7,8-Dihydroxyflavone, a small molecule TrkB agonist, improves spatial memory and increases thin spine density in a mouse model of Alzheimer disease-like neuronal loss

Abstract

Augmenting BDNF/TrkB signaling has been demonstrated to be a promising strategy for reversing cognitive deficits in preclinical models of Alzheimer disease (AD). Although these studies highlight the potential of targeting BDNF/TrkB signaling, this strategy has not yet been tested in a model that develops the disease features that are most closely associated with cognitive decline in AD: severe synaptic and neuronal loss. In the present study, we investigated the impact of 7,8-dihydroxyflavone (DHF), a TrkB agonist, in CaM/Tet-DTA mice, an inducible model of severe neuronal loss in the hippocampus and cortex. Systemic 7,8-DHF treatment significantly improved spatial memory in lesioned mice, as measured by water maze. Analysis of GFP-labeled neurons in CaM/Tet-DTA mice revealed that 7,8-DHF induced a significant and selective increase in the density of thin spines in CA1 of lesioned mice, without affecting mushroom or stubby spines. These findings suggest chronic upregulation of TrkB signaling with 7,8-DHF may be an effective and practical strategy for improving function in AD, even after substantial neuronal loss has occurred.

Figure 1. CaM/Tet-DT_A mice develop neuronal loss in Alzheimer-disease related brain areas.

A 25-day withholding of doxycycline from the diet of CaM/Tet-DT_A mice induces DT_A expression, which causes a selective loss of hippocampal and cortical neurons. Representative images depict a significant loss of NeuN immunoreactivity in the CA1 and DG of the hippocampus (A), and layers II/III of the entorhinal cortex (B, n = 5–6, quantified by optical densitometry in C). Crossing CaM/Tet-DT_A mice with mice carrying a Thy1-GFP-M transgene produces sparse GFP-labeling of CA1 pyramidal neurons (D). Scale bars = 1 mm.

Figure 2. 7,8-DHF treatment improves spatial learning and memory in CaM/Tet-DT_A mice.

Lesioned CaM/Tet-DT_A mice exhibit a significantly increased preference for the open arms of an elevated plus maze compared to controls (A, n = 9–12, p<0.0001). All groups made a similar total number of arm entries (B). 7,8-DHF treatment did not alter preference for open or closed arms (A–B). A rotarod test reveals no differences between groups in the time to fall off an accelerating rod (C). During water maze training, lesioned mice treated with 7,8-DHF were not significantly slower to reach the escape platform compared to controls, except for day 3 (D, n = 9–12, p<0.001). On day 4, lesioned mice treated with 7,8-DHF reached the platform significantly faster than lesioned mice that received vehicle (D, n = 9–12, #, p<0.05). On the 24 hr probe trial, latencies to reach and mean proximity to the former platform location were not significantly different between 7,8-DHF-treated lesioned mice and controls (E–F, n = 9–12, p>0.05). All groups had a similar average swim speed during the probe trial (G).

Figure 3. 7,8-DHF treatment increases thin spine density on dendrites in CA1 pyramidal neurons.

High-magnification confocal imaging and analysis of dendrites from GFP-expressing CA1 pyramidal neurons reveals elevated total spine density in 7,8-DHF-treated lesioned mice (A–B, n = 5–7, p<0.05). Classification of thin (T), mushroom (M), and stubby (S) spines indicates a specific elevation of thin spines in lesioned mice (C, n = 5–7, p<0.05), which is further elevated by 7,8-DHF treatment (n = 5–7, p<0.01 versus vehicle-treated lesioned mice). Analysis of synaptophysin immunoreactivity reveals a lesion-induced increase in presynaptic terminal packing density in CA1 stratum oriens (ori) and radiatum (rad) (D–E, n = 5, p<0.05), and loss of density in the stratum lacunosum-moleculare (D–E, n = 5, p<0.01), but no effect of 7,8-DHF. Labeling for MAP2 reveals a significant loss of CA1 pyramidal neuron dendrites in lesioned groups (F–G, n = 5–7, p<0.001), and no impact of 7,8-DHF treatment on dendrite density. Scale bar = 1 µm.