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Astrocytic adenosine receptor A2A and Gs-coupled signaling regulate memory.

Orr AG, Hsiao EC, Wang MM, Ho K, Kim DH, Wang X, Guo W, Kang J, Yu GQ, Adame A, Devidze N, Dubal DB, Masliah E, Conklin BR, Mucke L - Nat. Neurosci. (2015)

Bottom Line: Astrocytes express a variety of G protein-coupled receptors and might influence cognitive functions, such as learning and memory.However, the roles of astrocytic Gs-coupled receptors in cognitive function are not known.Together, these findings establish a regulatory role for astrocytic Gs-coupled receptors in memory and suggest that AD-linked increases in astrocytic A2A receptor levels contribute to memory loss.

View Article: PubMed Central - PubMed

Affiliation: 1] Gladstone Institute of Neurological Disease, San Francisco, California, USA. [2] Department of Neurology, University of California, San Francisco, California, USA.

ABSTRACT
Astrocytes express a variety of G protein-coupled receptors and might influence cognitive functions, such as learning and memory. However, the roles of astrocytic Gs-coupled receptors in cognitive function are not known. We found that humans with Alzheimer's disease (AD) had increased levels of the Gs-coupled adenosine receptor A2A in astrocytes. Conditional genetic removal of these receptors enhanced long-term memory in young and aging mice and increased the levels of Arc (also known as Arg3.1), an immediate-early gene that is required for long-term memory. Chemogenetic activation of astrocytic Gs-coupled signaling reduced long-term memory in mice without affecting learning. Like humans with AD, aging mice expressing human amyloid precursor protein (hAPP) showed increased levels of astrocytic A2A receptors. Conditional genetic removal of these receptors enhanced memory in aging hAPP mice. Together, these findings establish a regulatory role for astrocytic Gs-coupled receptors in memory and suggest that AD-linked increases in astrocytic A2A receptor levels contribute to memory loss.

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Conditional ablation of A2A receptors reduces memory deficits in aging hAPP mice(a– b) Representative photomicrographs (a) and quantification (b) of A2A receptor (green) immunostaining in hippocampal sections from 15–18-month-old loxP-Adora2a control mice (Con), hAPP/loxP-Adora2a mice (hAPP/loxP-A2A), and hAPP/loxP-Adora2a/Cre mice (hAPP/A2A-cKO). GFAP immunostaining is shown in red. Insets (i–iii) in (a) show magnified views of the boxed regions. DGgl: dentate gyrus granular layer. n = 7 Con, 6 hAPP/loxP-A2A, and 5 hAPP/A2A-cKO mice. Scale bar: 50 μm. (b) Quantification A2A receptor immunoreactivity in the hilus of the dentate gyrus. Levels were normalized to the average level in Con mice. One-way ANOVA: F(2, 17) = 6.52, P = 0.0091. n = 7 Con, 6 hAPP and 5 hAPP/A2A-cKO mice. *P < 0.05, **P < 0.01 vs. hAPP (Bonferroni test). (c–f)hAPP mice with or without conditional ablation of astrocytic A2A receptors were tested in the Morris water maze (c–e) and a habituation paradigm involving repeated exposure to the same open field (f) at 15–17 months of age. (c) Distance traveled to reach the platform during hidden platform training (two trials per session, two sessions per day for seven days). Two-way ANOVA revealed a significant difference between control (Con, singly transgenic loxP-A2A mice) and hAPP mice (F(1, 21) = 9.18, P = 0.0064), but not between hAPP and hAPP/A2A-cKO mice (F(1, 16) = 0.01, P = 0.90). n = 14 Con, 9 hAPP, and 9 hAPP/A2A-cKO mice. (d) Probe trial conducted one day after training in the Morris water maze. Left: Durations in target and non-target (other) quadrants. One-way ANOVA: F(2, 28) = 5.39, P = 0.0104; one-sample t-test vs. chance duration of 15 s with FDR correction for multiple comparisons (Target vs. chance): P = 0.0006 (Con), P = 0.618 (hAPP), P = 0.0054 (hAPP/A2A-cKO). n = 14 Con, 8 hAPP, and 9 hAPP/A2A-cKO mice. ##P < 0.01 vs. Con (Dunnett’s test); **P < 0.01, ***P < 0.001 (one-sample t-test). Right: Latency to reach target location. One-way ANOVA: F(2, 28) = 6.30, P = 0.0055. **P < 0.01 vs. Con (Dunnett’s test). n = 14 Con, 8 hAPP, and 9 hAPP/A2A-cKO mice. (e) Swim speeds during the probe trial. One-way ANOVA: F(2, 28) = 2.28, P = 0.12. P > 0.05 vs. Con (Dunnett’s test). n = 14 Con, 8 hAPP, and 9 hAPP/A2A-cKO mice. (f) Open-field habituation. Left: Mice were habituated to the open field in 5-min trials (1–2 trials per day) for two days and tested in the same arena on days 16 and 58. Repeated measures two-way ANOVA: F(2, 27) = 5.17, P = 0.0125 for genotype effect, F(5, 135) = 11.25, P < 0.0001 for time effect, F(10, 135) = 1.25, P = 0.27 for interaction between genotype and time. Right: Extent of dishabituation on day 58 (test trial 6 relative to trial 1). One-way ANOVA: F(2, 27) = 7.94, P = 0.0019. n = 14 Con, 8 hAPP, and 8 hAPP/A2A-cKO mice. #P < 0.05, ##P < 0.01 vs. Con (Dunnett’s test). Values are means ± s.e.m.
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Figure 8: Conditional ablation of A2A receptors reduces memory deficits in aging hAPP mice(a– b) Representative photomicrographs (a) and quantification (b) of A2A receptor (green) immunostaining in hippocampal sections from 15–18-month-old loxP-Adora2a control mice (Con), hAPP/loxP-Adora2a mice (hAPP/loxP-A2A), and hAPP/loxP-Adora2a/Cre mice (hAPP/A2A-cKO). GFAP immunostaining is shown in red. Insets (i–iii) in (a) show magnified views of the boxed regions. DGgl: dentate gyrus granular layer. n = 7 Con, 6 hAPP/loxP-A2A, and 5 hAPP/A2A-cKO mice. Scale bar: 50 μm. (b) Quantification A2A receptor immunoreactivity in the hilus of the dentate gyrus. Levels were normalized to the average level in Con mice. One-way ANOVA: F(2, 17) = 6.52, P = 0.0091. n = 7 Con, 6 hAPP and 5 hAPP/A2A-cKO mice. *P < 0.05, **P < 0.01 vs. hAPP (Bonferroni test). (c–f)hAPP mice with or without conditional ablation of astrocytic A2A receptors were tested in the Morris water maze (c–e) and a habituation paradigm involving repeated exposure to the same open field (f) at 15–17 months of age. (c) Distance traveled to reach the platform during hidden platform training (two trials per session, two sessions per day for seven days). Two-way ANOVA revealed a significant difference between control (Con, singly transgenic loxP-A2A mice) and hAPP mice (F(1, 21) = 9.18, P = 0.0064), but not between hAPP and hAPP/A2A-cKO mice (F(1, 16) = 0.01, P = 0.90). n = 14 Con, 9 hAPP, and 9 hAPP/A2A-cKO mice. (d) Probe trial conducted one day after training in the Morris water maze. Left: Durations in target and non-target (other) quadrants. One-way ANOVA: F(2, 28) = 5.39, P = 0.0104; one-sample t-test vs. chance duration of 15 s with FDR correction for multiple comparisons (Target vs. chance): P = 0.0006 (Con), P = 0.618 (hAPP), P = 0.0054 (hAPP/A2A-cKO). n = 14 Con, 8 hAPP, and 9 hAPP/A2A-cKO mice. ##P < 0.01 vs. Con (Dunnett’s test); **P < 0.01, ***P < 0.001 (one-sample t-test). Right: Latency to reach target location. One-way ANOVA: F(2, 28) = 6.30, P = 0.0055. **P < 0.01 vs. Con (Dunnett’s test). n = 14 Con, 8 hAPP, and 9 hAPP/A2A-cKO mice. (e) Swim speeds during the probe trial. One-way ANOVA: F(2, 28) = 2.28, P = 0.12. P > 0.05 vs. Con (Dunnett’s test). n = 14 Con, 8 hAPP, and 9 hAPP/A2A-cKO mice. (f) Open-field habituation. Left: Mice were habituated to the open field in 5-min trials (1–2 trials per day) for two days and tested in the same arena on days 16 and 58. Repeated measures two-way ANOVA: F(2, 27) = 5.17, P = 0.0125 for genotype effect, F(5, 135) = 11.25, P < 0.0001 for time effect, F(10, 135) = 1.25, P = 0.27 for interaction between genotype and time. Right: Extent of dishabituation on day 58 (test trial 6 relative to trial 1). One-way ANOVA: F(2, 27) = 7.94, P = 0.0019. n = 14 Con, 8 hAPP, and 8 hAPP/A2A-cKO mice. #P < 0.05, ##P < 0.01 vs. Con (Dunnett’s test). Values are means ± s.e.m.

Mentions: We next tested if astrocytic A2A receptors contribute to memory deficits in aging hAPP mice. We generated transgenic mice expressing hAPP with or without conditional ablation of astrocytic A2A receptors (Fig. 8a–b) and tested these mice in the Morris water maze and the open field. Specifically, we tested three groups of transgenic mice: singly transgenic control mice carrying two loxP-Adora2a alleles, doubly transgenic hAPP mice carrying human APP and one or two loxP-Adora2a alleles, and triply transgenic hAPP/A2A-cKO mice carrying human APP, two loxP-Adora2a alleles and Cre recombinase. Ablation of astrocytic A2A receptors did not affect the learning deficits of 15–17-month-old hAPP mice during hidden platform training (Fig. 8c), consistent with our observations that ablation of these receptors does not affect learning in young and aging mice without hAPP expression (Fig. 2). Notably, ablation of astrocytic A2A receptors enhanced the performance of hAPP mice in a probe trial 24 h after training (Fig. 8d). Specifically, hAPP mice bearing astrocytic A2A receptors showed a minimal preference for the target quadrant and an increased latency to reach the platform location compared with singly transgenic control mice without hAPP expression. In contrast, hAPP/A2A-cKO mice showed a significant preference for the target quadrant and a lower latency to reach the platform location. Swim speeds during the probe trial were comparable among the groups (Fig. 8e). Moreover, hAPP/A2A-KO mice showed less dishabituation in the open field than hAPP mice and singly transgenic controls (Fig. 8f), consistent with our findings in A2A-cHET and A2A-cKO mice without hAPP expression (Fig. 2c–f).


Astrocytic adenosine receptor A2A and Gs-coupled signaling regulate memory.

Orr AG, Hsiao EC, Wang MM, Ho K, Kim DH, Wang X, Guo W, Kang J, Yu GQ, Adame A, Devidze N, Dubal DB, Masliah E, Conklin BR, Mucke L - Nat. Neurosci. (2015)

Conditional ablation of A2A receptors reduces memory deficits in aging hAPP mice(a– b) Representative photomicrographs (a) and quantification (b) of A2A receptor (green) immunostaining in hippocampal sections from 15–18-month-old loxP-Adora2a control mice (Con), hAPP/loxP-Adora2a mice (hAPP/loxP-A2A), and hAPP/loxP-Adora2a/Cre mice (hAPP/A2A-cKO). GFAP immunostaining is shown in red. Insets (i–iii) in (a) show magnified views of the boxed regions. DGgl: dentate gyrus granular layer. n = 7 Con, 6 hAPP/loxP-A2A, and 5 hAPP/A2A-cKO mice. Scale bar: 50 μm. (b) Quantification A2A receptor immunoreactivity in the hilus of the dentate gyrus. Levels were normalized to the average level in Con mice. One-way ANOVA: F(2, 17) = 6.52, P = 0.0091. n = 7 Con, 6 hAPP and 5 hAPP/A2A-cKO mice. *P < 0.05, **P < 0.01 vs. hAPP (Bonferroni test). (c–f)hAPP mice with or without conditional ablation of astrocytic A2A receptors were tested in the Morris water maze (c–e) and a habituation paradigm involving repeated exposure to the same open field (f) at 15–17 months of age. (c) Distance traveled to reach the platform during hidden platform training (two trials per session, two sessions per day for seven days). Two-way ANOVA revealed a significant difference between control (Con, singly transgenic loxP-A2A mice) and hAPP mice (F(1, 21) = 9.18, P = 0.0064), but not between hAPP and hAPP/A2A-cKO mice (F(1, 16) = 0.01, P = 0.90). n = 14 Con, 9 hAPP, and 9 hAPP/A2A-cKO mice. (d) Probe trial conducted one day after training in the Morris water maze. Left: Durations in target and non-target (other) quadrants. One-way ANOVA: F(2, 28) = 5.39, P = 0.0104; one-sample t-test vs. chance duration of 15 s with FDR correction for multiple comparisons (Target vs. chance): P = 0.0006 (Con), P = 0.618 (hAPP), P = 0.0054 (hAPP/A2A-cKO). n = 14 Con, 8 hAPP, and 9 hAPP/A2A-cKO mice. ##P < 0.01 vs. Con (Dunnett’s test); **P < 0.01, ***P < 0.001 (one-sample t-test). Right: Latency to reach target location. One-way ANOVA: F(2, 28) = 6.30, P = 0.0055. **P < 0.01 vs. Con (Dunnett’s test). n = 14 Con, 8 hAPP, and 9 hAPP/A2A-cKO mice. (e) Swim speeds during the probe trial. One-way ANOVA: F(2, 28) = 2.28, P = 0.12. P > 0.05 vs. Con (Dunnett’s test). n = 14 Con, 8 hAPP, and 9 hAPP/A2A-cKO mice. (f) Open-field habituation. Left: Mice were habituated to the open field in 5-min trials (1–2 trials per day) for two days and tested in the same arena on days 16 and 58. Repeated measures two-way ANOVA: F(2, 27) = 5.17, P = 0.0125 for genotype effect, F(5, 135) = 11.25, P < 0.0001 for time effect, F(10, 135) = 1.25, P = 0.27 for interaction between genotype and time. Right: Extent of dishabituation on day 58 (test trial 6 relative to trial 1). One-way ANOVA: F(2, 27) = 7.94, P = 0.0019. n = 14 Con, 8 hAPP, and 8 hAPP/A2A-cKO mice. #P < 0.05, ##P < 0.01 vs. Con (Dunnett’s test). Values are means ± s.e.m.
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Figure 8: Conditional ablation of A2A receptors reduces memory deficits in aging hAPP mice(a– b) Representative photomicrographs (a) and quantification (b) of A2A receptor (green) immunostaining in hippocampal sections from 15–18-month-old loxP-Adora2a control mice (Con), hAPP/loxP-Adora2a mice (hAPP/loxP-A2A), and hAPP/loxP-Adora2a/Cre mice (hAPP/A2A-cKO). GFAP immunostaining is shown in red. Insets (i–iii) in (a) show magnified views of the boxed regions. DGgl: dentate gyrus granular layer. n = 7 Con, 6 hAPP/loxP-A2A, and 5 hAPP/A2A-cKO mice. Scale bar: 50 μm. (b) Quantification A2A receptor immunoreactivity in the hilus of the dentate gyrus. Levels were normalized to the average level in Con mice. One-way ANOVA: F(2, 17) = 6.52, P = 0.0091. n = 7 Con, 6 hAPP and 5 hAPP/A2A-cKO mice. *P < 0.05, **P < 0.01 vs. hAPP (Bonferroni test). (c–f)hAPP mice with or without conditional ablation of astrocytic A2A receptors were tested in the Morris water maze (c–e) and a habituation paradigm involving repeated exposure to the same open field (f) at 15–17 months of age. (c) Distance traveled to reach the platform during hidden platform training (two trials per session, two sessions per day for seven days). Two-way ANOVA revealed a significant difference between control (Con, singly transgenic loxP-A2A mice) and hAPP mice (F(1, 21) = 9.18, P = 0.0064), but not between hAPP and hAPP/A2A-cKO mice (F(1, 16) = 0.01, P = 0.90). n = 14 Con, 9 hAPP, and 9 hAPP/A2A-cKO mice. (d) Probe trial conducted one day after training in the Morris water maze. Left: Durations in target and non-target (other) quadrants. One-way ANOVA: F(2, 28) = 5.39, P = 0.0104; one-sample t-test vs. chance duration of 15 s with FDR correction for multiple comparisons (Target vs. chance): P = 0.0006 (Con), P = 0.618 (hAPP), P = 0.0054 (hAPP/A2A-cKO). n = 14 Con, 8 hAPP, and 9 hAPP/A2A-cKO mice. ##P < 0.01 vs. Con (Dunnett’s test); **P < 0.01, ***P < 0.001 (one-sample t-test). Right: Latency to reach target location. One-way ANOVA: F(2, 28) = 6.30, P = 0.0055. **P < 0.01 vs. Con (Dunnett’s test). n = 14 Con, 8 hAPP, and 9 hAPP/A2A-cKO mice. (e) Swim speeds during the probe trial. One-way ANOVA: F(2, 28) = 2.28, P = 0.12. P > 0.05 vs. Con (Dunnett’s test). n = 14 Con, 8 hAPP, and 9 hAPP/A2A-cKO mice. (f) Open-field habituation. Left: Mice were habituated to the open field in 5-min trials (1–2 trials per day) for two days and tested in the same arena on days 16 and 58. Repeated measures two-way ANOVA: F(2, 27) = 5.17, P = 0.0125 for genotype effect, F(5, 135) = 11.25, P < 0.0001 for time effect, F(10, 135) = 1.25, P = 0.27 for interaction between genotype and time. Right: Extent of dishabituation on day 58 (test trial 6 relative to trial 1). One-way ANOVA: F(2, 27) = 7.94, P = 0.0019. n = 14 Con, 8 hAPP, and 8 hAPP/A2A-cKO mice. #P < 0.05, ##P < 0.01 vs. Con (Dunnett’s test). Values are means ± s.e.m.
Mentions: We next tested if astrocytic A2A receptors contribute to memory deficits in aging hAPP mice. We generated transgenic mice expressing hAPP with or without conditional ablation of astrocytic A2A receptors (Fig. 8a–b) and tested these mice in the Morris water maze and the open field. Specifically, we tested three groups of transgenic mice: singly transgenic control mice carrying two loxP-Adora2a alleles, doubly transgenic hAPP mice carrying human APP and one or two loxP-Adora2a alleles, and triply transgenic hAPP/A2A-cKO mice carrying human APP, two loxP-Adora2a alleles and Cre recombinase. Ablation of astrocytic A2A receptors did not affect the learning deficits of 15–17-month-old hAPP mice during hidden platform training (Fig. 8c), consistent with our observations that ablation of these receptors does not affect learning in young and aging mice without hAPP expression (Fig. 2). Notably, ablation of astrocytic A2A receptors enhanced the performance of hAPP mice in a probe trial 24 h after training (Fig. 8d). Specifically, hAPP mice bearing astrocytic A2A receptors showed a minimal preference for the target quadrant and an increased latency to reach the platform location compared with singly transgenic control mice without hAPP expression. In contrast, hAPP/A2A-cKO mice showed a significant preference for the target quadrant and a lower latency to reach the platform location. Swim speeds during the probe trial were comparable among the groups (Fig. 8e). Moreover, hAPP/A2A-KO mice showed less dishabituation in the open field than hAPP mice and singly transgenic controls (Fig. 8f), consistent with our findings in A2A-cHET and A2A-cKO mice without hAPP expression (Fig. 2c–f).

Bottom Line: Astrocytes express a variety of G protein-coupled receptors and might influence cognitive functions, such as learning and memory.However, the roles of astrocytic Gs-coupled receptors in cognitive function are not known.Together, these findings establish a regulatory role for astrocytic Gs-coupled receptors in memory and suggest that AD-linked increases in astrocytic A2A receptor levels contribute to memory loss.

View Article: PubMed Central - PubMed

Affiliation: 1] Gladstone Institute of Neurological Disease, San Francisco, California, USA. [2] Department of Neurology, University of California, San Francisco, California, USA.

ABSTRACT
Astrocytes express a variety of G protein-coupled receptors and might influence cognitive functions, such as learning and memory. However, the roles of astrocytic Gs-coupled receptors in cognitive function are not known. We found that humans with Alzheimer's disease (AD) had increased levels of the Gs-coupled adenosine receptor A2A in astrocytes. Conditional genetic removal of these receptors enhanced long-term memory in young and aging mice and increased the levels of Arc (also known as Arg3.1), an immediate-early gene that is required for long-term memory. Chemogenetic activation of astrocytic Gs-coupled signaling reduced long-term memory in mice without affecting learning. Like humans with AD, aging mice expressing human amyloid precursor protein (hAPP) showed increased levels of astrocytic A2A receptors. Conditional genetic removal of these receptors enhanced memory in aging hAPP mice. Together, these findings establish a regulatory role for astrocytic Gs-coupled receptors in memory and suggest that AD-linked increases in astrocytic A2A receptor levels contribute to memory loss.

Show MeSH
Related in: MedlinePlus