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(A) Schematic connecting <t>neuronal</t> activity and CREB-mediated Eif4a3/Fus mRNA transcription for the subsequent circHomer1 biogenesis. (B-D) Relative to vehicle, circHomer1, Eif4a3 and Fus mRNA levels (quantifications were based on qRT-PCR and normalized to 18S rRNA) after treatment with CREB inhibitor (B-C) in two rounds of experiment at (B) dose < IC50 and (C) dose equal to IC50 in <t>mouse</t> primary <t>cortical</t> neurons (treatment was done for 24hours) and (D) after treatment with a CREB-CBP inhibitor at a dose equal to IC50. Mean ± SEM. ****p < 0.0001, ***p < 0.001, **p < 0.01, *p < 0.05, based on two tailed one sample t-test relative to vehicle. For circHomer1 after treatment with < IC50 dose (B): t = 3.106, df = 2, and p = 0.0899. For Eif4a3 mRNA after treatment with < IC50 dose (B): t = 3.441, df = 3, and p = 0.0412. For Fus mRNA after treatment with < IC50 dose (B): t = 10.53, df = 2, and p = 0.0089. For circHomer1 after treatment with = IC50 dose (C): t = 12.53, df = 4, and p = 0.0002. For Eif4a3 mRNA after treatment with = IC50 dose (C): t = 7.358, df = 4, and p = 0.0018. For Fus mRNA after treatment with = IC50 dose (C): t = 20.86, df = 4, and p < 0.0001. For circHomer1 after treatment with CREB-CBP inhibitor (D): t = 10.29, df = 4, and p = 0.0005. For Eif4a3 mRNA after treatment with CREB-CBP inhibitor (D): t = 8.238, df = 5, and p = 0.0004. For Fus mRNA after treatment with CREB-CBP inhibitor (D): t = 15.60, df = 5, and p < 0.0001. In all bar graphs the individual replicates are shown within the graph.
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(A) Schematic connecting <t>neuronal</t> activity and CREB-mediated Eif4a3/Fus mRNA transcription for the subsequent circHomer1 biogenesis. (B-D) Relative to vehicle, circHomer1, Eif4a3 and Fus mRNA levels (quantifications were based on qRT-PCR and normalized to 18S rRNA) after treatment with CREB inhibitor (B-C) in two rounds of experiment at (B) dose < IC50 and (C) dose equal to IC50 in <t>mouse</t> primary <t>cortical</t> neurons (treatment was done for 24hours) and (D) after treatment with a CREB-CBP inhibitor at a dose equal to IC50. Mean ± SEM. ****p < 0.0001, ***p < 0.001, **p < 0.01, *p < 0.05, based on two tailed one sample t-test relative to vehicle. For circHomer1 after treatment with < IC50 dose (B): t = 3.106, df = 2, and p = 0.0899. For Eif4a3 mRNA after treatment with < IC50 dose (B): t = 3.441, df = 3, and p = 0.0412. For Fus mRNA after treatment with < IC50 dose (B): t = 10.53, df = 2, and p = 0.0089. For circHomer1 after treatment with = IC50 dose (C): t = 12.53, df = 4, and p = 0.0002. For Eif4a3 mRNA after treatment with = IC50 dose (C): t = 7.358, df = 4, and p = 0.0018. For Fus mRNA after treatment with = IC50 dose (C): t = 20.86, df = 4, and p < 0.0001. For circHomer1 after treatment with CREB-CBP inhibitor (D): t = 10.29, df = 4, and p = 0.0005. For Eif4a3 mRNA after treatment with CREB-CBP inhibitor (D): t = 8.238, df = 5, and p = 0.0004. For Fus mRNA after treatment with CREB-CBP inhibitor (D): t = 15.60, df = 5, and p < 0.0001. In all bar graphs the individual replicates are shown within the graph.
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(A) Schematic connecting neuronal activity and CREB-mediated Eif4a3/Fus mRNA transcription for the subsequent circHomer1 biogenesis. (B-D) Relative to vehicle, circHomer1, Eif4a3 and Fus mRNA levels (quantifications were based on qRT-PCR and normalized to 18S rRNA) after treatment with CREB inhibitor (B-C) in two rounds of experiment at (B) dose < IC50 and (C) dose equal to IC50 in mouse primary cortical neurons (treatment was done for 24hours) and (D) after treatment with a CREB-CBP inhibitor at a dose equal to IC50. Mean ± SEM. ****p < 0.0001, ***p < 0.001, **p < 0.01, *p < 0.05, based on two tailed one sample t-test relative to vehicle. For circHomer1 after treatment with < IC50 dose (B): t = 3.106, df = 2, and p = 0.0899. For Eif4a3 mRNA after treatment with < IC50 dose (B): t = 3.441, df = 3, and p = 0.0412. For Fus mRNA after treatment with < IC50 dose (B): t = 10.53, df = 2, and p = 0.0089. For circHomer1 after treatment with = IC50 dose (C): t = 12.53, df = 4, and p = 0.0002. For Eif4a3 mRNA after treatment with = IC50 dose (C): t = 7.358, df = 4, and p = 0.0018. For Fus mRNA after treatment with = IC50 dose (C): t = 20.86, df = 4, and p < 0.0001. For circHomer1 after treatment with CREB-CBP inhibitor (D): t = 10.29, df = 4, and p = 0.0005. For Eif4a3 mRNA after treatment with CREB-CBP inhibitor (D): t = 8.238, df = 5, and p = 0.0004. For Fus mRNA after treatment with CREB-CBP inhibitor (D): t = 15.60, df = 5, and p < 0.0001. In all bar graphs the individual replicates are shown within the graph.

Journal: Research Square

Article Title: Regulation of neuronal circHomer1 biogenesis by PKA/CREB/ERK-mediated pathways and effects of glutamate and dopamine receptor blockade

doi: 10.21203/rs.3.rs-3547375/v1

Figure Lengend Snippet: (A) Schematic connecting neuronal activity and CREB-mediated Eif4a3/Fus mRNA transcription for the subsequent circHomer1 biogenesis. (B-D) Relative to vehicle, circHomer1, Eif4a3 and Fus mRNA levels (quantifications were based on qRT-PCR and normalized to 18S rRNA) after treatment with CREB inhibitor (B-C) in two rounds of experiment at (B) dose < IC50 and (C) dose equal to IC50 in mouse primary cortical neurons (treatment was done for 24hours) and (D) after treatment with a CREB-CBP inhibitor at a dose equal to IC50. Mean ± SEM. ****p < 0.0001, ***p < 0.001, **p < 0.01, *p < 0.05, based on two tailed one sample t-test relative to vehicle. For circHomer1 after treatment with < IC50 dose (B): t = 3.106, df = 2, and p = 0.0899. For Eif4a3 mRNA after treatment with < IC50 dose (B): t = 3.441, df = 3, and p = 0.0412. For Fus mRNA after treatment with < IC50 dose (B): t = 10.53, df = 2, and p = 0.0089. For circHomer1 after treatment with = IC50 dose (C): t = 12.53, df = 4, and p = 0.0002. For Eif4a3 mRNA after treatment with = IC50 dose (C): t = 7.358, df = 4, and p = 0.0018. For Fus mRNA after treatment with = IC50 dose (C): t = 20.86, df = 4, and p < 0.0001. For circHomer1 after treatment with CREB-CBP inhibitor (D): t = 10.29, df = 4, and p = 0.0005. For Eif4a3 mRNA after treatment with CREB-CBP inhibitor (D): t = 8.238, df = 5, and p = 0.0004. For Fus mRNA after treatment with CREB-CBP inhibitor (D): t = 15.60, df = 5, and p < 0.0001. In all bar graphs the individual replicates are shown within the graph.

Article Snippet: Mouse cortical neuronal cultures were purchased from ThermoFisher Scientific – Catalog number: A15585 (C57BL/6 mice).

Techniques: Activity Assay, Quantitative RT-PCR, Two Tailed Test