Autocrine BDNF–TrkB signalling within a single dendritic spine

Abstract

Brain-derived neurotrophic factor (BDNF) and its receptor TrkB are crucial for many forms of neuronal plasticity1, 2, 3, 4, 5, 6, including structural long-term potentiation (sLTP)7, 8, which is a correlate of an animal’s learning7, 9, 10, 11, 12. However, it is unknown whether BDNF release and TrkB activation occur during sLTP, and if so, when and where. Here, using a fluorescence resonance energy transfer-based sensor for TrkB and two-photon fluorescence lifetime imaging microscopy13, 14, 15, 16, we monitor TrkB activity in single dendritic spines of CA1 pyramidal neurons in cultured murine hippocampal slices. In response to sLTP induction9, 14, 15, 16, we find fast (onset < 1 min) and sustained (>20 min) activation of TrkB in the stimulated spine that depends on NMDAR (N-methyl-d-aspartate receptor) and CaMKII signalling and on postsynaptically synthesized BDNF. We confirm the presence of postsynaptic BDNF using electron microscopy to localize endogenous BDNF to dendrites and spines of hippocampal CA1 pyramidal neurons. Consistent with these findings, we also show rapid, glutamate-uncaging-evoked, time-locked BDNF release from single dendritic spines using BDNF fused to superecliptic pHluorin17, 18, 19. We demonstrate that this postsynaptic BDNF–TrkB signalling pathway is necessary for both structural and functional LTP20. Together, these findings reveal a spine-autonomous, autocrine signalling mechanism involving NMDAR–CaMKII-dependent BDNF release from stimulated dendritic spines and subsequent TrkB activation on these same spines that is crucial for structural and functional plasticity.

Rho GTPase complementation underlies BDNF-dependent homo- and heterosynaptic plasticity

“…To test whether crosstalk requires BDNF–TrkB–Rac1 signalling, we first used partial pharmacological inhibition of this pathway. Although strong inhibition of BDNF signalling has been shown to impair sLTP 15 , we found that weak inhibition of this pathway with a low concentration (0.25 μg ml −1 ) of TrkB-Ig preserved sLTP (Δ V supra sustained = 58 ± 13% (mean ± s.e.m. ), in which V denotes spine volume), but attenuated crosstalk (Δ V sub sustained = 20 ± 8%), suggesting that BDNF is required for this process (Fig.…”

“…We found that removal of postsynaptic BDNF significantly attenuated Rac1 and Cdc42 activation during sLTP without affecting RhoA (Fig. 1f, g), and reduced the associated expression of sLTP 15 (Extended Data Fig. 5a, b).…”

“…We next examined whether postsynaptic BDNF 15 , which also links NMDAR–CaMKII signalling and sLTP, is required for Rho GTPase activation during sLTP. To do this, we used a single-cell knockout technique 15, 16 with organotypic hippocampal slices from BDNF conditional knockout mice ( Bdnf fl/fl ) 17, 36 coupled with biolistic transfection of Cre recombinase alongside a Rho GTPase sensor.…”

“…To do this, we used a single-cell knockout technique 15, 16 with organotypic hippocampal slices from BDNF conditional knockout mice ( Bdnf fl/fl ) 17, 36 coupled with biolistic transfection of Cre recombinase alongside a Rho GTPase sensor. We found that removal of postsynaptic BDNF significantly attenuated Rac1 and Cdc42 activation during sLTP without affecting RhoA (Fig.…”

Imaging ERK and PKA Activation in Single Dendritic Spines during Structural Plasticity

“…Two-photon fluorescence lifetime imaging microscopy (2pFLIM), in combination with FRET biosensors, has been proven to be useful for imaging signaling cascades during structural and functional LTP (Harvey et al, 2008b, Lee et al, 2009, Murakoshi et al, 2011, Zhai et al, 2013, Bosch et al, 2014, Kim et al, 2015, Harward et al, 2016, Hedrick et al, 2016). When single spines are stimulated with glutamate uncaging or electrical stimulation, Ca 2+ influx through NMDA receptors triggers complex signaling cascades, including the activation of protein kinases and small GTPase proteins, in turn regulating actin polymerization and increasing spine volume and postsynaptic sensitivity to glutamate (Nishiyama and Yasuda, 2015).…”