Elucidating Pathway and Anesthetic Mechanism of Action of Clove Oil Nanoformulations in Fish

Abstract

Clove oil (CO), an essential oil of Syzygium aromaticum, has been reported as an anesthetic for many fish species. However, its insoluble properties require a suitable delivery system for its application. In the present study, nanoformulations of CO as a nanoemulsion (CO-NE), a self-microemulsifying drug-delivery system (CO-SMEDDS), and a self-nanoemulsifying drug-delivery system (CO-SNEDDS) were prepared for delivering CO. Zebrafish were used as a fish model to investigate oil pathways. The result shows fluorescence spots of fluorescence-labeled CO accumulate on the gills, skin, and brain. All CO nanoformulations significantly increased penetration flux compared to CO ethanolic solution. Investigation of the anesthetic mechanism of action using a rat brain γ-aminobutyric acid subtype A (GABAA) receptor-binding test demonstrates that CO and its major compound, eugenol, modulate [3H]muscimol binding. CO-NE exhibited a concentration-dependent binding activity with an EC50 value of 175 µg/mL, significantly higher than CO solution in dimethyl sulfoxide. In conclusion, CO enters the fish through the skin and gills. The anesthetic mechanism of action of CO is based on modulation of [3H] muscimol binding to GABAA receptors. Among three nanoformulations tested, CO-NE is the most effective at increasing permeability and enhancing the receptor-binding activity of the oil.

The effect of eugenol anesthesia on the electric organ discharge of the weakly electric fish Apteronotus leptorhynchus

“…A second mechanisms that may contribute to the effect of eugenol to induce a drop in EOD frequency is its ability to potentiate GABA A -receptor response. Such an effect has been observed in test systems based on GABA A receptors expressed in Xenopus oocytes (Aoshima and Hamamoto 1999 ; Sahin et al 2017 ) 1 and is consistent with the results obtained in a [ 3 H] muscimol-binding assay using rat cortical homogenate (Kheawfu et al 2022 ). Since GABA application to the SPPn lowers the frequency of the EOD, and thus the output frequency of the Pn (Metzner 1999 ), a potentiation of the response of GABA receptors received by input from the nE↓ would be expected to result in a decrease of the oscillation frequency of the Pn.…”

Efficacy and Physiological Responses of Payangka Fish (Ophieleotris aporos) to Anesthetization with Clove Oil

“…In general, clove oil was an efficient anesthetic at low concentrations (Fernandez et al, 2017). According to Kheawfu et al, (2022), the anesthetic mechanism of action of clove oil is based on the modulation of [ 3 H] muscimol binding to GABAA receptors located in the brain. Clove oil exerted a conspicuous depression of muscle contraction power (Fujimoto et al, 2018).…”

Clove Oil-Nanostructured Lipid Carriers: A Platform of Herbal Anesthetics in Whiteleg Shrimp (Penaeus vannamei)

“…In addition, anesthesia may inhibit neural and motor activity in the animals by blocking voltage-dependent K+ and Na+ channels [ 50 ]. Furthermore, the previous studies reported that eugenol and isoeugenol can activate the GABA A receptor in the central nervous system [ 30 , 51 , 52 ]. Therefore, we infer that the small particle and lipophilic nature of CO-NLCs have a tranquilizing effect on the brain and thereby potentiate the GABA A receptors’ response.…”

“…In this experiment, we investigated the biodistribution of fluorescently labeled clove oil in shrimp bodies visualized by bioluminescence imaging [ 30 ]. Shrimps were divided into three groups: control (untreated), STD-CO, and CO-NLCs.…”