Fig 1: Effects of selective deletion of AdipoR1 in 5-HT neurons on SERT and 5-HT immunoreactivity in mPFC and hippocampus and behavioral responses to fluoxetine and desipramine.a (Left panels) Representative immunoblots and quantitative analysis of SERT levels in the mPFC of male (upper) and female (lower) mice. Male mice: AdipoR1f/f, n = 6; AdipoR1f/f;ePet-Cre, n = 6. Female mice: AdipoR1f/f, n = 7. AdipoR1f/f;ePet-Cre, n = 7. (Right panels) Representative brain sections immunostained for 5-HT and quantitative analysis of the intensity of 5-HT-positive fibers in the mPFC of male (upper) and female (lower) mice. Male: 14 sections from three AdipoR1f/f mice; 14 sections from three AdipoR1f/f;ePet-Cre mice. Female: 14 sections from three AdipoR1f/f mice; 13 sections from three AdipoR1f/f;ePet-Cre mice. Scale bar = 50 µm. b (Left panels) Representative immunoblots and quantitative analysis of SERT levels in the hippocampus of male (upper) and female (lower) mice. Male mice: AdipoR1f/f, n = 6; AdipoR1f/f;ePet-Cre, n = 6. Female mice: AdipoR1f/f, n = 7. AdipoR1f/f;ePet-Cre, n = 7. (Right panels) Representative brain sections immunostained for 5-HT and quantitative analysis of the intensity of 5-HT-positive fibers in the hippocampus of both male (upper) and female (lower) mice. Male: 24 sections from three AdipoR1f/f mice; 23 sections from three AdipoR1f/f;ePet-Cre mice. Female: 21 sections from three AdipoR1f/f mice; 24 sections from three AdipoR1f/f;ePet-Cre mice. Scale bar = 100 µm for low magnification and 50 µm for high magnification. c Forced swim test performed 30 min after fluoxetine (5 or 10 mg kg-1, i.p.) or desipramine (10 mg kg-1, i.p.) injection in male (left) and female (right) mice. Male mice: AdipoR1f/f + saline, n = 7; AdipoR1f/f + fluoxetine (5 mg kg-1), n = 9; AdipoR1f/f + fluoxetine (10 mg kg-1), n = 7; AdipoR1f/f + desipramine, n = 9; AdipoR1f/f;ePet-Cre + saline, n = 7; AdipoR1f/f;ePet-Cre + fluoxetine (5 mg kg-1), n = 9; AdipoR1f/f;ePet-Cre + fluoxetine (10 mg kg-1), n = 8; AdipoR1f/f;ePet-Cre + desipramine, n = 7. Female mice: AdipoR1f/f + saline, n = 8; AdipoR1f/f + fluoxetine, n = 7; AdipoR1f/f + desipramine, n = 10; AdipoR1f/f;ePet-Cre + saline, n = 7; AdipoR1f/f;ePet-Cre + fluoxetine, n = 7; AdipoR1f/f;ePet-Cre + desipramine, n = 10. *P < 0.05, **P < 0.01, ***P < 0.001 compared with AdipoR1f/f littermate controls or saline-treated groups. d Schematic diagram illustrating the alterations in 5-HT system components and antidepressant responses induced by the loss of AdipoR1 in 5-HT neurons.
Fig 2: Summary of the effects of saffron on acute restraint stress (ARS)-induced neurobiological changes depending on the brain area. KMO: kynurenine 3-monooxygenase; KAT: kynurenine aminotransferase; 5-HT: serotonin; DA: dopamine; DRD1: Dopamine Receptor D1; SERT; serotonin transporter.
Fig 3: Effect of acute restraint stress (ARS) and oral pre-administration of Safr’Inside™ (6.25 mg/kg) on monoamine receptors, transporters and enzymes genes expression. Genes expression in (A) the Frontal Cortex (FCx); (B) Striatum (STR) and (C) Hippocampus (HPC). Data are represented as the foldchange calculated relative to the control group (baseline = 1). Results are shown as mean ± SEM. Control: n = 6–14; ARS: n = 9–15; Safr’Inside pre-ARS: n = 8–15. * p = 0.05, ** p = 0.01, *** p = 0.001 vs. Control. 5-HTR1a and b: Serotonin 1a and 1b Receptors; SERT: Serotonin transporter; MAO-A and B: Monoamine oxidase A and B; COMT: Catechol-O-methyltransferase; DRD1 and DRD2: Dopamine Receptors D1 and D2; DAT: Dopamine transporter.
Fig 4: Effect of acute restraint stress (ARS) and oral pre-administration of Safr’Inside™ (6.25 mg/kg) on protein expression of monoamine receptors and transporters. (A) Effect of Safr’Inside compared to ARS group on DA system proteins in the Frontal Cortex (FCx); (B) Effect of ARS compared to control group on DRD1 protein in the FCx; (C) Effect of Safr’Inside compared to ARS group on DA system proteins in the Striatum (STR); (D) Effect of ARS compared to control group on DRD1 proteins in the STR; (E) Effect of Safr’Inside compared to ARS group on 5-HT system proteins in the FCx; (F) Effect of Safr’Inside compared to ARS group on 5-HT system proteins in the Hippocampus (HPC). Results are shown as mean ± SEM. Control: n = 13–14; ARS: n = 11–14; Safr’Inside pre-ARS: n = 12–14. * p = 0.05. DRD1 and DRD2: Dopamine Receptors D1 and D2; DAT: Dopamine transporter; 5-HTR1a: Serotonin 1a Receptor; SERT: Serotonin transporter.
Fig 5: Effects of selective deletion of AdipoR1 in 5-HT neurons on expression levels of SERT, TPH2 and 5-HT in the dorsal raphe nucleus.a Immunoblots showing SERT and TPH2 protein levels in male (left) and female (right) AdipoR1f/f and AdipoR1f/f;ePet-Cre mice. Male mice: AdipoR1f/f, n = 6; AdipoR1f/f;ePet-Cre, n = 6. Female mice: AdipoR1f/f, n = 6; AdipoR1f/f;ePet-Cre, n = 6. b (Left panels) Representative immunohistochemical staining showing SERT, TPH2, and 5-HT immunofluorescence in the dorsal raphe nucleus of male AdipoR1f/f and AdipoR1f/f;ePet-Cre mice. (Middle panels) Relative fluorescence intensity normalized by control mice. (Right panels) The average number of immunopositive neurons per section in the dorsal raphe nucleus. SERT: 12 sections from three AdipoR1f/f mice; 12 sections from three AdipoR1f/f;ePet-Cre mice. TPH2: 11 sections from three AdipoR1f/f mice; 11 sections from three AdipoR1f/f;ePet-Cre mice. 5-HT: 12 sections from three AdipoR1f/f mice; 12 sections from three AdipoR1f/f;ePet-Cre mice. Scale bar = 100 µm. *P < 0.05, **P < 0.01 compared with AdipoR1f/f littermate controls.
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