Organochalcogens inhibit mitochondrial complexes I and II in rat brain: possible implications for neurotoxicity

Abstract

Organochalcogens, such as organoselenium and organotellurium compounds, can be neurotoxic to rodents. Since mitochondrial dysfunction plays a pivotal role in neurological disorders, the present study was designed to test the hypothesis that rat brain mitochondrial complexes (I, II, I-III, II-III and IV) could be molecular targets of organochalcogens. The results show that organochalcogens caused statistically significant inhibition of mitochondrial complex I activity, which was prevented by preincubation with NADH and fully blunted by reduced glutathione (GSH). Mitochondrial complex II activity remained unchanged in response to (PhSe)₂ treatment. Ebs and (PhTe)₂ caused a significant concentration-dependent inhibition of complex II that was also blunted by GSH. Mitochondrial complex IV activity was not modified by organochalcogens. Collectively, Ebs, (PhSe)₂ and (PhTe)₂ were more effective inhibitors of brain mitochondrial complex I than of complex II, whereas they did not affect complex IV. These observations are consistent with organochalcogens inducing mitochondrial complex I and II inhibition via their thiol-oxidase-like activity, with Ebs, (PhSe)₂ and (PhTe)₂ effectively oxidising critical thiol groups of these complexes.

Figure 1. Effect of organochalchogens on mitochondrial complex I activity

Brain mitochondria membranes were incubated (0.5 mg/ml) for 10 min at 30°C in a medium containing 100 mM phosphate buffer, pH 7.4, in the presence of the indicated concentrations of organochalcogens (Ebs 0–50 µM; [(PhSe)₂] 0–100 µM; [(PhTe)₂] 0–100 µM). Complex I activity is expressed as % of control (absent organochalcogens). — ▲— (PhSe)₂, —●— (PhTe)₂, —■— Ebs, —+— Rotenone (100µM). The control value (in the absence of organochalcogens) was 101.78 ±2.31. Data are expressed as means ± SEM (n=4). *p<0.05 from respective control by Duncan’s multiple range test. ^# p<0.05 from control by t test.

Figure 2. Effect of organochalchogens on mitochondrial complex I–III activity

The mitochondria membranes were incubated (0.5mg/ml) for 10 min at 30°C in a medium containing 100 mM TFK buffer, pH 7.4, in the presence of the indicated concentrations of organochalchogens. Complex I–III activity is expressed as % of control (in the absence of organochalchogens). —▲— (PhSe)₂, —●— (PhTe)₂, —■— Ebs. The control value (in the absence of organochalcogens) was 100.94 ± 1.92. Data are expressed as means ± SEM (n=5). *p<0.05 from respective control by Duncan’s multiple range test.

Figure 3. Effect of organochalchogens on mitochondrial complex I–III activity under different assay conditions

Mitochondrial membranes were incubated (0.5mg/ml) for 10 min at 30°C in a medium containing 100 mM TFK buffer, pH 7.4, in the presence of the indicated concentrations of organochalchogens in the presence (white bars) or absence (gray bars) of NADH (200µM). Complex I–III activity is expressed as % of control (a condition without organochalchogens addition). Control (in the absence of organocompounds); (PhSe)₂ (5µM); (PhTe)₂ (5µM); Ebs (5µM); Rotenone (100µM). Data are expressed as means ± SEM (n=5). *p<0.05 from respective control by t test. ^#p<0.05 condition 1 vs condition 2 by t test

Figure 4. Effect of organochalchogens on mitochondrial complex II activity

Mitochondria membranes were incubated (0.5mg/ml) for 10 min at 30°C in a medium containing 100 mM TFK buffer, pH 7.4, in the presence of the indicated concentrations of organochalchogens under two different conditions: Condition 1 (with succinate 5mM); Condition 2 (without succinate). Values of complex II activity are expressed as % of control (in the absence of organochalchogens). (A) Brain mitochondrial membranes under condition 1 (Control value 101.05 ± 1.81); (B) Brain mitochondrial membranes under condition 2 (Control value 99.88 ± 2.05). —▲— (PhSe)₂, —●— (PhTe)₂, —■— Ebs, —+— Malonate (8mM). Data are expressed as means ± SEM (n=5). *p<0.05 from respective control by Duncan’s multiple range test. ^#p<0.05 from control by t test.

Figure 4. Effect of organochalchogens on mitochondrial complex II activity

Mitochondria membranes were incubated (0.5mg/ml) for 10 min at 30°C in a medium containing 100 mM TFK buffer, pH 7.4, in the presence of the indicated concentrations of organochalchogens under two different conditions: Condition 1 (with succinate 5mM); Condition 2 (without succinate). Values of complex II activity are expressed as % of control (in the absence of organochalchogens). (A) Brain mitochondrial membranes under condition 1 (Control value 101.05 ± 1.81); (B) Brain mitochondrial membranes under condition 2 (Control value 99.88 ± 2.05). —▲— (PhSe)₂, —●— (PhTe)₂, —■— Ebs, —+— Malonate (8mM). Data are expressed as means ± SEM (n=5). *p<0.05 from respective control by Duncan’s multiple range test. ^#p<0.05 from control by t test.

Figure 5. Effect of organochalchogens on mitochondrial complex II–III activity

Mitochondria membranes were incubated (0.5mg/ml) for 10 min at 30°C in a medium containing 100 mM TFK buffer, pH 7.4, in the presence of the indicated concentrations of organochalchogens. Complex II–III activity is expressed as % of control (in the absence of organochalchogens). —▲— (PhSe)₂, —●— (PhTe)₂, —■— Ebs. The control value (in the absence of organochalcogens) was 101.91 ± 1.81. Data are expressed as means ± SEM (n=5). *p<0.05 from respective control by Duncan’s multiple range test.

Figure 6. Effect of organochalchogens on mitochondrial complex IV activity

The mitochondria membranes were incubated (0.5mg/ml) for 10min at 30°C in a medium containing 100 mM TFK buffer, pH 7.4, in the presence of indicated concentrations organochalchogens. Complex IV activity is expressed as % of control (in the absence of organochalchogens). —▲— (PhSe)₂, —●— (PhTe)₂, —■— Ebs, —◆— KCN (10mM). The control value (in the absence of organochalcogens) was 102.81 ± 2.07. Data are expressed as means ± SEM (n=5). *p<0.05 from respective control by Duncan’s multiple range test. ^#p<0.05 from control by t test.