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pT305-CaMKII stabilizes a learning-induced increase in AMPA receptors for ongoing memory consolidation after classical conditioning.

Naskar S, Wan H, Kemenes G - Nat Commun (2014)

Bottom Line: CaMKIINtide treatment significantly reduces the learning-induced elevation of both pT305-CaMKII and GluA1 levels and impairs associative long-term memory.Inhibition of proteasomal activity offsets the deleterious effects of CaMKIINtide on both GluA1 levels and long-term memory.These findings suggest that increased levels of pT305-CaMKII play a role in AMPAR-dependent memory consolidation by reducing proteasomal degradation of GluA1 receptor subunits.

View Article: PubMed Central - PubMed

Affiliation: 1] Sussex Neuroscience, School of Life Sciences, University of Sussex, Brighton BN1 9QG, UK [2].

ABSTRACT
The role of CaMKII in learning-induced activation and trafficking of AMPA receptors (AMPARs) is well established. However, the link between the phosphorylation state of CaMKII and the agonist-triggered proteasomal degradation of AMPARs during memory consolidation remains unknown. Here we describe a novel CaMKII-dependent mechanism by which a learning-induced increase in AMPAR levels is stabilized for consolidation of associative long-term memory. Six hours after classical conditioning the levels of both autophosphorylated pT305-CaMKII and GluA1 type AMPAR subunits are significantly elevated in the ganglia containing the learning circuits of the snail Lymnaea stagnalis. CaMKIINtide treatment significantly reduces the learning-induced elevation of both pT305-CaMKII and GluA1 levels and impairs associative long-term memory. Inhibition of proteasomal activity offsets the deleterious effects of CaMKIINtide on both GluA1 levels and long-term memory. These findings suggest that increased levels of pT305-CaMKII play a role in AMPAR-dependent memory consolidation by reducing proteasomal degradation of GluA1 receptor subunits.

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pT305-CaMKII and memory consolidation in Lymnaea(a) Timeline for the behavioral experiment (conducted concurrently with the biochemical assays shown in b and c) is shown above the diagram. The classically conditioned and CaMKIINtide-treated animals (N=23) as well as the Naïve animals (N=16) showed a significantly lower (asterisks) feeding response to the CS compared to the classically conditioned vehicle-treated animals (N=21). One-way ANOVA: P<0.0001. Tukey’s tests: CaMKIINtide and Naive versus Vehicle, both P<0.05, CaMKIINtide versus Naïve, P>0.05. (b, c) The selective effects of CaMKIINtide on T305 phosphorylation of CaMKII. Examples of western blot bands are shown above the graphs (full-length blots in Supplementary Fig. 7d and e). The means (±SEM) of pT305-CaMKII (b) or pT286-CaMKII (c) over CaMKII levels are shown (relative integrated density). Total CaMKII levels were not significantly different among the groups. The values thus calculated for each CaMKIINtide treated (b, N=7; c, N=7) and vehicle treated sample (b, N=6; c, N=6) were normalized to the mean of the density values obtained from the naïve samples (b, N=14; c, N=12), providing a baseline of 1 (dashed line). Vehicle treatment 6 h after paired training leaves the significantly raised level of pT305-CaMKII (see Fig. 6a) unaffected, whereas CaMKIINtide treatment significantly decreases it (b). Asterisks indicate significant differences both compared to the Naïve baseline and between the two paired groups. One-sample t tests: CaMKIINtide and Vehicle versus baseline, both P<0.03. Two-sample t-test: CaMKIINtide versus Vehicle, P<0.002). T286 phosphorylation remains unaffected by treatment with either vehicle or CaMKIINtide (c). One-sample t-tests: CaMKIINtide and Vehicle versus baseline, P=0.70 and 0.69, respectively. Two-sample t-test: CaMKIINtide vs. Vehicle, P=0.90). These experiments were replicated twice.
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Figure 7: pT305-CaMKII and memory consolidation in Lymnaea(a) Timeline for the behavioral experiment (conducted concurrently with the biochemical assays shown in b and c) is shown above the diagram. The classically conditioned and CaMKIINtide-treated animals (N=23) as well as the Naïve animals (N=16) showed a significantly lower (asterisks) feeding response to the CS compared to the classically conditioned vehicle-treated animals (N=21). One-way ANOVA: P<0.0001. Tukey’s tests: CaMKIINtide and Naive versus Vehicle, both P<0.05, CaMKIINtide versus Naïve, P>0.05. (b, c) The selective effects of CaMKIINtide on T305 phosphorylation of CaMKII. Examples of western blot bands are shown above the graphs (full-length blots in Supplementary Fig. 7d and e). The means (±SEM) of pT305-CaMKII (b) or pT286-CaMKII (c) over CaMKII levels are shown (relative integrated density). Total CaMKII levels were not significantly different among the groups. The values thus calculated for each CaMKIINtide treated (b, N=7; c, N=7) and vehicle treated sample (b, N=6; c, N=6) were normalized to the mean of the density values obtained from the naïve samples (b, N=14; c, N=12), providing a baseline of 1 (dashed line). Vehicle treatment 6 h after paired training leaves the significantly raised level of pT305-CaMKII (see Fig. 6a) unaffected, whereas CaMKIINtide treatment significantly decreases it (b). Asterisks indicate significant differences both compared to the Naïve baseline and between the two paired groups. One-sample t tests: CaMKIINtide and Vehicle versus baseline, both P<0.03. Two-sample t-test: CaMKIINtide versus Vehicle, P<0.002). T286 phosphorylation remains unaffected by treatment with either vehicle or CaMKIINtide (c). One-sample t-tests: CaMKIINtide and Vehicle versus baseline, P=0.70 and 0.69, respectively. Two-sample t-test: CaMKIINtide vs. Vehicle, P=0.90). These experiments were replicated twice.

Mentions: Next, the necessity for an elevated level of pT305-CaMKII in the consolidation of long-term memory was investigated in an experiment using three groups of animals: paired and CaMKIINtide-treated, paired and vehicle-treated and naïve (Fig. 7a). It was not necessary to use an unpaired group in this experiment because our previous experiments showed that animals in this control group did not respond to the CS and therefore the link between pharmacological treatments and inhibition of the conditioned response cannot be studied in them.


pT305-CaMKII stabilizes a learning-induced increase in AMPA receptors for ongoing memory consolidation after classical conditioning.

Naskar S, Wan H, Kemenes G - Nat Commun (2014)

pT305-CaMKII and memory consolidation in Lymnaea(a) Timeline for the behavioral experiment (conducted concurrently with the biochemical assays shown in b and c) is shown above the diagram. The classically conditioned and CaMKIINtide-treated animals (N=23) as well as the Naïve animals (N=16) showed a significantly lower (asterisks) feeding response to the CS compared to the classically conditioned vehicle-treated animals (N=21). One-way ANOVA: P<0.0001. Tukey’s tests: CaMKIINtide and Naive versus Vehicle, both P<0.05, CaMKIINtide versus Naïve, P>0.05. (b, c) The selective effects of CaMKIINtide on T305 phosphorylation of CaMKII. Examples of western blot bands are shown above the graphs (full-length blots in Supplementary Fig. 7d and e). The means (±SEM) of pT305-CaMKII (b) or pT286-CaMKII (c) over CaMKII levels are shown (relative integrated density). Total CaMKII levels were not significantly different among the groups. The values thus calculated for each CaMKIINtide treated (b, N=7; c, N=7) and vehicle treated sample (b, N=6; c, N=6) were normalized to the mean of the density values obtained from the naïve samples (b, N=14; c, N=12), providing a baseline of 1 (dashed line). Vehicle treatment 6 h after paired training leaves the significantly raised level of pT305-CaMKII (see Fig. 6a) unaffected, whereas CaMKIINtide treatment significantly decreases it (b). Asterisks indicate significant differences both compared to the Naïve baseline and between the two paired groups. One-sample t tests: CaMKIINtide and Vehicle versus baseline, both P<0.03. Two-sample t-test: CaMKIINtide versus Vehicle, P<0.002). T286 phosphorylation remains unaffected by treatment with either vehicle or CaMKIINtide (c). One-sample t-tests: CaMKIINtide and Vehicle versus baseline, P=0.70 and 0.69, respectively. Two-sample t-test: CaMKIINtide vs. Vehicle, P=0.90). These experiments were replicated twice.
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Related In: Results  -  Collection

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Figure 7: pT305-CaMKII and memory consolidation in Lymnaea(a) Timeline for the behavioral experiment (conducted concurrently with the biochemical assays shown in b and c) is shown above the diagram. The classically conditioned and CaMKIINtide-treated animals (N=23) as well as the Naïve animals (N=16) showed a significantly lower (asterisks) feeding response to the CS compared to the classically conditioned vehicle-treated animals (N=21). One-way ANOVA: P<0.0001. Tukey’s tests: CaMKIINtide and Naive versus Vehicle, both P<0.05, CaMKIINtide versus Naïve, P>0.05. (b, c) The selective effects of CaMKIINtide on T305 phosphorylation of CaMKII. Examples of western blot bands are shown above the graphs (full-length blots in Supplementary Fig. 7d and e). The means (±SEM) of pT305-CaMKII (b) or pT286-CaMKII (c) over CaMKII levels are shown (relative integrated density). Total CaMKII levels were not significantly different among the groups. The values thus calculated for each CaMKIINtide treated (b, N=7; c, N=7) and vehicle treated sample (b, N=6; c, N=6) were normalized to the mean of the density values obtained from the naïve samples (b, N=14; c, N=12), providing a baseline of 1 (dashed line). Vehicle treatment 6 h after paired training leaves the significantly raised level of pT305-CaMKII (see Fig. 6a) unaffected, whereas CaMKIINtide treatment significantly decreases it (b). Asterisks indicate significant differences both compared to the Naïve baseline and between the two paired groups. One-sample t tests: CaMKIINtide and Vehicle versus baseline, both P<0.03. Two-sample t-test: CaMKIINtide versus Vehicle, P<0.002). T286 phosphorylation remains unaffected by treatment with either vehicle or CaMKIINtide (c). One-sample t-tests: CaMKIINtide and Vehicle versus baseline, P=0.70 and 0.69, respectively. Two-sample t-test: CaMKIINtide vs. Vehicle, P=0.90). These experiments were replicated twice.
Mentions: Next, the necessity for an elevated level of pT305-CaMKII in the consolidation of long-term memory was investigated in an experiment using three groups of animals: paired and CaMKIINtide-treated, paired and vehicle-treated and naïve (Fig. 7a). It was not necessary to use an unpaired group in this experiment because our previous experiments showed that animals in this control group did not respond to the CS and therefore the link between pharmacological treatments and inhibition of the conditioned response cannot be studied in them.

Bottom Line: CaMKIINtide treatment significantly reduces the learning-induced elevation of both pT305-CaMKII and GluA1 levels and impairs associative long-term memory.Inhibition of proteasomal activity offsets the deleterious effects of CaMKIINtide on both GluA1 levels and long-term memory.These findings suggest that increased levels of pT305-CaMKII play a role in AMPAR-dependent memory consolidation by reducing proteasomal degradation of GluA1 receptor subunits.

View Article: PubMed Central - PubMed

Affiliation: 1] Sussex Neuroscience, School of Life Sciences, University of Sussex, Brighton BN1 9QG, UK [2].

ABSTRACT
The role of CaMKII in learning-induced activation and trafficking of AMPA receptors (AMPARs) is well established. However, the link between the phosphorylation state of CaMKII and the agonist-triggered proteasomal degradation of AMPARs during memory consolidation remains unknown. Here we describe a novel CaMKII-dependent mechanism by which a learning-induced increase in AMPAR levels is stabilized for consolidation of associative long-term memory. Six hours after classical conditioning the levels of both autophosphorylated pT305-CaMKII and GluA1 type AMPAR subunits are significantly elevated in the ganglia containing the learning circuits of the snail Lymnaea stagnalis. CaMKIINtide treatment significantly reduces the learning-induced elevation of both pT305-CaMKII and GluA1 levels and impairs associative long-term memory. Inhibition of proteasomal activity offsets the deleterious effects of CaMKIINtide on both GluA1 levels and long-term memory. These findings suggest that increased levels of pT305-CaMKII play a role in AMPAR-dependent memory consolidation by reducing proteasomal degradation of GluA1 receptor subunits.

Show MeSH
Related in: MedlinePlus