Figure 9
From: Levetiracetam mitigates doxorubicin-induced DNA and synaptic damage in neurons
Levetiracetam reduces DNA, synaptic and neurite damage caused by Dox.
(A) An example of cortical cultures pre-treated with 5 μM Lev and then Dox (0.01 μM, overnight), fixed and stained with antibodies against MAP2c. Scale bar is 20 μm. (B) Cortical neurons at 28–32 DIV were pre-treated with 5 μM Lev and then a vehicle or with Dox (0.01 μM for 3 days, or 0.1 μM, overnight) was added. Neurons were fixed and stained for MAP2c, synapsin and with the Hoechst dye and imaged. Note that red MAP2c staining also includes Dox’s red fluorescence. (C) Synaptically developed primary cortical cultures at 28−32 DIV were pre-treated with 5 mM Lev and then a vehicle or with Dox (0.01 μM for 3 days, or 0.1 μM, overnight) was added. Neurons were then fixed and stained with antibodies against γH2A.X and MAP2c and with the Hoechst dye. Blue staining was used by the algorithm to identify and analyze γH2A.X. (D) Quantification of fluorescent images of cortical cultures pre-treated with 5 mM Lev and then a vehicle or with Dox (0.01 μM for 3 days, or 0.1 μM, overnight). Neurons were fixed and stained with antibodies against BRCA1 and MAP2c and with the Hoechst dye. Blue staining was used by the algorithm to identify and analyze BRCA1. ***p<0.001 (Dunnett’s test). (E) Quantification of fluorescent images of cortical cultures pre-treated with 5 mM Lev and then a vehicle or with Dox (0.01 μM for 3 days, or 0.1 μM, overnight). Neurons were fixed and stained with antibodies against synapsin and MAP2c and with the Hoechst dye. MAP2c staining was used by the algorithm to identify and analyze synapses. ***p<0.001 (Dunnett’s test). (F) Quantification of fluorescent images of cortical cultures pre-treated with 5 mM Lev and then a vehicle or with Dox (0.01 μM for 3 days, or 0.1 μM, overnight). Neurons were fixed and stained with antibodies against MAP2c and with the Hoechst dye. MAP2c staining was used by the algorithm to identify and analyze neurites. ***p<0.001 (Dunnett’s test). Several thousand neurons were analyzed. Results were pooled from at least three independent experiments.
