The Action of Some Cholinergic Agonists and Anticholinesterase Agents on the Dorsal Muscle of the Leech

Abstract

The leech muscle is one of the oldest preparations for the bioassay of acetylcholine and has served in this capacity on many venerable occasions. The last systematic pharmacological study was carried out by Minz (1932). Minz found the contraction of the eserinized leech muscle practically specific for acetylcholine. Several other agents either were not potentiated by physostigmine or were simply inactive save in very high concentrations.No information is available concerning the effects on the leech muscle of a number of agents which are known to, or assumed to, interact with acetylcholine receptors and which have become available only within the past 20 years. We consider it of sufficient interest and importance to try and fill in these gaps. The results of these investigations indicate an even closer pharmacological similarity between the leech muscle and the endplate of vertebrate skeletal muscle than previously recognized. Succinylcholine and decamethonium were found to be agonists in the leech, but could clearly be differentiated from acetylcholine, carbachol, and nicotine.In addition, the ability of some anticholinesterase agents to potentiate the effect of acetylcholine and related agonists has been studied. These studies, too, take on an added significance in view of the close pharmacological similarity between vertebrate endplate and leech muscle. METHODSThe dorsal muscle of the leech (Hirudo medicinalis) was dissected free of all tissue adhering to its inner surface as described by MacIntosh & Perry (1950). Two strips of muscle were prepared by dividing along the midline. The paired muscles were mounted vertically in identical Lucite organ baths of 10 ml. capacity and suspended in a bicarbonate Ringer solution. The lower end was secured to a hook at the bottom of the bath and the upper end attached by a length of inelastic thread to a strain gauge tension transducer (Grass model FT 03) mounted on an adjustable rack. On isolating and mounting, the muscles tend to go into spasm. This could be overcome by perfusing the organ bath with the Ringer solution at a rate of approximately 3 ml./min.

Localization of the myomodulin-like immunoreactivity in the leech CNS

“…It has been established that the AE and L motor neurons use acetylcholine ( ACh) as a neurotransmitter (Flacke and Yeoh, 1968). More recent experiments demonstrate that in addition to ACh, L motor neurons may use FMRF-amide as a chemical signal (Kuhlman et al, 1985a,b).…”

Differentiation of Acetylcholine and Succinylcholine Receptors in Leech Muscle

“…1 for concentrations of ( +)-tubocurarine of 3 x 10-6-3 x 10-5M. Physostigmine (10-6M) was present because, in the absence of any enzyme inhibitor, the muscle was very insensitive to ACh (Flacke & Yeoh, 1968). Carbachol and nicotine were found to be antagonized in the same way as ACh, but the effect on succinylcholine and decamethonium was quite different and will be described later.…”

“…values are given in Table 1. It has been shown (Flacke & Yeoh, 1968) that, in the leech muscle, succinylcholine was capable of eliciting only about 70% of the maximum response obtained with carbachol.…”

“…We have presented evidence (Flacke & Yeoh, 1968) that, in the leech muscle, acetylcholine, carbachol and nicotine form one group of agonists, and succinylcholine and decamethonium, the so-called long-acting depolarizers, form another. The grouping was based on the differences observed in potency and in size of the maximal responses.…”

“…The methods used have already been described (Flacke & Yeoh, 1968). The effects of different concentrations of an antagonist agent were determined after exposure of at least 30 min to each concentration.…”