Rapid brain-derived neurotrophic factor-dependent sequestration of amygdala and hippocampal GABAA receptors via different tyrosine receptor kinase B-mediated phosphorylation pathways

L. Mou, Scott Heldt, K. J. Ressler

Research output: Contribution to journalArticle

23 Citations (Scopus)

Abstract

During the consolidation of fear memory, it has been shown that GABAA receptors (GABAAR) are rapidly downregulated in amygdala. This rapid decrease in GABAAR functioning may permit transient hyperexcitablity, contributing to cellular mechanisms of memory consolidation. Memory consolidation also requires brain-derived neurotrophic factor (BDNF) activation of tyrosine receptor kinase B (TrkB) receptors in the amygdala and hippocampus. We hypothesized that rapid internalization of GABAARα1 is mediated via TrkB activation of PKA and PKC-dependent processes. Primary neuronal cell cultures, from postnatal day 14-21 mouse amygdala and hippocampus, were analyzed with immunofluorescence using cell-surface, whole-cell permeabilization, and antibody internalization techniques, as well as with 3H-muscimol binding assays. In both hippocampal and amygdala cultures, we found a >60% reduction in surface GABAARα1 within 5 min of BDNF treatment. Notably, the rapid decrease in surface GABAARα1 was confirmed biochemically using surface biotinylation assays followed by western blotting. This rapid effect was accompanied by TrkB phosphorylation and increased internal GABAARα1 immunofluorescence, and was blocked by k252a, a broad-spectrum tyrosine kinase antagonist. To further demonstrate TrkB specificity, we used previously characterized TrkBF616A mice, in which the highly selective TrkB-mutant specific antagonist, 1NMPP1, prevented the BDNF-dependent GABAARα1 internalization. In hippocampus, we found both PKA and PKC inhibition, using Rp-8-Br-cAMP and Calphostin C, respectively, blocked GABAARα1 internalization, whereas inhibition of MAPK (U0126) and PI3K (LY294002) did not prevent rapid internalization. By contrast in amygdala cultures, Rp-8-Br-cAMP had no effect. Together, these data suggest that rapid GABAAR internalization during memory consolidation is BDNF-TrkB dependent. Further, it appears that hippocampal GABAAR internalization is PKA and PKC dependent, while it may be primarily PKC dependent in amygdala, implying differential roles for TrkB-dependent kinase activation in BDNF-dependent memory formation.

Original languageEnglish (US)
Pages (from-to)72-85
Number of pages14
JournalNeuroscience
Volume176
DOIs
StatePublished - Mar 10 2011

Fingerprint

Brain-Derived Neurotrophic Factor
Receptor Protein-Tyrosine Kinases
GABA-A Receptors
Amygdala
Phosphorylation
Hippocampus
Protein-Tyrosine Kinases
Fluorescent Antibody Technique
trkB Receptor
Biotinylation
Muscimol
2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one
Primary Cell Culture
Phosphatidylinositol 3-Kinases
Fear
Phosphotransferases
Down-Regulation
Western Blotting
Memory Consolidation
Antibodies

All Science Journal Classification (ASJC) codes

  • Neuroscience(all)

Cite this

Rapid brain-derived neurotrophic factor-dependent sequestration of amygdala and hippocampal GABAA receptors via different tyrosine receptor kinase B-mediated phosphorylation pathways. / Mou, L.; Heldt, Scott; Ressler, K. J.

In: Neuroscience, Vol. 176, 10.03.2011, p. 72-85.

Research output: Contribution to journalArticle

@article{e018741f50a9467ab8553c9546996f01,
title = "Rapid brain-derived neurotrophic factor-dependent sequestration of amygdala and hippocampal GABAA receptors via different tyrosine receptor kinase B-mediated phosphorylation pathways",
abstract = "During the consolidation of fear memory, it has been shown that GABAA receptors (GABAAR) are rapidly downregulated in amygdala. This rapid decrease in GABAAR functioning may permit transient hyperexcitablity, contributing to cellular mechanisms of memory consolidation. Memory consolidation also requires brain-derived neurotrophic factor (BDNF) activation of tyrosine receptor kinase B (TrkB) receptors in the amygdala and hippocampus. We hypothesized that rapid internalization of GABAARα1 is mediated via TrkB activation of PKA and PKC-dependent processes. Primary neuronal cell cultures, from postnatal day 14-21 mouse amygdala and hippocampus, were analyzed with immunofluorescence using cell-surface, whole-cell permeabilization, and antibody internalization techniques, as well as with 3H-muscimol binding assays. In both hippocampal and amygdala cultures, we found a >60{\%} reduction in surface GABAARα1 within 5 min of BDNF treatment. Notably, the rapid decrease in surface GABAARα1 was confirmed biochemically using surface biotinylation assays followed by western blotting. This rapid effect was accompanied by TrkB phosphorylation and increased internal GABAARα1 immunofluorescence, and was blocked by k252a, a broad-spectrum tyrosine kinase antagonist. To further demonstrate TrkB specificity, we used previously characterized TrkBF616A mice, in which the highly selective TrkB-mutant specific antagonist, 1NMPP1, prevented the BDNF-dependent GABAARα1 internalization. In hippocampus, we found both PKA and PKC inhibition, using Rp-8-Br-cAMP and Calphostin C, respectively, blocked GABAARα1 internalization, whereas inhibition of MAPK (U0126) and PI3K (LY294002) did not prevent rapid internalization. By contrast in amygdala cultures, Rp-8-Br-cAMP had no effect. Together, these data suggest that rapid GABAAR internalization during memory consolidation is BDNF-TrkB dependent. Further, it appears that hippocampal GABAAR internalization is PKA and PKC dependent, while it may be primarily PKC dependent in amygdala, implying differential roles for TrkB-dependent kinase activation in BDNF-dependent memory formation.",
author = "L. Mou and Scott Heldt and Ressler, {K. J.}",
year = "2011",
month = "3",
day = "10",
doi = "10.1016/j.neuroscience.2010.12.041",
language = "English (US)",
volume = "176",
pages = "72--85",
journal = "Neuroscience",
issn = "0306-4522",
publisher = "Elsevier Limited",

}

TY - JOUR

T1 - Rapid brain-derived neurotrophic factor-dependent sequestration of amygdala and hippocampal GABAA receptors via different tyrosine receptor kinase B-mediated phosphorylation pathways

AU - Mou, L.

AU - Heldt, Scott

AU - Ressler, K. J.

PY - 2011/3/10

Y1 - 2011/3/10

N2 - During the consolidation of fear memory, it has been shown that GABAA receptors (GABAAR) are rapidly downregulated in amygdala. This rapid decrease in GABAAR functioning may permit transient hyperexcitablity, contributing to cellular mechanisms of memory consolidation. Memory consolidation also requires brain-derived neurotrophic factor (BDNF) activation of tyrosine receptor kinase B (TrkB) receptors in the amygdala and hippocampus. We hypothesized that rapid internalization of GABAARα1 is mediated via TrkB activation of PKA and PKC-dependent processes. Primary neuronal cell cultures, from postnatal day 14-21 mouse amygdala and hippocampus, were analyzed with immunofluorescence using cell-surface, whole-cell permeabilization, and antibody internalization techniques, as well as with 3H-muscimol binding assays. In both hippocampal and amygdala cultures, we found a >60% reduction in surface GABAARα1 within 5 min of BDNF treatment. Notably, the rapid decrease in surface GABAARα1 was confirmed biochemically using surface biotinylation assays followed by western blotting. This rapid effect was accompanied by TrkB phosphorylation and increased internal GABAARα1 immunofluorescence, and was blocked by k252a, a broad-spectrum tyrosine kinase antagonist. To further demonstrate TrkB specificity, we used previously characterized TrkBF616A mice, in which the highly selective TrkB-mutant specific antagonist, 1NMPP1, prevented the BDNF-dependent GABAARα1 internalization. In hippocampus, we found both PKA and PKC inhibition, using Rp-8-Br-cAMP and Calphostin C, respectively, blocked GABAARα1 internalization, whereas inhibition of MAPK (U0126) and PI3K (LY294002) did not prevent rapid internalization. By contrast in amygdala cultures, Rp-8-Br-cAMP had no effect. Together, these data suggest that rapid GABAAR internalization during memory consolidation is BDNF-TrkB dependent. Further, it appears that hippocampal GABAAR internalization is PKA and PKC dependent, while it may be primarily PKC dependent in amygdala, implying differential roles for TrkB-dependent kinase activation in BDNF-dependent memory formation.

AB - During the consolidation of fear memory, it has been shown that GABAA receptors (GABAAR) are rapidly downregulated in amygdala. This rapid decrease in GABAAR functioning may permit transient hyperexcitablity, contributing to cellular mechanisms of memory consolidation. Memory consolidation also requires brain-derived neurotrophic factor (BDNF) activation of tyrosine receptor kinase B (TrkB) receptors in the amygdala and hippocampus. We hypothesized that rapid internalization of GABAARα1 is mediated via TrkB activation of PKA and PKC-dependent processes. Primary neuronal cell cultures, from postnatal day 14-21 mouse amygdala and hippocampus, were analyzed with immunofluorescence using cell-surface, whole-cell permeabilization, and antibody internalization techniques, as well as with 3H-muscimol binding assays. In both hippocampal and amygdala cultures, we found a >60% reduction in surface GABAARα1 within 5 min of BDNF treatment. Notably, the rapid decrease in surface GABAARα1 was confirmed biochemically using surface biotinylation assays followed by western blotting. This rapid effect was accompanied by TrkB phosphorylation and increased internal GABAARα1 immunofluorescence, and was blocked by k252a, a broad-spectrum tyrosine kinase antagonist. To further demonstrate TrkB specificity, we used previously characterized TrkBF616A mice, in which the highly selective TrkB-mutant specific antagonist, 1NMPP1, prevented the BDNF-dependent GABAARα1 internalization. In hippocampus, we found both PKA and PKC inhibition, using Rp-8-Br-cAMP and Calphostin C, respectively, blocked GABAARα1 internalization, whereas inhibition of MAPK (U0126) and PI3K (LY294002) did not prevent rapid internalization. By contrast in amygdala cultures, Rp-8-Br-cAMP had no effect. Together, these data suggest that rapid GABAAR internalization during memory consolidation is BDNF-TrkB dependent. Further, it appears that hippocampal GABAAR internalization is PKA and PKC dependent, while it may be primarily PKC dependent in amygdala, implying differential roles for TrkB-dependent kinase activation in BDNF-dependent memory formation.

UR - http://www.scopus.com/inward/record.url?scp=79851508614&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=79851508614&partnerID=8YFLogxK

U2 - 10.1016/j.neuroscience.2010.12.041

DO - 10.1016/j.neuroscience.2010.12.041

M3 - Article

VL - 176

SP - 72

EP - 85

JO - Neuroscience

JF - Neuroscience

SN - 0306-4522

ER -