1- Department of Biology, Faculty of Basic Sciences, University of Zabol, Zabol, Iran.
2- Department of Chemistry, Faculty of Basic Sciences, University of Zabol, Zabol, Iran.
Abstract:
Introduction: Methamphetamine (MA) acts as a powerful oxidant agent, while Rosmarinic Acid (RA) is an effective herbal antioxidant. Oxidative stress-mediated by MA results in apoptosis, and caspase-3 is one of the critical enzymes in the apoptosis process. MA can epigenetically alter gene regulation. In this paper, to investigate the effects of RA on MA-mediated oxidative stress, changes in the level of casp3a mRNA were demonstrated in zebrafish.
Methods: The animals were grouped in 3 treatment conditions for the behavioral test: control, MA, MA pretreated by RA, and 6 treatment conditions for the molecular test: control, RA, MA, MA co-treated with RA, MA co-treated with RA/ZnO/chitosan nanoparticle, and ZnO/chitosan nanoparticle. Then molecular and behavioral investigations were carried out, and critical comparisons were made between the groups.
MA solution was prepared with a concentration of 25 mg/L, and RA solution was prepared by DPPH test with the antioxidant power of about 97%. Each solution was administered by immersing 20 zebrafish for 20 minutes, once per day for 7 days. The level of casp3a mRNA was quantified by using qRT-PCR. One-sided trapezoidal tank diving test was applied to study behavioral alterations.
Results: The qPCR analysis demonstrated the high potential of RA/ZnO/chitosan in counteracting the MA-mediated elevation in casp3a mRNA level. Based on the diving test results of MA-treated fish, MA was found to be anxiolytic compared to the control. While the resulted diving pattern of the MA-treated animals pretreated by RA was novel and different from both the control and MA-treated groups.
Conclusion: The potential of RA combined with a suitable nanoparticle against MA-induced oxidative stress was supported. The high efficiency of ZnO/chitosan in increasing RA penetration to the brain cells was evident. MA at a dose of 25 mg/L is anxiolytic for zebrafish. However, the molecular mechanisms involved in these processes should be studied.
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Highlights
● Methamphetamine (MA) induces excess casp3a transcript in the brain tissue in zebrafish.
● Immediate effect of chronic exposure to 25 mg/L MA is anxiolytic for zebrafish.
● Rosmarinic Acid (RA) counteracts the effects of MA-mediated oxidative stress.
● RA alone does not reduce the anxiolytic effects of MA and even enhances it.
● ZnO/chitosan nanoparticles increase RA penetration to the blood-brain barrier as fast as MA.
Plain Language Summary
Methamphetamine (MA) called ice or crystal in streets, is a drug of abuse. It has been associated with a variety of neurotoxic effects including oxidative stress, which leads to characteristic cell changes and death. The caspase-3 enzyme is one of the key enzymes of this process. There are storage forms of inactive Caspase-3 in any cell called procaspase-3. It’s been demonstrated previously that MA treatment causes activation of the procaspase-3 resource. The present study was undertaken to investigate whether MA can change the regulation of Caspase-3 gene expression and increase the synthesis of casp3a mRNA in zebrafish. qRT-PCR results demonstrated that MA increases the casp3a mRNA levels up to 18-fold compare to the control group. We were curious to know whether treatment by an antioxidant solution can counteract MA-mediated oxidative stress. So in the second part of this research, we investigated the herbal antioxidant molecule, Rosmarinic Acid (RA). Since the penetration of RA across the Blood-Brain Barrier is not as fast as MA, RA was combined with ZnO/chitosan nanoparticles and then provided to zebrafish as an animal model. The qRT-PCR analysis demonstrated the high potential of RA/ZnO/chitosan in counteracting the MA-mediated elevation in the casp3a mRNA level and brought it back much closer to the level of the control group (2.8-fold). On the other hand, zebrafish is known as a non-mammalian anxiety model system. The immediate effects of treatment of RA on MA-mediated psycho-behaviors were studied by applying the one-sided trapezoidal tank diving test. The diving test of MA-treated zebrafish resulted that the dose of 25 mg/l of MA was anxiolytic compared to the test of saline-treated zebrafish as the control. But the diving pattern of the zebrafish exposed to RA as well as MA was novel and varied from both the control and MA-treated groups. The diving pattern was not similar to any of them and even the stress and speed of diving were less than the MA-treated group.
Type of Study:
Original |
Subject:
Behavioral Neuroscience Received: 2020/04/10 | Accepted: 2020/12/1 | Published: 2021/03/1