Persistent phosphorylation parallels long-term desensitization of cerebellar purkinje cell AMPA-type glutamate receptors.
This study is aimed at testing the hypothesis that sustained phosphorylation underlies long-term desensitization of AMPA receptors, which is thought to be the mechanism of long-term synaptic depression in cerebellar Purkinje cells (PCs). We induced long-term desensitization of AMPA receptors in rat cerebellar slices by (1) a 4-min bath application of quisqualate (0.1 mM) or (2) a 15-min bath application of a protein kinase C (PKC) activator, phorbol-12,13-diacetate (0.5 microM) or -dibutyrate (0.6 microM), followed by a 4-min AMPA (0.1 mM) application. In slices so treated, labeling with an antibody (12P3) against a peptide corresponding to part of AMPA receptor subunit GluR2 including serine 696 and phosphorylated at this serine site revealed phosphorylation of the AMPA receptors in PC dendrites that was sustained for at least 1 hr. At an early phase, within 20 min after the chemical stimulation, the phosphorylation was resistant to an Ca2+ chelator (BAPTA-AM), a metabotropic glutamate receptor antagonist (MCPG), and a PKC inhibitor (calphostin C), whereas at a late phase, 30 min or more after the chemical stimulation, it was blocked by these reagents similarly to long-term desensitization of AMPA receptors. Taken together with data obtained previously using different protocols of chemical stimulation, the present results strongly support the above-mentioned hypothesis.
Nakazawa
K
K
Laboratory for Memory and Learning, Institute of Physical and Chemical Research (RIKEN), Saitama, Japan.
Mikawa
S
S
Ito
M
M
D000818:Animals; D000906:Antibodies / Q000187:drug effects; D002118:Calcium / Q000378:metabolism; D002531:Cerebellum / Q000166:cytology / Q000378:metabolism*; D002614:Chelating Agents / Q000494:pharmacology; D003712:Dendrites / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D018690:Excitatory Amino Acid Agonists / Q000494:pharmacology; D066298:In Vitro Techniques; D010446:Peptide Fragments / Q000276:immunology; D010703:Phorbol Esters / Q000494:pharmacology; D010766:Phosphorylation / Q000187:drug effects; D010770:Phosphotransferases / Q000037:antagonists & inhibitors; D011233:Precipitin Tests; D011689:Purkinje Cells / Q000378:metabolism*; D016318:Quisqualic Acid / Q000494:pharmacology; D011817:Rabbits; D018091:Receptors, AMPA / Q000187:drug effects / Q000276:immunology / Q000378:metabolism / Q000502:physiology*
D000906:Antibodies; D002614:Chelating Agents; D004791:Enzyme Inhibitors; D018690:Excitatory Amino Acid Agonists; D010446:Peptide Fragments; D010703:Phorbol Esters; D018091:Receptors, AMPA; D016318:Quisqualic Acid; D010770:Phosphotransferases; C104722:glutamate receptor ionotropic, AMPA 2; D002118:Calcium