Fig 2: Sulpiride, pure D2R antagonist, robustly upregulates circHomer1, Eif4a3 and Fus in brain regions that dopaminergic receptors are abundant.(A-D) Relative to vehicle circHomer1, Eif4a3 and Fus mRNA levels (based on qRT-PCR and normalized to 18S rRNA) after treatment with Sulpiride in (A) Putamen - dorsal striatum, (B) Nucleus Accumbens – ventral striatum, (C) Orbitofrontal cortex and (D) Cerebellum. Mean ± SEM. Mice were sacrificed 24hrs after the last injection – delayed time point to clarify if the possible change of circHomer1 levels after pharmacological intervention is persistent over time. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001, two tailed one sample t-test relative to vehicle. For circHomer1 in (A): p = 0.0390, t = 2.375, df = 10. For Eif4a3 in (A): p < 0.0001, t = 10.34, df = 10. For Fus in (A): p = 0.0008, t = 4.914, df = 9. For circHomer1 in (B): p = 0.0199, t = 2.720, df = 11. For Eif4a3 in (B): p < 0.0001, t = 10.63, df = 7. For Fus in (B): p = 0.0158, t = 3.051, df = 8. For circHomer1 in (C): p = 0.0014, t = 4.358, df = 9. For Eif4a3 in (C): p < 0.0001, t = 7.270, df = 9. For Fus in (C): p < 0.0001, t = 7.151, df = 9. In all bar graphs the individual replicates are shown within the graph.

Fig 3: MAPK cascade, PKA and the glutamatergic system can modulate circHomer1 and its upstream regulators (primary cortical neurons and mouse OFC)(A) Experimental Design – Primary mouse cortical neurons were plated in multiple laminin-coated 24-well plates, and on DIV13 (mature), potent and selective small molecule kinase inhibitors were added – treatments were done for 24hours. (B-E) Relative to vehicle, circHomer1, Eif4a3 and Fus mRNA levels (based on qRT-PCR and normalized to 18S rRNA) after treatment with (B) ERK inhibitor (C) MEK1,2 inhibitor – kinase upstream of ERK (D) PKA inhibitor and (E) PKC inhibitor. Mean ± SEM. *p < 0.05, **p < 0.01, ****p < 0.0001, based on two tailed one sample t-test relative to vehicle. For circHomer1 in (B): p < 0.0001, t = 107.4, df = 3. For Eif4a3 in (B): p < 0.0001, t = 36.50, df = 5. For Fusin (B): p < 0.0001, t = 16.23, df = 5. For circHomer1 in (C): p = 0.0083, t = 4.226, df = 5. For Eif4a3 in (C): p = 0.0001, t = 8.624, df = 4 For Fus in (C): p = 0.0034, t = 8.539, df = 3. For circHomer1in (D): p = 0.0016, t = 6.220, df = 5. For Eif4a3in (D): p = 0.0091, t = 4.124, df = 5. For Fusin (D): p = 0.0027, t = 5.521, df = 5. In all bar graphs the individual replicates are shown within the graph. (F-H) Relative to vehicle circHomer1, Eif4a3 and Fus mRNA levels (based on qRT-PCR and normalized to 18S rRNA) after treatment with (F) CHPG – mGluR5 agonist (treatment for 2–12-24hours), (G) MK801 – NMDAR antagonist (treatment for 2–12-24hours) and (H) ANA12 – TrkB antagonist (treatment for 24hours). Mean ± SEM. *p < 0.05, **p < 0.01, ****p < 0.0001, based on two tailed one sample t-test relative to vehicle. For circHomer1in (F) after 24 hours of treatment: p = 0.0040, t = 5.038, df = 5. For Eif4a3 in (F) after 12 hours of treatment: p = 0.0047, t = 4.851, df = 5. For Fus in (F) after 12 hours of treatment: p = 0.0055, t = 4.666, df = 5. For circHomer1 in (G) after 24 hours of treatment: p = 0.0032, t = 5.303, df = 5. For Eif4a3 in (G) after 12 hours of treatment: p = 0.0025, t = 5.591, df = 5. For Fus in (G) after 12 hours of treatment: p < 0.0001, t = 15.38, df = 5. For Eif4a3in (H): p = 0.0275, t = 4.029, df = 3. (I-K) Relative to vehicle circHomer1, Eif4a3 and Fus mRNA levels (based on qRT-PCR and normalized to 18S rRNA) after treatment with (I) CDPPB – mGluR5 positive allosteric modulator, (J) MK801 – NMDAR antagonist and (J) a combination of both MK801 and CDPPB as a rescue experiment – all treatments were done for 7 days. Mean ± SEM. *p < 0.05, **p < 0.01, ****p < 0.0001, based on two tailed one sample t-test relative to vehicle. For Eif4a3 in (I): p = 0.0378, t = 2.392, df = 10. For Fus in (I): p = 0.0110, t = 3.190, df = 9. For circHomer1 in (J): p = 0.0006, t = 4.708, df = 11. For Eif4a3 in (J): p = 0.0023, t = 3.928, df = 11. For Fus in (J): p = 0.0178, t = 2.784, df = 11. In all bar graphs the individual replicates are shown within the graph.

Fig 4: MDL100907, pure 5T2AR antagonist, robustly downregulates circHomer1, Eif4a3and Fus in brain regions that serotonergic receptors are highly present.(A-D) Relative to vehicle circHomer1, Eif4a3 and Fus mRNA levels (based on qRT-PCR and normalized to 18S rRNA) after treatment with MDL100907 in (A) Putamen - dorsal striatum, (B) Nucleus Accumbens – ventral striatum, (C) Orbitofrontal cortex and (D) Cerebellum. Mean ± SEM. Mice were sacrificed 24hrs after the last injection – delayed time point to clarify if the possible change of circHomer1 levels after pharmacological intervention is persistent over time. #0.10 < p < 0.05, *p < 0.05, **p < 0.01, ****p < 0.0001, based on two tailed one sample t-test relative to vehicle. For circHomer1 in (A): p = 0.0030, t = 4.034, df = 9. For Eif4a3 in (A): p = 0.0041, t = 3.606, df = 11. For Fus in (A): p = 0.0614, t = 2.082, df = 11. For circHomer1 in (B): p = 0.0238, t = 2.714, df = 9. For Eif4a3 in (B): p = 0.0366, t = 2.427, df = 11. For Fus in (B): p = 0.0848, t = 1.894, df = 11. For circHomer1 in (C): p = 0.0004, t = 5.134, df = 10. For Eif4a3in (C): p = 0.0102, t = 3.096, df = 11. For Fusin (C): p = 0.0499, t = 2.229, df = 10. For Fusin (D): p = 0.0848, t = 1.894, df = 11. In all bar graphs the individual replicates are shown within the graph.

Fig 5: circHomer1is indirectly regulated by CREB stimulation.(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 circHomer1after 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.