Investigation of the effects of BRL37344 on TRPA1 and TRPC1 channels in experimental ichemia-reperfusion damaged rat brain tissue

Ischemic cerebrovascular disease is a serious neurological problem causing neurological deficit as a result of a reduction or blockage in blood flow to the brain tissue, also known as the leading cause of disability or death in the world. Although neuronal damage is commonly associated with several conditions including cytotoxic edema caused by the disruption of the Na⁺/K⁺-ATPase pump after ischemia, lactic acidosis due to anaerobic glycolysis, protease and phospholipase activation due to increased intracellular calcium, and the formation of free radicals due to phospholipase activation, the mechanism and pathogenesis of neuronal destruction has not been fully elucidated. The ion currents in cells mediated by non-selective cation channels are held responsible for neuronal cell death. The TRP family is considered to be upregulated in ischemic stroke and to play a role in Ca2+-mediated necrotic and apoptotic cell death. The aim of this study was to investigate the effects of BRL37344 on serum levels of TRPA1 and TRPC1 and immunoreactivity in rat brain tissue in an experimental ischemia-reperfusion (I/R) injury model. A total of 35 Sprague-Dawley adult male rats aged 8-12 weeks were randomly divided into five groups: (i) control, (ii) sham, (iii) BRL37344, (iv) I/R, and (v) I/R + BRL37344. No treatment was administered in the control group. A single dose of BRL37344 (5 mcg/kg) was administered to the BRL37344 group 120 minutes before decapitation. Surgery was performed in all rats in the ischemia-reperfusion and sham groups. At the end of the experiment, all the rats were decapitated using ketamine (75 mg/kg) + xylazine (10 mg/kg) i.p. Blood was drawn from the rats and brain tissues were rapidly removed after decapitation. Immunohistochemical staining was performed for TRPA1 and TRPC1 under light microscopy, and TUNEL staining was performed for apoptosis. Blood samples were centrifuged and serums were stored in Eppendorf tubes at -80 °C for ELISA examination (total antioxidant status [TAS], total oxidant status [TOS], TRPA1, and TRPC1) until the analysis. In biochemical examinations, no significant difference was found among the control, sham, and BRL37344 groups with regard to TAS, TOS, TRPA1, and TRPC1 levels. In the I/R group, there was a significant increase in the TOS, TRPA1, and TRPC1 levels and a significant decrease in the TAS level compared to the control group. Compared to the I/R group, the IR+BRL37344 group showed a significant decrease in the TOS, TRPA1, and TRPC1 levels and a significant increase in the TAS level. In immunohistochemical examination TRPA1 and TRPC1 under light microscopy, immunoreactivity was similar in the control, sham, and BRL37344 groups. In contrast, a significant increase in immunoreactivity was observed in the I/R group compared to the control and a significant decrease was observed in TRPA1 and TRPC1 immunoreactivity in the IR+BRL37344 group compared to the I/R group. The exact causes of neuronal damage after I/R injury remain unclear. Findings from the present study show that TRPA1 and TRPC1 cation channels may play a significant role in neuronal damage and that BRL37344 may have a protective effect on these channels. Key words: Brain, ischemia/reperfusion injury, TRPA1, TRPC1, BRL37344