Ama“Zinc” Link between TrkB Transactivation and Synaptic Plasticity

While Trk receptors can be activated in a neurotrophin-independent manner through “transactivation” by GPCR ligands, its physiological significance in the brain remains unknown. Huang et al. have now identified a novel mechanism of TrkB transactivation. They show that zinc ions can transactivate TrkB independent of neurotrophins and that such a transactivation is important for mossy fiber long-term potentiation (LTP)

Multiple functions of precursor BDNF to CNS neurons: negative regulation of neurite growth, spine formation and cell survival

<p>Abstract</p> <p>Background</p> <p>Proneurotrophins and mature neurotrophins elicit opposite effects via the p75 neurotrophin receptor (p75^NTR) and Trk tyrosine kinase receptors, respectively; however the molecular roles of proneurotrophins in the CNS are not fully understood.</p> <p>Results</p> <p>Based on two rare single nucleotide polymorphisms (SNPs) of the <it>human brain-derived neurotrophic factor (BDNF) </it>gene, we generated R125M-, R127L- and R125M/R127L-BDNF, which have amino acid substitution(s) near the cleavage site between the pro- and mature-domain of BDNF. Western blot analyses demonstrated that these BDNF variants are poorly cleaved and result in the predominant secretion of proBDNF. Using these cleavage-resistant proBDNF (CR-proBDNF) variants, the molecular and cellular roles of proBDNF on the CNS neurons were examined. First, CR-proBDNF showed normal intracellular distribution and secretion in cultured hippocampal neurons, suggesting that inhibition of proBDNF cleavage does not affect intracellular transportation and secretion of BDNF. Second, we purified recombinant CR-proBDNF and tested its biological effects using cultured CNS neurons. Treatment with CR-proBDNF elicited apoptosis of cultured cerebellar granule neurons (CGNs), while treatment with mature BDNF (matBDNF) promoted cell survival. Third, we examined the effects of CR-proBDNF on neuronal morphology using more than 2-week cultures of basal forebrain cholinergic neurons (BFCNs) and hippocampal neurons. Interestingly, in marked contrast to the action of matBDNF, which increased the number of cholinergic fibers and hippocampal dendritic spines, CR-proBDNF dramatically reduced the number of cholinergic fibers and hippocampal dendritic spines, without affecting the survival of these neurons.</p> <p>Conclusion</p> <p>These results suggest that proBDNF has distinct functions in different populations of CNS neurons and might be responsible for specific physiological cellular processes in the brain.</p

Pro-BDNF–induced synaptic depression and retraction at developing neuromuscular synapses

Postsynaptic cells generate positive and negative signals that retrogradely modulate presynaptic function. At developing neuromuscular synapses, prolonged stimulation of muscle cells induces sustained synaptic depression. We provide evidence that pro–brain-derived neurotrophic factor (BDNF) is a negative retrograde signal that can be converted into a positive signal by metalloproteases at the synaptic junctions. Application of pro-BDNF induces a dramatic decrease in synaptic efficacy followed by a retraction of presynaptic terminals, and these effects are mediated by presynaptic pan-neurotrophin receptor p75 (p75NTR), the pro-BDNF receptor. A brief stimulation of myocytes expressing cleavable or uncleavable pro-BDNF elicits synaptic potentiation or depression, respectively. Extracellular application of metalloprotease inhibitors, which inhibits the cleavage of endogenous pro-BDNF, facilitates the muscle stimulation–induced synaptic depression. Inhibition of presynaptic p75NTR or postsynaptic BDNF expression also blocks the activity-dependent synaptic depression and retraction. These results support a model in which postsynaptic secretion of a single molecule, pro-BDNF, may stabilize or eliminate presynaptic terminals depending on its proteolytic conversion at the synapses

BDNF Facilitates L-LTP Maintenance in the Absence of Protein Synthesis through PKMζ

Late-phase long term potentiation (L-LTP) is thought to be the cellular basis for long-term memory (LTM). While LTM as well as L-LTP is known to depend on transcription and translation, it is unclear why brain-derived neurotrophic factor (BDNF) could sustain L-LTP when protein synthesis is inhibited. The persistently active protein kinase ζ (PKMζ) is the only molecule implicated in perpetuating L-LTP maintenance. Here, in mouse acute brain slices, we show that inhibition of PKMζ reversed BDNF-dependent form of L-LTP. While BDNF did not alter the steady-state level of PKMζ, BDNF together with the L-LTP inducing theta-burst stimulation (TBS) increased PKMζ level even without protein synthesis. Finally, in the absence of de novo protein synthesis, BDNF maintained TBS-induced PKMζ at a sufficient level. These results suggest that BDNF sustains L-LTP through PKMζ in a protein synthesis-independent manner, revealing an unexpected link between BDNF and PKMζ

ProNGF promotes neurite growth from a subset of NGF-dependent neurons by a p75(NTR)-dependent mechanism

The somatosensory and sympathetic innervation of the vertebrate head is derived principally from the neurons of trigeminal and superior cervical ganglia (SCG), respectively. During development, the survival of both populations of neurons and the terminal growth and branching of their axons in the tissues they innervate is regulated by the supply of nerve growth factor (NGF) produced by these tissues. NGF is derived by proteolytic cleavage of a large precursor protein, proNGF, which is recognised to possess distinctive biological functions. Here, we show that proNGF promotes profuse neurite growth and branching from cultured postnatal mouse SCG neurons. In marked contrast, proNGF does not promote the growth of trigeminal neurites. Studies using compartment cultures demonstrated that proNGF acts locally on SCG neurites to promote growth. The neurite growth-promoting effect of proNGF is not observed in SCG neurons cultured from p75NTR-deficient mice, and proNGF does not phosphorylate the NGF receptor tyrosine kinase TrkA. These findings suggest that proNGF selectively promotes the growth of neurites from a subset of NGF-responsive neurons by a p75NTR-dependent mechanism during postnatal development when the axons of these neurons are ramifying within their targets in vivo

BDNF modulates activity-dependent PKMζ level to sustain L-LTP in the absence of protein synthesis.

<p>(A, B) Rescuing L-LTP impairment by BDNF in the presence of anisomycin is PKMζ-dependent. (A) Applications of various drugs were indicated by horizontal bars. Anisomycin (40 µM) was used throughout the entire experiment. BDNF (200 ng/ml) was applied 3 minutes after 12TBS stimulation and successfully rescued L-LTP impairment. ZIP was applied at 1 hour after stimulation and completely abolished L-LTP. (B) Quantification of the initial slope from the last 5 minutes of recording. (C) PKMζ protein level of hippocampal CA1 derived from WT mice at 1 hour and 3 hours after 12TBS stimulation. Tubulin was used as loading control. The 12 TBS group was normalized against control group. The 12 TBS plus BDNF and anisomycin treatment groups were normalized against that without BDNF treatment. Representative blots are shown on top of the quantification of data (3–5 slices per treatment, n = 3 independent experiments).</p

Steady-State PKMζ protein level is not regulated by BDNF.

<p>(A) PKMζ protein level in cortex and hippocampus of BDNF KO and WT littermates. At postnatal day 18, cortex or hippocampus from BDNF KO and WT littermates were dissected and subjected to Western blot. Representative blots and quantification of data were shown. GAPDH was used as loading control. (n = 5–8 independent experiments). (B) PKMζ expression in primary neuron cultures derived from different genotypes after BDNF treatment. DIV 7 cortical primary cultures of WT, Het or KO genotype were separately treated with BDNF (100 ng/ml) or vehicle for 24 hours. For each experiment, BDNF treatment group was normalized against vehicle treatment group. Representative blots are shown on top of the quantification of data. (n = 5 independent experiments).</p

BDNF-dependent late phase LTP is mediated by PKMζ.

<p>(A, B) 12 TBS-induced L-LTP was reversed by PKMζ inhibitor ZIP. Field EPSP (fEPSP) was evoked in CA1 stratum radiatum by stimulating Schaffer Collateral in adult C57BL/6 mice. (A) After a stable baseline was obtained, 12 TBS was conducted. LTP was sustained at least for 3 hours. ZIP (5 µM) or scrambled ZIP peptide (5 µM) was applied at 1 hour after stimulation. (B) Quantification of the initial slope value from the last 5 minutes recording. (C, D) Forskolin-induced L-LTP was abolished by PKMζ inhibitor ZIP. The experiments were done identically as in (A), except that L-LTP was induced by a transient perfusion of forskolin (50 µM) and IBMX (30 µM) for 15 minutes. ZIP or scrambled ZIP was applied at 80 minutes after chemical induction when stable L-LTP was fully established. Numbers of slices and mice used in each condition are indicated at the top of each plot. In this and all other figures, data are presented as mean ± s.e.m. * p<0.05, ** p<0.01,*** p<0.001, Student's t-Test.</p