Hesperidin Interacts With CREB-BDNF Signaling Pathway to Suppress Pentylenetetrazole-Induced Convulsions in Zebrafish

Abstract

Hesperidin (3,5,7-trihydroxyflavanone 7-rhamnoglucoside) is a β-7-rutinoside of hesperetin (4'-methoxy-3',5,7-trihydroxyflavanone), abundantly found in citrus fruits and known to interact with various cellular pathways to show a variety of pharmacological effects. The present study was envisaged to understand the anticonvulsant effect of hesperidin in a zebrafish model of pentylenetetrazole (PTZ)-induced convulsions, with the support of in silico docking. Healthy zebrafish larvae were preincubated with hesperidin (1, 5, and 10 µM) for 1 h, before PTZ exposure. Hesperidin treatment significantly increased the seizure latency and minimized PTZ-induced hyperactive responses. A significant reduction in c-fos expression further supported the suppression of neuronal excitation following hesperidin incubation in the larvae exposed to PTZ. The treatment also modulated larval bdnf expression and reduced the expression of il-10. The results of in vivo studies were further supported by in silico docking analysis, which showed the affinity of hesperidin for the N-methyl-d-aspartate receptor, the gamma-aminobutyric acid receptor, Interleukin 10 and the TrkB receptor of brain-derived neurotrophic factor. The results concluded that hesperidin suppresses PTZ-mediated seizure in zebrafish larvae through interaction with the central CREB-BDNF pathway.

Keywords: brain-derived neurotrophic factor; c-fos; clonic-like seizures; epilepsy; flavanone glycoside; il-10; in silico docking; zebrafish larva.

FIGURE 1

Docking results obtained from LeadIT for the four receptors with hesperidin. (A) 3D and 2D interaction diagram of Hesperidin with NMDAR receptor. (B) 3D and 2D interaction diagram of Hesperidin with TRK receptor. (C) 3D and 2D interaction diagram of Hesperidin with GABA receptor. (D) 3D and 2D interaction diagram of Hesperidin with Interleukin-10 receptor; (i) 3D pose, and (ii) 2D pose.

FIGURE 2

Snapshots of the complexes at 0 ns (start), 25 ns (midway), and 50 ns (end) of the simulation visualised in Discovery studio. (A) NMDAR-Hesperidin, (B) TRK-Hesperidin, (C) GABA-Hesperidin, and (D) Interleukin10-Hesperidin.

FIGURE 3

Effect of hesperidin on PTZ-induced hyperactivity in 7 dpf zebrafish larvae. (A) Mean swimming speed, (B) total distance travelled, and (C) larva movement track. ^a P < 0.05 as compared to veh/naive (naïve) group and; ^b P < 0.05 as compared to veh/ptz (vehicle control) group. The results expressed as mean ± SEM. veh/naive: system water incubated group not exposed to PTZ; veh/ptz: system water incubated group exposed to PTZ; hes1/ptz: Hesperidin 1 μM incubated group exposed to PTZ; hes5/ptz: Hesperidin 5 μM incubated group exposed to PTZ and; hes10/ptz: Hesperidin 10 μM incubated group exposed to PTZ.

FIGURE 4

Effect of hesperidin on the latency to PTZ-induced seizures (Stage 3). veh/naïve group was not exposed to PTZ, hence not shown in the figure. ^a P < 0.05 as compared to veh/ptz (vehicle control) group; ^b P < 0.05 as compared to hes1/ptz group and; ^c P < 0.05 as compared to hes5/ptz group. The results expressed as mean ± SEM. veh/ptz: system water incubated group exposed to PTZ; hes1/ptz: Hesperidin 1 μM incubated group exposed to PTZ; hes5/ptz: Hesperidin 5 μM incubated group exposed to PTZ and; hes10/ptz: Hesperidin 10 μM incubated group exposed to PTZ.

FIGURE 5

Effect of hesperidin on PTZ-mediated alterations in mRNA levels of (A) c-fos, (B) bdnf, and (C) il-10. ^a P < 0.05 as compared to veh/naive-s (naive) group and; ^b P < 0.05 as compared to veh/ptz-s (vehicle control). The results expressed as mean ± SD. veh/naïve-s: system water incubated group not exposed to PTZ; veh/ptz-s: system water incubated group exposed to PTZ; and; hes10/ptz-s: Hesperidin 10 μM incubated group exposed to PTZ.