Long-term desensitization of nicotinic acetylcholine receptors is regulated via protein kinase A-mediated phosphorylation.
During prolonged application of transmitter, ligand-gated ion channels enter a nonconducting desensitized state. Studies on Torpedo electroplax nicotinic acetylcholine (ACh) receptors have shown that entry into the desensitized state is accelerated by protein kinase A-dependent (PKA) receptor phosphorylation. To examine the effects of phosphorylation on desensitization of muscle-type ACh receptors, we expressed the frog embryonic receptor type in Xenopus oocytes. Treatment of embryonic muscle ACh receptors with 8-Br cAMP had no measurable effect on the rate of entry into a desensitized state, but it greatly accelerated the recovery from desensitization. Three complementary approaches to reduce the levels of receptor phosphorylation provided additional evidence for a role of PKA-dependent phosphorylation in rescuing receptors from long-term desensitization. Inactivation of the endogenous PKA activity by coexpression of an inhibitor protein, treatment of receptors with phosphatase, and removal of phosphorylation sites by site-specific subunit mutation all resulted in slowed recovery. Our findings point to the existence of two distinct desensitized states: one requiring several seconds for full recovery and a second state from which recovery requires minutes. Receptors lacking PKA phosphorylation sites exhibit a pronounced increase in the slowly recovering component of desensitization, suggesting that receptor phosphorylation speeds overall recovery by reducing the entry into a deep desensitized state. This newly described effect of phosphorylation on ACh receptor function may serve as an important modulator of postsynaptic receptor sensitivity.
Paradiso
K
K
Department of Neurobiology and Behavior, State University of New York at Stony Brook, Stony Brook, New York 11790, USA.
Brehm
P
P
D015124:8-Bromo Cyclic Adenosine Monophosphate / Q000494:pharmacology; D000109:Acetylcholine / Q000494:pharmacology; D000367:Age Factors; D000818:Animals; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism*; D004594:Electrophysiology; D015640:Ion Channel Gating / Q000187:drug effects / Q000502:physiology*; D018482:Muscle, Skeletal / Q000737:chemistry / Q000201:enzymology; D009865:Oocytes / Q000502:physiology; D018408:Patch-Clamp Techniques; D010749:Phosphoprotein Phosphatases / Q000378:metabolism; D010766:Phosphorylation; D011978:Receptors, Nicotinic / Q000378:metabolism*; D013569:Synapses / Q000737:chemistry / Q000378:metabolism; D014981:Xenopus
D011978:Receptors, Nicotinic; D015124:8-Bromo Cyclic Adenosine Monophosphate; D017868:Cyclic AMP-Dependent Protein Kinases; D010749:Phosphoprotein Phosphatases; D000109:Acetylcholine