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Membranes were then incubated for 30 min at RT in HRP-conjugated secondary antibody (Santa Cruz Biotechnology, Inc, USA) diluted in 5% milk or BSA in TBS-T

Brain tissue homogenates and serum samples were collected to quantify intestinal permeability markers (zonulin and occluding), oxidative-stress markers (GSH, MDA, CAT), and GABA-receptor gene expression using ELISA and biochemical assays

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Lastly, the effects of AST appeared to be mediated through the benzodiazepine and opioid receptors, which are known to play roles in the management of neuropathic pain (Figure 11)