Venlafaxine

Fluoxetine (N-methyl-3-[(4-trifluoromethyl) phenoxy]-3-phenylpropylamine hydrochloride), venlafaxine (1-[2-(dimethylamino)-1-(4-methoxyphenyl) ethyl] cyclohexanol hydrochloride) and amitriptyline (3-(10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5-ylidene)-N,N-dimethyl-1-propanamine hydrochloride) (Sigma-Aldrich) were prepared in ultrapure water to a concentration of 10 mM. Further dilutions were made in appropriate culture media as described in the text. These concentrations are similar to therapeutic serum concentrations of these drugs in humans. Treatment with drugs (or vehicle control [VEH]) was commenced at the beginning of each differentiation processes and maintained through all subsequent media changes. Representative light micrographs (magnification×10) were taken at week two and three of differentiation.

Drugs utilized were as follows: Vitamin-B6, 100 mg/kg (pyridoxine HCl, 100 mg/ml, Caspian Tamin Industry, Iran) (6); clomipramine, 10 mg/kg (Sigma-Aldrich, Germany); venlafaxine, 16 mg/kg (Sigma-Aldrich, Germany); fluoxetine HCl, 20 mg/kg (gift from Pars Daru, Iran) (10). All drugs were diluted by normal saline and injections were adjusted for 10 mL/kg mice body weight. All drugs were administered i.p. 30 min before testing. Vitamin B6 was administered daily for 3, 6 or 9 days in order to evaluate the best timing. Each experimental group comprised of 8 mice as follows: control group, vitamin B6 groups (3, 6 or 9 days), vitamin B6 plus clomipramine, fluoxetine, venlafaxine or vehicle groups.

Venlafaxine was kindly gifted from Mash premiere company, Ni (NO₃)₂·6H₂O and Co(NO₃)₂·6H₂O were obtained from Sigma Aldrich. Venlaxin® tablet (containing 75 mg VEN per tablet) and Venllamash® capsule (containing 75 mg VEN per capsule) were purchased from local market.
Graphene oxide (GO) was synthesized following the modified Hummer's method.^{38 (link)} Potassium ferrocyanide with purity of 98% was purchased from Alpha Chemika, India.
All chemicals were of analytical reagent grade and were used as received. Deionized water was used for the preparation of standard solutions and electrolytes.

Five drugs were tested in this study: duloxetine, celecoxib (Tokyo Chemical Industry Co., Ltd, Tokyo, Japan), pregabalin (ChemFaces, Hubei, China), venlafaxine, and tramadol (Sigma-Aldrich, St. Louis, MO, USA). All drugs were dissolved in distilled water (Otsuka Pharmaceutical Factory Inc, Tokushima, Japan) containing 0.5% w/v methylcellulose 400 solution (Fujifilm Wako Pure Chemical Corporation, Osaka, Japan) and administered orally (5 mL/kg) on Day 7 after SNL.

GBR12909 dihydrochloride was purchased from Tocris Bioscience (Bristol, UK). Venlafaxine and dopamine were obtained from Sigma-Aldrich Chemical Co (St. Louis, MO, USA). [³H]-DA and [³H]-5-HT were purchased from PerkinElmer Life Sciences (Waltham, MA, USA). [³H]-Sulpiride (75.7 Ci/mmol) was purchased from PerkinElmer Life Sciences. The synthetic 4-benzylpiperidine carboxamides used in this study have been described in a previous study (Paudel et al., 2015 (link)). Human embryonic kidney (HEK)-293 cells were obtained from the American Type Culture Collection (Manassas, VA, USA) and maintained in minimal essential medium containing 10% fetal bovine serum, 100 U/mL penicillin, and 100 μg/mL streptomycin.

The mixed-gender 10 donor pooled cryopreserved primary human hepatocytes with 5 million cells, hepatocyte thawing buffer, and incubation buffer were purchased from BIOIVT elevating science (Baltimore, MD). Gefitinib, losartan, SSRIs (citalopram, fluoxetine, fluvoxamine, paroxetine, sertraline, and venlafaxine), and CYP-specific substrates (phenacetin, mephenytoin, tolbutamide, dexamethorphan, nifedipine) were purchased from Sigma-Aldrich (St. Louis, MO). Recombinant pooled human supersomes for CYP1A2, CYP2C19, CYP2C9, CYP2D6, and CYP3A4, and the NADPH regeneration system solutions A and B were obtained from Corning Life and Sciences (Tewksbury, MA).