Choroid Neurons Research Articles

Analysis of quantal content and quantal conductance in two populations of neurons in the avian ciliary ganglion

The avian ciliary ganglion contains two populations of parasympathetic cells, termed the ciliary and choroid neurons. We have estimated the quantal contents of nicotinic excitatory postsynaptic potentials in both populations of neurons by several methods. The singly innervated ciliary neurons have quantal contents of 15–30. In contrast, the multiply innervated choroid cells have quantal contents of 4–7. Quantal conductance was also determined, using a parallel conductance model which takes into account the capacitance of the cell membrane. This analysis indicates that in both populations of neurons one quantum activates approximately 100 postsynaptic receptors. It is concluded that in autonomie ganglia singly innervated cells demonstrate a larger quantal content, consistent with a higher safety factor for neurotransmission, while quantal content in multiply innervated cells is generally much lower, allowing for considerable summation of presynaptic inputs. Further, in autonomic neurons many fewer postsynaptic receptors are activated by a single quantum than is the case at the neuromuscular junction.

Properties of choroid and ciliary neurons in the avian ciliary ganglion and evidence for substance P as a neurotransmitter

Intracellular recordings were made from identified choroid and ciliary neurons in the ciliary ganglion of the embryonic chick. Choroid neurons, which were innervated by multiple preganglionic fibers, frequently displayed noncholinergic slow excitatory postsynaptic potentials (EPSPs) following repetitive stimulation of the preganglionic nerve trunk. These slow potentials were blocked by high Mg2+/low Ca2+ buffer and were closely mimicked by bath application of substance P, which is known to be present within both populations of preganglionic nerve terminals. Substance P-induced depolarizations desensitized during prolonged exposure, at which time it was no longer possible to evoke slow synaptic potentials. Following manual voltage clamp to resting membrane potential, parallel increases in input resistance were seen during the slow EPSP and the response to substance P, suggesting that the two responses share common mechanisms. Ciliary neurons, which were innervated by a single preganglionic fiber and displayed dual electrical-chemical synapses, did not exhibit slow synaptic potentials and were unaffected by bath application of substance P. The magnitude and time course of fast nicotinic EPSPs elicited in ciliary neurons by 0.5 Hz presynaptic stimulation were also unchanged in the presence of 1 to 3 microM substance P. Although the ciliary and choroid neurons share a common embryological origin in the neural crest, they are specialized for quite different physiological roles. Integration of multiple presynaptic inputs occurs at choroid synapses, mediated by the presence of both subthreshold fast nicotinic EPSPs and the slow EPSP. In contrast, synapses on ciliary neurons have specializations which preclude any integrative function, including single innervation, a high quantal content, electrical coupling potentials, and a lack of slow synaptic potentials.

Gamma-Aminobutyric acid receptors on chick ciliary ganglion neurons in vivo and in cell culture

In the chick ciliary ganglion, preganglionic terminals maintain cholinergic synapses on the choroid neurons and both cholinergic and electrical synapses on the ciliary neurons. The preganglionic terminals also contain enkephalin- and substance P-like immunoreactivity, suggesting that transmission through the ganglion is more complicated than is indicated by the known synaptic connections. We report here that embryonic chick ciliary ganglion neurons also have gamma-aminobutyric acid (GABA) receptors and that GABA applied to the ganglion can block transmission elicited by preganglionic stimulation. Studies on the neurons in cell culture indicate that the GABA response is mediated by GABAA receptors: GABA activates a Cl- conductance, and the response can be mimicked by muscimol and blocked by bicuculline or picrotoxin. The GABA receptors are regulated independently from acetylcholine (ACh) receptors on the neurons since the levels of ACh and GABA sensitivity are influenced differently by culture age and by chronic exposure to GABA or elevated K+ concentrations. Application of GABA to intact ciliary ganglia increases the membrane conductance of ganglionic neurons (as in culture), reduces to subthreshold the amplitude of excitatory postsynaptic potentials in the neurons elicited by preganglionic stimulation and completely blocks transmission through the ganglion. A native source of ligand for the receptors in vivo has yet to be identified.

Surface AChE in the chick ciliary ganglion neurons: Ultrastructural localization and possible relations to α-bungarotoxin receptors

The localization of acetylcholinesterase activity in the chick ciliary ganglion was investigated by ultrastructural cytochemistry. Both ganglionic cell populations, i.e. the ciliary and the choroid neurons, showed similar distribution patterns of the enzymic activity in the cytoplasm as well as at the neuronal surface. As indicated by specific inhibition tests, the whole enzymic activity was attributable to specific acetylcholinesterase. While the endocellular activity was mainly localized in the rough endoplasmic reticulum, the surface activity occurred at postsynaptic level and at extrasynaptic areas, where the neuronal membrane comes into contact with the plasma membrane of the satellite cell (boundary neuron-satellite cell). Enzymic activity also uniformly occurred at the surface of preganglionic nerve terminals. The surface localization of specific acetylcholinesterase recalls that recently described for α-bungarotoxin receptors, which suggests that acetylcholinesterase and α-bungarotoxin receptors can be distributed together, not only at postsynaptic level but also in extrasynaptic neuronal areas and at presynaptic level. The possibility that α-bungarotoxin receptors and acetylcholinesterase form a ·receptive’ system not engaged in ganglionic transmission and not exclusively confined to postsynaptic level is discussed in relation to the electrophysiological data existing in literature.

The effects of α- and β-neurotoxins from the venoms of various snakes on transmission in autonomic ganglia

We have previously shown that certain commercially available lots of α-bungarotoxin block transmission in ciliary and choroid neurons of both pigeon and chicken ciliary ganglia at a concentration of 10 μg/ml (1.2 μM). The blockade is antagonized by pre-incubation with 100 μM tubocurarine. Further evidence that this blockade is produced by a postsynaptic action, as one would expect of an α-neurotoxin, are our findings that: (a) exposure to the toxin prevents the depolarization of ganglion cells normally seen in response to the cholinergic agonist, carbachol; and (b) the blocking activity of the toxin is removed by treatment with membranes purified from Torpedo electric organ containing an excess of α-neurotoxin binding sites. A high affinity binding site for [ 125I]α-bungarotoxin was characterized in the chicken ciliary ganglion. However, since it is labelled equally well by lots of α-bungarotoxin which block transmission and those that do not, this site does not appear to be involved in the blockade of transmission. α-Cobratoxin (from Naja naja siamensis), the α-neurotoxin L.s. III (from Laticauda semifasciata) and certain lots of α-bungarotoxin produce a partial blockade of transmission in ciliary neurons of the pigeon ciliary ganglion at a concentration of 10 μg/ml (1.2 μM), but have no effect on transmission in choroid neurons. Two other α-neurotoxins from Laticauda semifasciata, erabutoxin a and erabutoxin b, have no effect on transmission in either cell population at this concentration. None of the α-neurotoxins tested had any effect on transmission in either the rat superior cervical ganglion or the rat pelvic ganglion at concentrations up to 100 μg/ml (12 μM). Collagenase treatment of these ganglia, in an attempt to increase access of the toxins to ganglion cells, did not alter these negative results. β-Bungarotoxin (0.5 μg/ml, 0.02 μM) produces a complex blockade of transmission in both avian ciliary ganglia and rat superior cervical ganglia. Unlike the action of α-bungarotoxin, the blockade of ciliary ganglion transmission by β-bungarotoxin is irreversible and is not prevented by pretreatment with tubocurarine.

Immunocytochemical localization of α-bungarotoxin receptors in the chick ciliary ganglion: Synaptic and extrasynaptic sites?

The immunocytochemical localization of nicotinic acetylcholine receptors in the chick ciliary ganglion was investigated at the ultrastructural level with a procedure utilizing binding of α-bungarotoxin visualized by immunoperoxidase histology. Both ganglionic cell populations, i.e. the ciliary and choroid neurons, showed specific immunoreactivity for receptors. In both types of neurons evident stain was found on the postsynaptic membrane. Often the reaction product filled discrete regions of the synaptic cleft. Furthermore, specific staining was also observed on large areas of the neuronal surface devoid of presynaptic nerve endings. These data probably indicate the occurrence of both synaptic and extrasynaptic nicotinic receptors on the neuronal plasma membrane. Immunoreactivity was also observed on the membrane of the presynaptic nerve endings, and on the part of the satellite plasma membrane which is adjacent to the neuron. These last results are discussed in relation to the occurrence of possible artifacts or, alternatively, to the presence of presynaptic and glial receptors. Immunoreactivity at all sites was prevented almost completely by ganglion incubation in 1 mM d-tubocurarine prior to and during treatment with toxin.

α-Bungarotoxin-acetylcholine receptors in the chick ciliary ganglion during development

[125I]α-Bungarotoxin binds in a saturable and practically irreversible fashion to membrane-associated receptors in the ciliary ganglion of the chick embryo and young chick. Nicotinic but not muscarinic ligands are potent inhibitors of toxin binding. Thus, the binding site for toxin in the ganglion appears to have the properties of a nicotinic acetylcholine receptor. The ontogeny of the receptors was studied from day 7in ovo to day 70 after hatching. When expressed per ganglion, the developmental pattern of the receptors may be divided into three stages: a peak of high receptor level (days 7–15in ovo) with a maximum at day 12in ovo, a period of constant level (days 15in ovo to day 8 after hatching) and a second increase in receptor number (days 8–50 after hatching).Light-microscope autoradiography suggests that different developmental changes in receptor distribution occur in the two neuronal populations of the ganglion (i.e. the ciliary and choroid neurons). While choroid neurons show an adult-like receptor distribution from late embryonic stages, the ciliary neurons do not fully develop their distribution pattern until after hatching.These findings are discussed in relation to morphological and physiological events which are known to occur in the ganglion during development, in particular those concerning synaptic transmission.

α-Bungarotoxin-acetylcholine receptors in the chick ciliary ganglion: Effects of deafferentation and axotomy

alpha-Bungarotoxin (alpha-BuTX) binds in a saturable and practically irreversible fashion to membrane-associated receptors in the ciliary ganglion of the adult chick. The binding of toxin to receptors is competitively inhibited by nicotinic cholinergic ligands, and for these properties the receptors are regarded as acetylcholine receptors of the nicotinic type (alpha-buTX-AChRs). The rate constant of association (K1) and dissociation (K-1) of the toxin-receptor reaction has been estimated to be K1 = 7.4 x 104 M(-1) sec(-1) and K-1 = 9.6 X 10(-6) sec(-1), respectively. Light autoradiography shows that most, if not all, the receptors are related to surface membrane, probably to synaptic areas of both choroid and ciliary neurons. The choroid neurons contain more receptors than the ciliary ones. A single chick ciliary ganglion binds specifically 47 fmole of alpha-BuTX in situ corresponding to 2.83 x 1010 alpha-BuTX-AChRs/ganglion. No changes in number and distribution of the toxin receptors occur following preganglionic denervation. Conversely, postganglionic axotomy causes a rapid disappearance of the receptors in situ. Since binding experiments in vitro revealed a partial, instead of a total, loss of the receptors, it is suggested that the disappearance of the receptors in situ includes both a partial loss of the original receptors and the masking of the residual ones.

Alpha-Bungarotoxin blocks nicotinic transmission in the avian ciliary ganglion.

alpha-Bungarotoxin binds to nicotinic receptors in skeletal muscle, blocking neuromuscular transmission. Because this toxin has recently been shown to bind to chicken ciliary ganglia, an attempt has been made to determine whether it also blocks nicotinic transmission in this ganglion, alpha-Bungarotoxin (1 micrometer) completely blocked nicotinic transmission in both the ciliary and choroid neurons of chicken and pigeon ciliary ganglia. The effect of the toxin could be partially reversed by prolonged washing (2--8 hr). Incubation of ganglia with d-tubocurarine (0.1 mM) prior to the addition of alpha-bungarotoxin significantly decreased the duration of the washout period necessary to restore transmission. These results suggest that d-tubocurarine and alpha-bungarotoxin are interacting with the same receptor. Under similar conditions, alpha-bungarotoxin did not block nicotinic transmission in the rat superior cervical ganglion, in agreement with previous reports. The avian ciliary ganglion is the only vertebrate autonomic ganglion in which both alpha-bungarotoxin binding and alpha-bungarotoxin blockade of transmission have been shown to occur. This ganglion therefore provides a model system for using alpha-bungarotoxin to study neuronal nicotinic receptors.