cAMP response element-binding protein is essential for the upregulation of brain-derived neurotrophic factor transcription, but not the behavioral or endocrine responses to antidepressant drugs

Abstract

Antidepressant drugs activate the cAMP signal transduction pathway through a variety of monoamine neurotransmitter receptors. Recently, molecular studies have identified a role for cAMP response element-binding protein (CREB) in the mechanism of action of chronically administered antidepressant drugs. However, the function of CREB in the behavioral and endocrine responses to these drugs has not been thoroughly investigated. We have used CREB-deficient mice to study the effects of two antidepressants, desipramine (DMI) and fluoxetine (FLX), in behavioral, endocrine, and molecular analyses. Behaviorally, CREB-deficient mice and wild-type mice respond similarly to DMI and FLX administration in the forced swim test and tail suspension test. Furthermore, the ability of DMI to suppress an acute corticosterone response after swim stress is maintained in CREB-deficient mice. However, upregulation of a molecular target of CREB, BDNF, is abolished in the CREB-deficient mice after chronic administration of DMI. These data are the first to demonstrate that CREB activation is upstream of BDNF mechanistically in response to antidepressant drug treatment. Therefore, although behavioral and endocrine responses to antidepressants may occur by CREB-independent mechanisms, CREB is critical to target gene regulation after chronic drug administration, which may contribute to long-term adaptations of the system to antidepressant drug treatment.

Fig. 1.

Behavioral response of wild-type and CREB^αΔ mice in the FST to subchronic antidepressant treatment. After a 15 min preswim on day 1, animals were administered saline or DMI (10.0 mg/kg, i.p.). On day 2 animals were administered saline or DMI (10.0 mg/kg, i.p.) 5 and 1 hr (20.0 mg/kg, s.c.) before a 6 min test swim. Saline-treated CREB^αΔ mutant mice demonstrated significantly lower immobility times than saline-treated wild-type mice. Subchronic DMI treatment significantly reduced immobility times in wild-type and CREB^αΔ mutant mice compared with respective saline controls. Results are presented as mean immobility ± SEM (in seconds). ANOVA and post hoc Fisher's pairwise comparisons revealed the following differences: *p< 0.05 versus wild-type saline; #p < 0.05 versus mutant saline (F_(1,27) = 54.6); **p < 0.05 versus wild-type saline (F_(1,27) = 24.2).

Fig. 2.

Behavioral response of wild-type and CREB^αΔ mice to chronic antidepressant treatment. Animals were administered DMI (10.0 mg/kg, i.p.) twice daily with 20.0 mg/kg, subcutaneously administered on testing days and measured for immobility in the forced swimming test. CREB^αΔmutant mice administered saline had significantly lower immobility times than saline-treated wild type mice. DMI administration significantly reduced immobility times in both wild-type and CREB mutant mice on days 1 and 14 of the chronic treatment paradigm. Results are presented as mean immobility ± SEM (in seconds). ANOVA andpost hoc Newman–Keuls pairwise comparisons revealed the following differences: +p < 0.05 versus saline, corresponding gene and day; *p < 0.05 versus different gene, same treatment and day.

Fig. 3.

Behavioral response of wild-type and CREB^αΔ mice in the TST to subchronic antidepressant treatment. Animals were administered saline, DMI, or FLX (20 mg/kg, i.p.) 30 min before a 6 min test session. Saline-treated CREB^αΔ mutant mice demonstrated significantly lower immobility times than saline-treated wild-type mice. Subchronic FLX and DMI significantly reduced immobility times in both wild-type and CREB^αΔ mutant mice compared with respective saline controls. Results are presented as mean immobility ± SEM (in seconds). ANOVA and post hoc Fisher's pairwise comparisons revealed the following differences: **p< 0.05 versus wild-type saline; ***p < 0.05 versus wild-type saline; +p < 0.05 versus wild-type fluoxetine; #p < 0.05 versus mutant saline (F_(2,48) = 16.3); *p < 0.05 versus wild-type saline (F_(1,48) = 3.8).

Fig. 4.

Regulation of corticosterone after exposure to the forced swimming test. Animals were examined for changes in the stress hormone corticosterone with and without acute exposure to the FST. In wild-type and CREB^αΔ mutant mice, swim stress induced a significant elevation in corticosterone compared with non-swim stressed control mice. Subchronic administration of DMI significantly blunted the elevation in corticosterone in both wild-type and mutant mice compared with respective saline controls. Values are plotted as mean corticosterone ± SEM (in micrograms per decaliter). ANOVA and post hoc Student–Newman–Keuls pairwise comparisons revealed the following differences: *p = 0.05 versus corresponding non-swim stressed saline group (F_(1,50) = 31) and **p < 0.05 versus corresponding swim-stressed saline group (F_(1,50) = 2).

Fig. 5.

RNase protection analysis of BDNF gene expression after chronic antidepressant treatment. The steady-state levels of BDNF and TBP (as internal standard) mRNAs were determined by RNase protection assay after chronic (21 d) drug administration in the frontal cortex and hippocampus of wild-type and CREB^αΔ mutant mice. Twenty micrograms of total RNAs were cohybridized with both riboprobes and analyzed as described in Materials and Methods. Three representative lanes are shown for saline and DMI treatment groups from wild-type and CREB^αΔ mutant mice (A, B, top). Radioactive bands for BDNF were quantified using a phosphorimager, and signals were normalized to those of TBP and plotted (A, B, bottom). DMI administration increased BDNF mRNA in the frontal cortex (A) and hippocampus (B) of wild-type mice compared with saline controls. P, Free probes; t, tRNA as control. ANOVA and post hoc Fisher's pairwise comparisons revealed the following differences: *p< 0.05 versus wild-type saline (F_(2,43) = 3.1); **p < 0.05 versus wild-type saline (F_(2,54) = 3.6). In CREB^αΔ mutant mice, however, DMI administration did not alter BDNF mRNA in the frontal cortex or hippocampus (A, B, respectively). No increases in BDNF mRNA were observed in either the frontal cortex or hippocampal regions after chronic FLX in wild-type or CREB^αΔ mutant mice (A, B, respectively).