromide and desipramine were from Tocris (Madrid, Spain). To assess the local effects in microdialysis procedures, drugs were dissolved in artificial cerebrospinal fluid (aCSF) and were administered by reverse dialysis at the stated concentrations [9, 10]. All other drugs dissolved in saline or aCSF as required. Concentrated solutions (1mM; pH adjusted to 6.5-7 with NaHCO3 when necessary) were stored at -80ºC and working solutions were prepared daily by dilution in aCSF.

Fullana M.N., Ferrés-Coy A., Ortega J.E., Ruiz-Bronchal E., Paz V., Meana J.J., Artigas F, & Bortolozzi A. (2019). Selective Knockdown of TASK3 Potassium Channel in Monoamine Neurons: a New Therapeutic Approach for Depression. Molecular neurobiology, 56(4).

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