ANTI-STRESS POTENTIALS AND THE MECHANISM OF ACTION OF MORIN HYDRATE IN SWISS MICE

Abstract

Stress, is an integral component of life that distorts body homeostasis. The available anti-stress drugs are often ineffective in combating its multiple effects. Adaptogens with antioxidant and neuroprotective effects are known to relieve stress. Morin hydrate (MH), a flavonoid from Morus alba with known antioxidant and neuroprotective properties has not been investigated for its anti-stress potential. The study was designed to investigate the anti-stress property of MH and its mechanisms of action in mice. Eighty male Swiss mice (22.0 ± 2.5 g) were used for acute studies: Swimming Endurance Test (SET), Anoxic Tolerance Test (ATT) and Acute Restraint Stress (ARS). The SET and ATT consisted of 5 treatment groups each (n = 5): vehicle (normal saline, 10 mg/mL), MH (5, 10, 20 mg/kg) and adaptogen (ginseng, 25 mg/kg), administered intrapertioneally. Thirty minutes later immobility time was measured in SET and convulsion latency in ATT. In ARS, thirty male mice were allotted into treatment groups I-VI (n = 5): vehicle(10 mg/mL), vehicle-stress control (10 mg/mL), MH (5, 10, 20 mg/kg) and ginseng (25 mg/kg), and treated for 7 days prior to being restrained except group 1. Thereafter, mice were assessed for anxiety and depression in Elevated-Plus Maze (EPM) and Forced-Swim Test (FST), respectively. For chronic studies, ninety male mice were used in 3 models [Chronic- Restraint Stress (CRS), Paradoxical Sleep-deprivation (PSD) and Chronic-Unpredictable Stress (CUS) and grouped, respectively as in ARS. In CRS, mice were pre-treated and restrained for 14 days, and thereafter assessed for anxiety and depression in Mice were sleep-deprived for 48 hours in PSD, and exposed to stressors for 14 days in CUS before testing for memory and anxiety behaviours using Y-maze and EPM, respectively. Brain glutathione (GSH), malondialdehyde, nitric-oxide and blood glucose were determined spectrophotometrically in ARS, CRS, PSD and CUS. Serum corticosterone, brain tumor necrosis factor-alpha (TNF-α) and interleukin-l beta were measured in CUS using ELISA. Brain nuclear factor-kB (NF-kB) and inducible nitric-oxide synthase (iNOS) expressions in CUS were measured using immunohistochemistry. Data were analysed using ANOVA at α0.05. Morin hydrate (5, 10, 20 mg/kg) significantly reduced immobility (10.8 ± 0.20, 7.04 ± 0.77, 9.16± 0.59 s against 11.9 ± 0.25 s) in SET and prolonged convulsion latency ATT (33.1 ± 1.26, 34.1 ±2.40, 34.8 ± 1.06 s against 21.9 ± 1.15 s).The MH decreased anxiety, depression-like symptoms, malondialdehyde and nitric-oxide but increased GSH in ARS and CRS. The MH reversed memory impairment (65.87 ± 3.59, 69.17 ± 6.51, I 62.0± 5.12% against 50.87 ± 2.87%) and anxiety (64.75 ± 5.36, 57.75 ± 2.95, 61.00 ± 1.68 s against 45.0 ± 1.87 s) in PSD. Also, MH increased GSH (104.6 ± 8.50, 97.3 ±6.51, 91.5 ± 7.70 µmol/g tissue against 50.22 ±1.41 µmol/g tissue) but decreased malondialdehyde (7.57± 0.25, 4.50 ± 0.13, 3.16 ± 0.22 µmol/g tissue against 8.60 ± 0.14 µmol/g tissue) and nitric-oxide (163.0 ± 8.67, 121.3 ± 6.67, 124.7± 3.33 µmol/g tissue against 338.0 ± 16.77 µmol/g tissue). The MH increased GSH concentration in CUS and ameliorated CUS-induced increases in glucose, malondialdehyde and nitric oxide Ievels compared to controls. Morin hydrate reduced corticosterone (7.93 ± 0.19, 7.18 ±0.21, 7.46 ± 0.20 ng/mL. against 8.54 ±0.14 ng/mL), TNF-α (49.19 ± 0.55, 47.60± 2.48, 35.22 ± I .77 pg/mL against 92.37± 7.90 pg/mL), and interleukin-I beta (74.45 ± 2.18. 46.45 ± 2.71 43.12 ± 1.55 pg/mL, against 98.72 ± 4.03 pg/mL). Morin hydrate reduced iNOS and NF-kB protein levels in CUS. Morin hydrate exhibited anti-stress potential via mechanisms related to inhibition of hypothalamic-pitutary-adrenal axis hyperactivation, oxidative stress and neuroinflammation.