β-Adrenergic Receptors Regulate the Acquisition and Consolidation Phases of Aversive Memory Formation Through Distinct, Temporally Regulated Signaling Pathways

Abstract

Memory formation requires the temporal coordination of molecular events and cellular processes following a learned event. During Pavlovian threat (fear) conditioning (PTC), sensory and neuromodulatory inputs converge on post-synaptic neurons within the lateral nucleus of the amygdala (LA). By activating an intracellular cascade of signaling molecules, these G-protein-coupled neuromodulatory receptors are capable of recruiting a diverse profile of plasticity-related proteins. Here we report that norepinephrine, through its actions on β-adrenergic receptors (βARs), modulates aversive memory formation following PTC through two molecularly and temporally distinct signaling mechanisms. Specifically, using behavioral pharmacology and biochemistry in adult rats, we determined that βAR activity during, but not after PTC training initiates the activation of two plasticity-related targets: AMPA receptors (AMPARs) for memory acquisition and short-term memory and extracellular regulated kinase (ERK) for consolidating the learned association into a long-term memory. These findings reveal that βAR activity during, but not following PTC sets in motion cascading molecular events for the acquisition (AMPARs) and subsequent consolidation (ERK) of learned associations.

Figure 1

Pretraining, but not post-training activation of βAR with ISO enhances memory formation for PTC. (a) Intra-LA ISO administration 15 min before training enhances freezing responses to the CS at short-term memory measured 3 h after training and at long-term memory measured 48 h after training. Enhanced freezing to the CS is also evident during the third CS of the training session. STM ISO (n=8) and vehicle (n=9); LTM ISO (n=9) and vehicle (n=12). (b) Post-training ISO administration does not affect freezing behavior. ISO (n=8) and vehicle (n=8). Error bars indicate±SEM. *p<0.05.

Figure 2

βAR activation leads to GluA1 phosphorylation at S845. (a) Bilateral intra-LA infusion of ISO (n=4) increases GluA1 phosphorylation at S845 compared with vehicle (n=3). All values are normalized to the loading control α-tubulin. ISO values are presented as a percent increase from the control condition. (b) Bilateral intra-LA ISO administration enhances GluA1 S845 phosphorylation in all animals independent of learning 5 min after weak PTC training. Training for weak PTC does not induce GluA1 S845 phosphorylation. For 5 min, vehicle box (n=9), vehicle conditioning (n=12), ISO box (n=12), ISO conditioning (n=10). Insets show sample bands from each condition. Error bars represent±SEM. *p<0.05.

Figure 3

Enhanced short-term and long-term memory by ISO administration depends on CP-AMPARs. (a) Timeline of experimental procedures. (b) Animals treated with ISO alone (n=13) show significantly enhanced freezing at STM measured 3 h after training compared with vehicle or ISO+NASPM: vehicle (n=9), ISO+NASPM (n=12), NASPM alone (n=11). (c) Animals treated with ISO alone (n=12) show significantly enhanced freezing at LTM measured 48 h after training compared with vehicle or ISO+NASPM: vehicle (n=8), ISO+NASPM (n=12), NASPM alone (n=8). Error bars represent±SEM. *p<0.05.

Figure 4

Enhanced long-term memory by ISO administration depends on ERK activity during consolidation. (a) Timeline of experimental procedures (left panel). For short-term memory test 3 h after PTC training (middle panel), animals treated with ISO (n=10) or ISO+SL327 (n=13) show significantly enhanced freezing compared with animals treated with vehicle (n=11). For long-term memory test 48 h after PTC training (right panel), animals treated with ISO (n=14) show significantly enhanced freezing compared to all other groups. Animals treated with vehicle (n=8), SL327 (n=11), or SL327+ISO (n=10) display equivalent freezing levels. (b) Enhanced freezing due to pretraining ISO administration (n=9) is prevented by post-training administration of an ERK inhibitor (n=6). Error bars represent±SEM. *p<0.05.