Cholinergic Transmission in the Frog Spinal Cord

Abstract

The effect of acetylcholine and other drugs has been tested on the isolated and sagittally hemisected spinal cord of the frog (Rana temporaria). The release of acetylcholine from this preparation during stimulation of the spinal roots and whole hemicord has also been studied. After inactivation of the tissue cholinesterases acetylcholine was released spontaneously from the preparation at a steady rate of about 5.5 pmole/15 min/preparation. No increase in this release was obtained by stimulating the dorsal roots, but antidromic stimulation of the ventral roots always gave an increase in the rate of acetylcholine release up to 2.3 times the spontaneous level. Direct stimulation of the spinal cord did not alter the rate of acetylcholine release. Acetylcholine (1 mm) and eserine (10 mM) had weak excitatory actions and prolonged the reflex response of the preparation during dorsal root stimulation, but dihydro-,8-erythroidine and atropine had no effect. The response evoked by antidromic ventral root stimulation was potentiated by eserine (10 rM) and depressed by acetylcholine (10 MM) and dihydro-18-erythroidine (1 lAM) as in the toad preparation.The evidence supporting cholinergic transmission at some synapses in the spinal cord is discussed in relation to these results.In a previous paper (Kiraly & Phillis, 1961) it was suggested that the slow depolarizing dorsal root potentials evoked by stimulation of adjacent ventral roots of the toad spinal cord were generated along a neuronal pathway containing at least one cholinergic synapse. It was further suggested that this synapse might be similar, anatomically and pharmacologically, to the synapse between motor axon collaterals and Renshaw cells in the cat.It is generally considered (Crossland, 1957;Paton, 1958) that, to qualify as a synaptic transmitter agent, the postulated substance should be released during stimulation of the appropriate pre-synaptic elements. Hence it was felt that the demonstration of an increased acetylcholine content in the solution bathing the isolated frog spinal cord during ventral root stimulation would provide further evidence of cholinergic synapses in the ventral root-dorsal root pathway.

Biochemical Aspects of Transmission at Inhibitory Synapses: The Role of Glycine

“…On the basis of the distribution of the content of glycine within the spinal cord of seven vertebrates , the isolated amphibian spinal cord (Mitchell and Phillis, 1962;Tebecis and Phillis, 1969) seemed to be a useful preparation since the biochemical data support the transmitter role of glycine in this tissue. Aprison (1970a,b) conducted some of these experiments in which the spinal cord of the toad was isolated, hemisectioned, and placed in a modified Lucite bath for transmitter release studies.…”

Demonstrations

“…Stimulation of the ventral roots (1 Hz; 0X1 msec) during a 10 min collecting period yields an increase in the ACh output (+ 123 %) which is absent if the preparation is bathed in Ca-free Ringer or treated with tetrodotoxin. Stimulation of the dorsal roots fails to elevate the ACh release as already described by Mitchell & Phillis (1962 Histamine can increase rat gastric mucosal blood flow (MBF) even when its stimulatory effect on gastric acid secretion has been inhibited by the histamine H2-receptor antagonist, burimamide, or an antisecretory prostaglandin E2 methyl analogue (Main & Whittle, 1974). We have further investigated the nature of the vascular histamine responses in the gastric mucosa using selective histamine Hi-and H2-receptor agonists.…”

“…However, specialized synaptic structures are rarely seen in gastropod nervous systems. Acetylcholine (ACh) is present in the frog spinal cord (Cammelli, De Bellis & Nistri, 1974) and is released following stimulation of the ventral roots (Mitchell & Phillis, 1962). The amphibian spinal cord is much used for neuropharmacological studies (Curtis, Phillis & Watkins, 1961;Tebecis & Phillis, 1969).…”

PROCEEDINGS OF THE British Pharmacological Society

“…In view of similarities in the transport systems, these findings may also throw light on gastrointestinal function. It is well known, for instance, that ricinoleic acid has a pronounced cathartic action, although it is uncertain whether this is due to effects on fluid transport or on smooth muscle (Fingl, 1970 In the frog spinal cord the motor axon collaterals are cholinergic fibres (Mitchell & Phillis, 1962) which can be directly activated by antidromic ventral root stimulation. Since small concentrations of morphine depressed spontaneous ACh output but failed to reduce the electrically-evoked ACh output, it is suggested that morphine reduced ACh release through an indirect mechanism probably mediated by interneurones.…”