Regulation of CREB function in rat frontal cortex after combined treatment with Fluoxetine and Olanzapine

Statement of the Study: Generally, drugs used in the treatment of depression exert their therapeutic effect after 4/6 weeks and only in 60–65% of patients. The search for an adequate and faster treatment of major depression is one of the main challenges in neuropsychopharmacology. Recently, a clinical study of treatmentresistant depressed patients without a psychotic component, showed that after only one week of treatment, Fluoxetine (a selective serotonin reuptake inhibitor antidepressant) and Olanzapine (an atypical antipsychotic agent) produced a higher level of improvement than either monotherapy alone (Shelton et al., American Journal of Psychiatry 158(1), 131–134, 2001). Furthermore, preclinical data, using microdialysis, indicated that the combination of Olanzapine and Fluoxetine resulted in an increase in the extracellular levels of dopamine and norepinephrine in the rat prefrontal cortex, an effect that was significantly bigger than after treatment with either drug alone (Zhang et al., Neuropsychopharmacology 23(3),250– 262, 2000). However, it is not yet completely understood which intracellular signaling pathway could be involved in the fast response (seen in clinical trials) to Fluoxetine plus Olanzapine co-treatment. Methods: Since antidepressant and antipsychotic drugs affect the cyclic adenosine monophosphate (cAMP) pathway, including the expression of the cAMP response element binding protein (CREB), the levels of CREB mRNA and CREB nuclear protein, total and phosphorylated, were studied in the frontal cortex of rats using RNase protection assay and Western Blotting analysis respectively. Four experimental groups were used: rats were treated for one, five or ten days with either saline, Fluoxetine (ip 10 mg/Kg), Olanzapine (sc 1 mg/Kg) or combined Fluoxetine plus Olanzapine (10 mg/Kg and 1 mg/Kg). Summary of Results: Our results show that the level of phosphorylated CREB Ser133 was significantly increased in the frontal cortex of rats receiving the combined treatment regimen (Fluoxetine plus Olanzapine) for five days. No effect was observed in acutely and ten day treated rats. Conclusion: This specific effect on CREB phosphorylation levels after five days of combined treatment with Fluoxetine plus Olanzapine might represent one of the mechanisms underlying the faster response to this therapy recently observed in several clinical trials.