Localization of acetylcholine receptors during synaptogenesis in retina.

Abstract

Nicotinic acetylcholine receptors are synthesized in chick embryo retina before synapses appear. Most of the receptors are found associated with neurites in the inner synaptic layer of the retina; later in development the receptors appear in the outer synaptic laver.Cells dissociated from retina and cultured in vitro form aggregates and also sort out into regions comprised either of neurites or cell bodies. Most of the nicotinic acetylcholine receptors of aggregates are found associated with neurite regions. The receptor distribution of cultured retina cells thus resembles that of intact retina.125I-Labeled a-bungarotoxin, a specific ligand of the nicotinic acetylcholine (AcCh) receptor, was used to detect AcCh receptors in the developing chick embryo retina. This species of AcCh receptor is a synaptic marker, since the AcCh receptors of striated muscle cells (1, 2) and certain parasympathetic neurons (3) are found in abundance only at the site of the synapse. After denervation, the receptors are found over the entire surface of the cell.Most of the recent information regarding the nicotinic AcCh receptor has been obtained with the receptors of the electric organ of fish, and striated muscle (for reviews see refs. 4-6). a-Bungarotoxin has been used to detect AcCh receptors in sympathetic ganglia (7) and brain (8-10), but the nicotinic AcCh receptors of retina have not been studied in this manner. The vertebrate retina is a model system for studies on synaptic communication, since relatively few types of neurons are present and cells dissociated from retina form synapses in vitro (11,12). The retina contains acetylcholine and enzymes that catalyze the synthesis and the hydrolysis of this compound (13-17). In addition, electrophysiologic studies show that some neurons in retina respond to acetylcholine and that both nicotinic and muscarinic AcCh receptors are present in the retina of certain organisms (18)(19)(20)(21)(22).In this report, the concentration of nicotinic AcCh receptors and their location in chick embryo retina are described as a function of the developmental age of the retina. H-21) with 20 mM Hepes (N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid), pH 7.4, instead of NaHCO3, and 2 mg of serum albumin per ml of medium]. Pieces of retina were incubated for 60 min at 370 in 0.5-1.0 ml of solution A with 10 nM 125I-aBT and then were washed five times with 5 ml of cold solution A and twice with solution A without serum albumin. The tissue was fixed with 10% (vol/vol) Abbreviations: aBT, a-bungarotoxin; l25I-aBT, a-bungarotoxin labeled with 2 atoms of 125I per toxin molecule; AcCh, acetylcholine. MATERIALS AND METHODS Assay of

Neurotransmitter-related features of the retinal pigment epithelium

“…These include receptors for benzodiazepine (Altstein et al, 1981 ;Osborne, 1980;Paul et al, 1980), strychnine (Borbe, Muller and Wollert, 1981;Schaeffer and Anderson, 1981 ), muscarinic acetylcholine (Hruska et al, 1978;Vogel and Nirenberg, 1976), dopamine (Magistretti and Schordoret, 1979;Redburn and Kyles, 1980) and GABA (Osborne, 1981, 198la;Schaeffer, 1980;Yazulla and Brecha, 1981 ;Pourcho, 1981 ). These findings imply the presence of a variety of neurotransmitter species within the neural retina and several of these have been described including GABA, glycine, acetylcholine and dopamine (Watling, 1981 ;Yazulla and Schmidt, 1976), norepinephrine (Osborne, 1981 b), glutamate and aspartate (Berger et al, 1977;Altschuler et al, 1982), and serotonin (Suzuki et al, 1977;Osborne et al, 1981 ;Parkinson and Rando, 1981).…”

Synapse turnover: the formation and termination of transient synapses.

“…All synapses between retina neurons and muscle cells are terminated in 3-10 days depending on the developmental age of the neurons. The half-lives of synapses between muscle cells and retina neurons from 8-, 12-, or (3)(4)(5)(6)(7)(8).…”

“…All synapses between retina neurons and muscle cells are terminated in 3-10 days depending on the developmental age of the neurons. The half-lives of synapses between muscle cells and retina neurons from 8-, 12-, or (3)(4)(5)(6)(7)(8).In this report we examine the specificity of synapse formation by retina neurons. The neurons were cultured with striated muscle cells which also possess abundant nicotinic ACh receptors, to determine whether retina neurons form synapses with muscle cells and whether muscle cells and retina neurons displace one another or are competitive targets for synapse formation by retina neurons that synthesize ACh.…”

Synapse turnover: a mechanism for acquiring synaptic specificity.

“…Nicotinic acetylcholine receptors are present in 6-day chick embryo retina in low concentration and increase in concentration throughout embryo development (8,9). Thus, synapses mediated by nicotinic acetylcholine receptors probably form in chick retina between the 6th and 16th day of embryonic development.…”

“…High choline acetyltransferase (EC 2.3.1.6) (5) and acetylcholinesterase (EC 3.1.1.7) (6) activities, acetylcholine (6), a high-affinity choline uptake mechanism (7), and nicotinic (8,9) and .muscarinic (10) acetylcholine receptors are present in chick retina.…”