CPEB2-activated axonal translation of VGLUT2 mRNA promotes glutamatergic transmission and presynaptic plasticity.

BACKGROUND: Local translation at synapses is important for rapidly remodeling the synaptic proteome to sustain long-term plasticity and memory. While the regulatory mechanisms underlying memory-associated local translation have been widely elucidated in the postsynaptic/dendritic region, there is no direct evidence for which RNA-binding protein (RBP) in axons controls target-specific mRNA translation to promote long-term potentiation (LTP) and memory. We previously reported that translation controlled by cytoplasmic polyadenylation element binding protein 2 (CPEB2) is important for postsynaptic plasticity and memory. Here, we investigated whether CPEB2 regulates axonal translation to support presynaptic plasticity. METHODS: Behavioral and electrophysiological assessments were conducted in mice with pan neuron/glia- or glutamatergic neuron-specific knockout of CPEB2. Hippocampal Schaffer collateral (SC)-CA1 and temporoammonic (TA)-CA1 pathways were electro-recorded to monitor synaptic transmission and LTP evoked by 4 trains of high-frequency stimulation. RNA immunoprecipitation, coupled with bioinformatics analysis, were used to unveil CPEB2-binding axonal RNA candidates associated with learning, which were further validated by Western blotting and luciferase reporter assays. Adeno-associated viruses expressing Cre recombinase were stereotaxically delivered to the pre- or post-synaptic region of the TA circuit to ablate Cpeb2 for further electrophysiological investigation. Biochemically isolated synaptosomes and axotomized neurons cultured on a microfluidic platform were applied to measure axonal protein synthesis and FM4-64FX-loaded synaptic vesicles. RESULTS: Electrophysiological analysis of hippocampal CA1 neurons detected abnormal excitability and vesicle release probability in CPEB2-depleted SC and TA afferents, so we cross-compared the CPEB2-immunoprecipitated transcriptome with a learning-induced axonal translatome in the adult cortex to identify axonal targets possibly regulated by CPEB2. We validated that Slc17a6, encoding vesicular glutamate transporter 2 (VGLUT2), is translationally upregulated by CPEB2. Conditional knockout of CPEB2 in VGLUT2-expressing glutamatergic neurons impaired consolidation of hippocampus-dependent memory in mice. Presynaptic-specific ablation of Cpeb2 in VGLUT2-dominated TA afferents was sufficient to attenuate protein synthesis-dependent LTP. Moreover, blocking activity-induced axonal Slc17a6 translation by CPEB2 deficiency or cycloheximide diminished the releasable pool of VGLUT2-containing synaptic vesicles. CONCLUSIONS: We identified 272 CPEB2-binding transcripts with altered axonal translation post-learning and established a causal link between CPEB2-driven axonal synthesis of VGLUT2 and presynaptic translation-dependent LTP. These findings extend our understanding of memory-related translational control mechanisms in the presynaptic compartment.

Proximodistal Heterogeneity in Learning-promoted Pathway-specific Plasticity at Dorsal CA1 Synapses.

Contextual learning requires the delivery of AMPA receptors to CA1 synapses in the dorsal hippocampus. However, proximodistal heterogeneity of pathway-specific plasticity remains unclear. Here, we examined the proximodistal heterogeneity in learning-induced plasticity at the CA1 synapses with inputs from the entorhinal cortex layer III (ECIII) or from CA3. We subjected male rats to an inhibitory avoidance task and prepared acute hippocampal slices for whole-cell patch clamp experiments, where we stimulated ECIII-CA1 or CA3-CA1 input fibers to analyze evoked excitatory postsynaptic currents (EPSCs). Compared to untrained controls, trained rats exhibited higher AMPA/NMDA current ratios at CA3-CA1 synapses of proximal and intermediate, but not distal CA1 neurons, which suggested that region-specific plasticity occurred after learning. Moreover, trained rats exhibited higher AMPA/NMDA current ratios at ECIII-CA1 synapses of intermediate and distal, but not proximal CA1 neurons. These findings suggested the presence of proximodistal heterogeneity in pathway-specific postsynaptic plasticity. Regarding presynaptic plasticity, training slightly, but significantly increased the paired-pulse ratios of CA3-CA1 synapses of proximal and intermediate, but not distal CA1 neurons. Moreover, trained rats exhibited higher paired-pulse ratios at ECIII-CA1 synapses of intermediate and distal, but not proximal CA1 neurons, which suggested region-specific presynaptic plasticity. Finally, learning was clearly prevented by the bilateral microinjection of a plasticity blocker in the proximal or intermediate, but not distal CA1 subfields, which suggested functional heterogeneity along the proximodistal axis. Understanding region- and pathway-specific plasticity at dorsal CA1 synapses could aid in controlling encoded memory.

Impaired function of α2-containing nicotinic acetylcholine receptors on oriens-lacunosum moleculare cells causes hippocampus-dependent memory impairments.

Children of mothers who smoked during pregnancy are at significantly greater risk for cognitive impairments including memory deficits, but the mechanisms underlying this effect remain to be understood. In rodent models of smoking during pregnancy, early postnatal nicotine exposure results in impaired long-term hippocampus-dependent memory, functional loss of α2-containing nicotinic acetylcholine receptors (α2∗ nAChRs) in oriens-lacunosum moleculare (OLM) cells, increased CA1 network excitation, and unexpected facilitation of long-term potentiation (LTP) at Schaffer collateral-CA1 synapses. Here we demonstrate that α2 knockout mice show the same pattern of memory impairment as previously observed in wild-type mice exposed to early postnatal nicotine. However, α2 knockout mice and α2 knockout mice exposed to early postnatal nicotine did not share all of the anomalies in hippocampal function observed in wild-type mice treated with nicotine during development. Unlike nicotine-treated wild-type mice, α2 knockout mice and nicotine-exposed α2 knockout mice did not demonstrate increased CA1 network excitation following Schaffer collateral stimulation and facilitated LTP, indicating that the effects are likely adaptive changes caused by activation of α2∗ nAChRs during nicotine exposure and are unlikely related to the associated memory impairment. Thus, the functional loss of α2∗ nAChRs in OLM cells likely plays a critical role in mediating this developmental-nicotine-induced hippocampal memory deficit.

Early postnatal nicotine exposure causes hippocampus-dependent memory impairments in adolescent mice: Association with altered nicotinic cholinergic modulation of LTP, but not impaired LTP.

Fetal nicotine exposure from smoking during pregnancy causes long-lasting cognitive impairments in offspring, yet little is known about the mechanisms that underlie this effect. Here we demonstrate that early postnatal exposure of mouse pups to nicotine via maternal milk impairs long-term, but not short-term, hippocampus-dependent memory during adolescence. At the Schaffer collateral (SC) pathway, the most widely studied synapses for a cellular correlate of hippocampus-dependent memory, the induction of N-methyl-D-aspartate receptor-dependent transient long-term potentiation (LTP) and protein synthesis-dependent long-lasting LTP are not diminished by nicotine exposure, but rather unexpectedly the threshold for LTP induction becomes lower after nicotine treatment. Using voltage sensitive dye to visualize hippocampal activity, we found that early postnatal nicotine exposure also results in enhanced CA1 depolarization and hyperpolarization after SC stimulation. Furthermore, we show that postnatal nicotine exposure induces pervasive changes to the nicotinic modulation of CA1 activity: activation of nicotinic receptors no longer increases CA1 network depolarization, acute nicotine inhibits rather than facilitates the induction of LTP at the SC pathway by recruiting an additional nicotinic receptor subtype, and acute nicotine no longer blocks LTP induction at the temporoammonic pathway. These findings reflect the pervasive impact of nicotine exposure during hippocampal development, and demonstrate an association of hippocampal memory impairments with altered nicotinic cholinergic modulation of LTP, but not impaired LTP. The implication of our results is that nicotinic cholinergic-dependent plasticity is required for long-term memory formation and that postnatal nicotine exposure disrupts this form of plasticity.