Selective loss of brain-derived neurotrophic factor in the dentate gyrus attenuates antidepressant efficacy

Abstract

Background: Brain-derived neurotrophic factor (BDNF) plays an important role in neural plasticity in the adult nervous system and has been suggested as a target gene for antidepressant treatment. The neurotrophic hypothesis of depression suggests that loss of BDNF from the hippocampus contributes to an increased vulnerability for depression, whereas upregulation of BDNF in the hippocampus is suggested to mediate antidepressant efficacy.

Methods: We have used a viral-mediated gene transfer approach to assess the role of BDNF in subregions of the hippocampus in a broad array of behavioral paradigms, including depression-like behavior and antidepressant responses. We have combined the adeno-associated virus (AAV) with the Cre/loxP site-specific recombination system to induce the knockout of BDNF selectively in either the CA1 or dentate gyrus (DG) subregions of the hippocampus.

Results: We show that the loss of BDNF in either the CA1 or the DG of the hippocampus does not alter locomotor activity, anxiety-like behavior, fear conditioning, or depression-related behaviors. However, the selective loss of BDNF in the DG but not the CA1 region attenuates the actions of desipramine and citalopram in the forced swim test.

Conclusions: These data suggest that the loss of hippocampal BDNF per se is not sufficient to mediate depression-like behavior. However, these results support the view that BDNF in the DG might be essential in mediating the therapeutic effect of antidepressants.

Figure 1

Region Specific Deletion of BDNF in Subregions of the Hippocampus. (A, B) BDNF and Cre recombinase expression was analyzed by double fluorescent in situ hybridization in mice injected with AAV-GFP or AAV-Cre virus into either CA1 (A) or DG (B). Probes for BDNF and Cre recombinase were coupled to Cy3 and FITC epifluorescence, respectively. In AAV-GFP injected mice, BDNF was strongly present in the CA1 and DG subregions of the hippocampus while no detectable expression of Cre recombinase was detected. In contrast, in the AAV-Cre injected mice, the localized expression of Cre was observed in either the CA1 or DG depending on the injection site, and in these regions the levels of BDNF expression was reduced. DAPI staining was used to show that the loss of BDNF expression was not due to changes in cell number. (C, D) The entire region of the CA1 (C) and DG (D) were laser micro-dissected out from brain sections of AAV-GFP and AAV-Cre injected mice. The sections were subjected to QRT-PCR analyses for quantitation of Cre and BDNF mRNA levels. Cre expression was virtually absent in AAV-GFP injected mice but significantly increased in AAV-Cre injected animals. AAV-Cre injected animals in both CA1 and DG had significant reduction (52% and 62%, respectively) in BDNF mRNA compared to AAV-GFP mice (CA1, AAV-GFP, n = 6; AAV-CRE, n = 7: DG, AAV-GFP, n = 10; AAV-Cre, n = 13).

Figure 2

Localized Deletion of BDNF in the CA1 and DG Subregions Does Not Alter Locomotor Activity. (A) Male mice injected into the CA1 with either AAV-Cre or AAV-GFP exhibited no significant difference in locomotor activity (ambulation), as assessed by a consecutive horizontal beam break, over a two-hour period in either 5 minute increments or over the entire two-hour period (see insert; AAV-GFP, n = 12; AAV-Cre, n = 11). (B) The male mice injected into the DG region with either AAV-Cre or AAV-GFP exhibited a similar level of locomotor activity during the testing period whether analyzed as 5 minute increments or total number of beam breaks over the entire two-hour period (see insert; AAV-GFP, n = 23; AAV-Cre, n = 24).

Figure 3

Localized Deletion of BDNF in the CA1 and DG Does Not Alter Anxiety Related Behavior. (A) The CA1 injected mice with AAV-Cre displayed a similar level of anxiety, as determined by the time spent in the center, dark side, and light side of the elevated plus maze compared to AAV-GFP injected mice (AAV-GFP, n = 12; AAV-Cre, n = 11). (B) DG injected mice with AAV-Cre exhibited normal anxiety, as assessed in the elevated plus maze, compared to AAV-GFP controls (CTL) (AAV-GFP, n = 21; AAV-Cre, n = 22).

Figure 4

Context and Cue-Dependent Fear Conditioning is Unaltered by the loss of BDNF Selectively in the CA1 or DG region. (A) The selective loss of BDNF in the CA1 did not result in any difference in context-dependent fear conditioning 24 hours after training compared to AAV-GFP injected mice. No significant difference was observed in baseline freezing behavior. Cue-dependent fear conditioning was also indistinguishable in AAV-Cre or AAV-GFP injected mice (AAV-GFP, n = 12; AAV-Cre, n = 11). (B) The loss of BDNF in the DG did not produce any significant difference in context-dependent or cue-dependent fear conditioning compared to AAV-GFP injected animals (AAV-GFP, n = 17; AAV-Cre, n = 16). No significant difference was observed in baseline freezing behavior.

Figure 5

The Localized Deletion of BDNF in the CA1 or DG Region does not Alter Preference for a Natural Reward in the Sucrose Preference Paradigm. (A) Male floxed BDNF mice injected with either AAV-Cre or AAV-GFP in the CA1 display a similar preference for the sucrose solution (AAV-GFP, n = 12; AAV-Cre, n = 11). (B) Male floxed BDNF mice with a localized deletion of BDNF in the DG region have indistinguishable levels of intake of the sucrose solution compared to injected AAV-GFP mice. The total intake of liquid (water plus sucrose solution) was unchanged between the groups (p > 0.05). Results are presented as mean of sucrose preference (the percentage of sucrose solution ingested relative to the total amount of liquid consumed) ± SEM (AAV-GFP, n = 12; AAV-Cre, n = 13).

Figure 6

The Loss of BDNF in the DG Attenuates Antidepressant Responses in the Forced Swim Test. (A–C) Male floxed BDNF mice injected with AAV-Cre in the DG or CA1 subregion of the hippocampus display a similar % of immobility in the FST as compared to AAV-GFP mice (CTL). (A) Antidepressant treatment with desipramine significantly reduced immobility time in AAV-GFP mice (p < 0.05) as well as mice with a selective reduction of BDNF in the CA1 subregion of the hippocampus (p < 0.05). (AAV-GFP, n = 12; AAV-Cre, n = 11). (B) Desipramine treatment significantly reduced immobility time in AAV-GFP mice (p < 0.05) but not in the mice with a selective loss of BDNF in the DG (AAV-GFP, n = 12; AAV-Cre, n = 13). (C) Using the serotonin-selective reuptake inhibitor, citalopram, we found that this antidepressant also significantly reduced immobility time in AAV-GFP mice (p < 0.05) but not in the mice with a selective loss of BDNF in the DG (AAV-GFP, n = 11; AAV-Cre, n = 11).