TrkB Signaling in Dorsal Raphe Nucleus is Essential for Antidepressant Efficacy and Normal Aggression Behavior

Abstract

Brain-derived neurotrophic factor (BDNF) and its high affinity receptor, tropomyosin receptor kinase B (TrkB), have important roles in neural plasticity and are required for antidepressant efficacy. Studies examining the role of BDNF-TrkB signaling in depression and antidepressant efficacy have largely focused on the limbic system, leaving it unclear whether this signaling is important in other brain regions. BDNF and TrkB are both highly expressed in the dorsal raphe nucleus (DRN), a brain region that has been suggested to have a role in depression and antidepressant action, although it is unknown whether BDNF and TrkB in the dorsal raphe nucleus are involved in these processes. We combined the adeno-associated virus (AAV) with the Cre-loxP site-specific recombination system to selectively knock down either Bdnf or TrkB in the DRN. These mice were then characterized in several behavioral paradigms including measures of depression-related behavior and antidepressant efficacy. We show that knockdown of TrkB, but not Bdnf, in the DRN results in loss of antidepressant efficacy and increased aggression-related behavior. We also show that knockdown of TrkB or Bdnf in this brain region does not have an impact on weight, activity levels, anxiety, or depression-related behaviors. These data reveal a critical role for TrkB signaling in the DRN in mediating antidepressant responses and normal aggression behavior. The results also suggest a non-cell autonomous role for BDNF in the DRN in mediating antidepressant efficacy.

Figure 1

Localized knockdown of Bdnf in dorsal raphe nucleus (DRN). (a) Schematic of a coronal section containing DRN, highlighted in gray. (b, c) DRN containing green fluorescent protein (GFP) epifluorescence were laser micro-dissected and subjected to real-time PCR for quantification of Cre and Bdnf mRNA levels (adeno-associated virus expressing GFP (AAV-GFP), N=12; AAV-GFP tagged to Cre recombinase (AAV-GFP-Cre), N=12).

Figure 2

Mice with dorsal raphe nucleus (DRN)-specific Bdnf knockdown display overall normal behavioral phenotypes. (a) Mice injected with adeno-associated virus expressing GFP tagged to Cre recombinase (AAV-GFP-Cre) exhibited a significant increase in locomotor activity in the first 5 min, in comparison with those with AAV-GFP. However, the locomotor activity for the rest of 2 h testing period was similar between both groups of animals. The total number of beam breaks during the test period was indistinguishable between AAV-GFP and AAV-GFP-Cre mice (see the inset). (b) Anxiety-like behavioral phenotype was assessed by the open-field test. (c) In a resident-intruder paradigm, latency to attack a male intruder was measured in flBdnf mice injected with either AAV-GFP or AAV-GFP-Cre into DRN. (d) Depression-like behavior was assessed by the sucrose consumption test. (AAV-GFP, N=12; AAV-GFP-Cre, N=12 for a, b, c, and d.) (e) Responses to an antidepressant, desipramine, were examined by the forced swim test (FST). In either AAV injection, sub-chronic treatment with desipramine resulted in significantly less immobile behavior in comparison with saline treatment, indicating a normal response to the antidepressant. (AAV-GFP/saline, N=9; AAV-GFP/desipramine, N=10; AAV-GFP-Cre/saline, N=8; AAV-GFP-Cre/desipramine, N=7; two-way ANOVA (F_{1, 30})=10.34, p=0.0031 for treatment followed by multiple comparison with t-test, *p<0.05.)

Figure 3

Localized knockdown of tropomyosin receptor kinase B (TrkB) in dorsal raphe nucleus (DRN). (a, b) DRN containing green fluorescent protein (GFP) epifluorescence were laser micro-dissected out and subjected to real-time PCR for quantification of (a) Cre and (b) TrkB mRNA levels. (Adeno-associated virus expressing GFP (AAV-GFP), N=12; AAV-GFP tagged to Cre recombinase (AAV-GFP-Cre), N=14.)

Figure 4

Mice with localized knockdown of tropomyosin receptor kinase B (TrkB) in dorsal raphe nucleus (DRN) display normal locomotor, anxiety-like, and depression-like behavior, but have an attenuated response to antidepressants. (a) Locomotor activity as assessed by the number of horizontal beam breaks over a 2 h testing period in 5 min increments. Total beam breaks during the test period were indistinguishable between adeno-associated virus expressing GFP (AAV-GFP) and AAV-GFP tagged to Cre recombinase (AAV-GFP-Cre)-injected flTrkB mice (see the inset). (b) The animals were assessed for anxiety-like behavior using the open-field test. (c) Aggression-like behavior of flTrkB mice injected with either AAV-GFP or AAV-GFP-Cre was examined in the resident-intruder test. Latency to attack the intruder was scored. (d) Depression-like behavior was measured by the sucrose consumption test (AAV-GFP, N=15; AAV-GFP-Cre, N=16 for a, b, c, and d). (e) Antidepressant responses to desipramine were examined by forced swim test (FST). Floxed TrkB mice injected with AAV-GFP, but not AAV-GFP-Cre, showed less immobility in response to sub-chronic treatment with desipramine, in comparison with saline-treated animals (AAV-GFP/saline, N=8; AAV-GFP/desipramine, N=7; AAV-GFP-Cre/saline, N=8; AAV-GFP-Cre/desipramine, N=8; two-way ANOVA (F_{1, 27})=11.24, p=0.0024 for treatment followed by multiple comparison with t-test, *p<0.05). (f) Chronic treatment with fluoxetine resulted in less immobility in flTrkB mice with AAV-GFP, but not in AAV-GFP-Cre. (AAV-GFP/saline, N=7; AAV-GFP/desipramine, N=7; AAV-GFP-Cre/saline, N=6; AAV-GFP-Cre/desipramine, N=6; a multiple t-test, followed by Holm–Sidak post-hoc analysis to correct for multiple comparisons, revealed that for GFP-SA vs GFP-Flx, p=0.03, with all other group comparisons are nonsignificant.)