Robert W. Baughman

Biochemical characterization and cellular localization of the cholinergic system in the chicken retina

The biochemical properties and morphological localization of the cholinergic system in the chicken retina were studied with the following results. (1) The uptake of [3H]choline saturated with increasing choline concentration, and could be accounted for by the presence of two saturable processes, a ‘high affinity’ system with aKmof1.1 μM and Vmax of 8.0 pmole/min/mg protein, and a ‘low affinity’ system with aKmof214 μM and Vmax of578 pmole/min/mg protein. (2) Following an incubation with5 × 10−7 M[3H]choline for 10 min, the intracellular composition of labeled products consisted of 44% acetylcholine, 46% choline and 10% phosphorylcholine. (3) Incubation in the presence of 5 × 10−5 M hemicholinium-3 or in the absence of sodium blocked synthesis of acetylcholine and reduced the total uptake of [3H]choline by an amount which could be accounted for by blockage of the high affinity system. (4) Labeled acetylcholine synthesized from [3H]choline could be released by increasing the extracellular potassium concentration, and this release was calcium dependent. (5) By the use of freeze drying and autoradiography with a dry emulsion film, the [3H]choline taken up was found to be concentrated in 6% of the cell bodies in the inner nuclear layer, in 19% of the cell bodies in the ganglion cell layer and in two bands in the inner plexiform layer. (6) When autoradiography was repeated following an incubation carried out in the presence of hemicholinium-3, no localization of3H uptake was observed. (7) Following an incubation with5 μM [3H]choline, the total uptake of choline and the contributions of the high and low affinity uptake systems were estimated from an autoradiograph by grain counting, and the values obtained closely matched those found with the biochemical methods.

GABAERGIC TRANSCALLOSAL NEURONS IN DEVELOPING RAT NEOCORTEX

In the mature cerebral cortex the interhemispheric connections across the corpus callosum appear to be essentially completely excitatory on the basis of both immunocytochemical and electrophysiological studies. During late embryonic development, however, immunocytochemical staining reveals numerous GABA‐positive fibres in the callosum, which later largely disappear. The origin of these fibres and whether they represent functional GABAergic neurons has not been established. In the present study we used a combination of retrograde labelling in vivo with electrophysiology and immnunocytochemistry in cell culture to show that transiently at birth in rat pups a substantial number of transcallosal cortical cells are functional GABAergic neurons. Possible roles and fates for these neurons are discussed.

Choline acetyltransferase immunocytochemistry of Edinger-Westphal and ciliary ganglion afferent neurons in the cat

The distribution of cholinergic neurons in the region of the cat Edinger-Westphal nucleus (EW) was determined by immunocytochemical localization of the acetylcholine-synthesizing enzyme choline acetyltransferase (ChAT). Neurons containing ChAT-like immunoreactivity (ChAT-LI) were densely distributed within EW, the anteromedian nucleus (AM), and the oculomotor nucleus (III), and were also present in immediately adjacent regions of the periaqueductal gray and ventral tegmental region. The majority of labelled neurons in EW and AM showed a markedly lower intensity of ChAT-LI than the labelled neurons in III and adjacent regions. To determine the relationship of cells with ChAT-LI to the distribution of ciliary ganglion afferent neurons, a double labelling immunocytochemistry/retrograde transport technique was also used. These experiments showed that many of the cells located outside of III that stained intensely for ChAT-LI project to ciliary ganglion. Very few ciliary ganglion afferent neurons were found in EW or AM itself; instead, the distribution of lightly labelled ChAT-LI-positive neurons in EW and AM more closely matched the known distribution of peptide-containing cells that have descending, central projections.

Cholinergic Innervation in Cerebral Cortex

This chapter summarizes current knowledge about the anatomical organization of cholinergic innervation in cerebral cortex. Many investigations over the last five decades have contributed to establishing ACh as a neurotransmitter in cerebral cortex. It was realized early that cerebral cortex contains all of the components of cholinergic metabolism in moderately high amounts, including ACh itself, the ACh-synthesizing enzyme choline acetyltransferase (ChAT, EC 2.3.1.6), and the ACh-degrading enzyme acetylcholinesterase (AChE, EC 3.1.1.7). Later, the presence in cortex of both muscarinic and nicotinic binding sites (probably reflecting ACh receptors) was described (for review see Fonnum, 1973, 1975; Kuhar, 1976; Emson and Lindvall, 1979; Fibiger, 1982; Parnavelas and McDonald, 1983). Cortical synaptosomes, subcellular fractions containing mainly isolated synaptic complexes, have been shown to contain ACh, ChAT, AChE, and sodium-dependent high-affinity choline uptake (HACU) (Yamamura and Snyder, 1973; Kuhar and Murrin, 1978), and ACh can be released from cortex in vivo (Mitchell, 1963: Collier and Mitchell, 1966) and from synaptosomes in a Ca2+-dependent manner (Kuhar and Murrin, 1978). Many studies have investigated the pharmacological and physiological effects of ACh in cortex (Krnjevic and Phillis, 1963a–c; Crawford, 1970; Lamour et al. 1982b). Altogether, the evidence strongly suggests that ACh acts as a neurotransmitter in cerebral cortex, although the complex anatomical organization of this tissue has not permitted experiments as straightforward and conclusive as those that have characterized cholinergic transmission at the neuromuscular junction.

The National Institutes of Health Blueprint for Neuroscience Research

The National Institutes of Health (NIH) Blueprint for Neuroscience Research is a collaborative effort among the NIH Office of the Director and 16 NIH institutes and centers.[a][1] Its aim is to develop research tools, create research resources shared by the entire neuroscience community, train a new

The Cholinergic System in the Chicken Retina: Cellular Localization and Development

The cellular localization of the cholinergic system of the chicken retina was determined by means of freeze-drying and dry autoradiography following incubation with [3H]choline under conditions favoring high-affinity uptake. The cholinergic cells were localized to the inner nuclear and ganglion cell layers, and they extended processes in two bands in the inner plexiform layer. During embryogenesis in the chick, the cholinergic system was found to develop in two stages; the first occurs relatively early in retinal differentiation and is associated with increased ACh synthesis and storage and with a large rise in CAT activity; and the second occurs just before hatching, coincident with synaptogenesis and the appearance of visual function, and is associated with further increases in ACh synthesis and storage and with the development of high-affinity choline uptake.