Neuronal acetylcholine receptors: fate of surface and internal pools in cell culture

Abstract

Chick ciliary ganglion neurons have nicotinic ACh receptors that mediate synaptic input to the cells. Ultrastructural studies with a monoclonal antibody that recognizes the neuronal ACh receptor have previously shown that, in addition to a predominantly synaptic location for the receptors on the neuron surface in vivo, substantial amounts of intracellular receptor are present as well. Here we report that intracellular receptor and smaller receptor-related components make up at least two-thirds of the total antibody binding sites associated with the ciliary ganglion neurons in cell culture. The intracellular sites for the most part represent integral membrane components that bind to concanavalin A when solubilized, indicating that the components are glycosylated. Sucrose gradient analysis shows that the intracellular material includes a 10 S component, likely to represent assembled receptor, along with species sedimenting in the 5-9 S range. Blocking the surface sites with unlabeled antibody and measuring the appearance of the new receptor on the cell surface with radiolabeled antibody indicates that the receptors are inserted into the plasma membrane at a rate equivalent to about 4% of the total surface receptor per hour. The transit time for newly synthesized receptor to reach the cell surface appears to be 2-3 hr. These observations suggest that about 5% of the intracellular receptors are transported to the cell surface in culture. The half-life of ACh receptors in the plasma membrane was estimated by 2 different approaches to be about 22 hr. Surface and internal sites respond in a qualitatively similar way to external agents that specifically modulate cholinergic receptors.(ABSTRACT TRUNCATED AT 250 WORDS)