238 J. Physiol. (I953) I22, 238-251 MOTOR END-PLATE DIFFERENCES AS A DETERMINING FACTOR IN THE MODE OF ACTION OF NEUROMUSCULAR BLOCKING SUBSTANCES By ELEANOR J. ZAIMIS From the Department of Pharmacology, School of Pharmacy, 17 Bloomsbury Square, W.C. 1 (Received 26 February 1953) Work on cat, human, avian and frog skeletal muscle (Paton & Zaimis, 1952) led to the conclusion that true neuro-muscular block is mainly produced by two different processes: (a) by competition with acetylcholine at the motor end-plates; and (b) by long lasting depolarization of the motor end-plates. Experiments carried out upon other mammalian species reveal the possibility of a third mode of neuromuscular block which combines the two processes mentioned above. These experiments show the characteristics of such a neuro- muscular block and provide evidence that differences in the motor end-plates are a determining factor in the mode of action of a substance at the neuro- muscular junction. A preliminary account of this work has already appeared (Zaimis, 1952). METHODS The mammalian species used have been the cat, monkey, dog, rabbit and hare. Chloralose alone was used as an anaesthetic for the cats, the dose given being 80 mg/kg. To the other species between 100 and 120 mg/kg of chloralose was given, but in a few experiments it proved necessary to supplement this anaesthetic with a small dose of pentobarbitone sodium. The injections of chloralose were made into the saphenous vein. Twitches and tetani of the tibialis, soleus and flexor digitorum sublimis muscles were excited by twice-maximal shocks of 02 msec duration, applied to the tied sciatic nerve in the thigh. In some experiments records of the mechanical response of two muscles in the same leg were recorded on the same drum. Injections were made intravenously or intra-arterially. For the intra-arterial injection the external iliac of the non-operated leg was cannulated towards the bifurcation of the aorta. The aorta was ligatured below the origin of the external iliac arteries. Under such conditions the dose injected was carried to the operated leg. Respiration was recorded by the apparatus described by Paton (1949). For experiments on chicks, birds a few days old were used and injections were made into the jugular vein. NEUROMUSCULAR BLOCK AND SPECIES DIFFERENCES 239 RESULTS Decamethonium Monkeys. The investigations with which this paper deals started with the surprising observation that while a first intravenous dose of 03 mg/kg of decamethonium into a monkey produced a transitory 95 % paralysis of the tibialis muscle, a subsequent dose injected after the complete recovery of the twitch was without effect (Fig. 1 a-c). This is in contrast with the effect of 3 4 _ 1g1 k<g I 0*5 - 04 - 0-3 - 0.2 _ O Fig. 1. a-c, monkey, 3-3 kg. Contractions of tibialis excited by supramaximal shocks to the sciatic nerve, every 10 sec. At (1) and 30 min later at (2), intravenous injection of 1 mg of decamethonium iodide; 10 min later at (3) and (4) injections of 1 mg of decamethonium iodide. At (5) tetanic stimulus to motor nerve, 50/sec. d, from another experiment. Monkey, 8 kg. Chloralose. Tibialis: nerve shock every 10 sec. At (1) intravenous injection of 10 mg of decamethonium iodide (following three smaller doses of decamethonium). At (2) 2 mg neostigmine methylsulphate I.v. At (3) tetanic stimulus to motor nerve, 50/sec. decamethonium on the tibialis muscle of the cat, where a further dose produces a greater paralysis. But this is not the end of the story, for in a monkey the block is rarely preceded by potentiation of the twitch and then only to a very slight degree; the tetanus is not well maintained and antagonizes the block which is also antagonized by neostigmine (Fig. 1 d). In short, the characteristics 240 ELEANOR J. ZAIMIS of this block differ from those of a block produced by pure depolarization, which is usually preceded by a brief period of fasciculation of the muscle and potentiation of the single twitches, and in which the tension of a tetanus is well maintained during the block while neostigmine and tetanus are not antagonists. If one records simultaneously the contractions of the tibialis and soleus muscles in a cat, results show that the tibialis is much more sensitive to decamethonium than the soleus. The same muscles differ in sensitivity to 1 2 3 4 56~~~~~~~~kg7 X. ]~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~. Fig. 2. Dog, 10 kg. Simultaneous records of supramaximal motor nerve twitches of tibialis (upper record) and flexor digitorum sublimiis (lower record). At (1), (2), (4) and (5) intravenous injection of 1 mg of decamethonium. At (3) 1 mg neostigmine methylsuiphate I.V. At (6) and (7) tetanic stimulus to motor nerve, 50/sec. D-tubocurarine but in the opposite way (Paton & Zaimis, 1951). If one now records the contractions of the tibialis and soleus muscles in a monkey the first dose of decamethonium causes a paralysis of tibialis, but has little effect on soleus. When the doses are progressively increased it is found that tibialis becomes less and less sensitive but soleus is increasingly affected. When the paralysis of the soleus starts spontaneous respiration stops. Briefly we start with the picture as in a cat given by decamethonium and finish with that given by D-tubocurarine. On the other hand, the block produced in the monkey by D-tubocurarine has all the well-known characteristics of that produced by competition with acetylcholine; it is not preceded by stimulation and is antagonized by a tetanus or neostigmine. During the block the muscles cannot maintain a tetanus and show an increasing sensitivity to subsequent doses of D-tubocurarine. Dogs, rabbits and hares. A number of tests were also made upon dogs and rabbits. The results show that the neuromuscular block produced by decame- thonium in these species has the same characteristics as that in the monkey. This is clearly indicated in Figs. 2 and 3. The muscles show the same decreasing sensitivity and under the influence of decamethonium they cannot maintain NEUROMUSCULAR BLOCK AND SPECIES DIFFERENCES 241 a tetanus. Moreover, the block is antagonized by tetanus and by neostigmine, and potentiated by D-tubocurarine. The only difference between the dog and rabbit is that the block produced by the first dose of decamethonium is always preceded in the rabbit (Fig. 3, 1) by potentiation of the twitch. 1 2 3 4 |E_0X1 0~~~~~~~~~~~~~~~~~0 S8 67 9 10 11~~~~~~~~~~10 Fig. 3. Rabbit, 3 kg. Contractions of tibialis excited by supramaximal shocks to the sciatic nerve, every 10 sec. At (1), (3) and (4) intravenous injection of 0-6 mg of decamethonium iodide. At (2) and (7), tetanic stimulus to motor nerve, 50/sec. At (5) 2 mg atropine sulphate i.v. At (6) 1-8 mg decamethonium iodide i.v. At (8) and (11) intravenous injection of 05 mg neostigmine methylsulphate. At (9) 1 mg decamethonium iodide and at (10) 0 5 mg D-tubocurarine chloride i.v. The following phenomenon deserves attention as it frequently occurs and is difficult to explain. It has been observed that whereas a first dose of neo- stigmine given during a neuromuscular block antagonizes the block effectively a second dose given during a subsequent dose is a far less effective antagonizer. This is clearly indicated in Fig. 3. The same phenomenon occurs when the block is produced by D-tubocurarine. Results of an experiment carried out on a single wild hare are illustrated in Fig. 4 and are sufficiently clear-cut to be worth recording. Here again the block produced by decamethonium has the same peculiar characteristics: decreasing sensitivity of the muscles to subsequent doses, the tetanus very poorly sustained, neostigmine effectively antagonizing the block and no sign of stimulation preceding it. PH. CXXII. 16 242 ELEANOR J. ZAIMIIS Other depolarizing substances An attempt was then made to study the actions of succinylcholine, a neuro- muscular blocking substance believed to produce its actions in the cat and in avian muscle by pure depolarization. The results obtained with this substance in the monkey and the dog are shown in Figs. 5 and 6. It is clear that in these species succinylcholine produces a block similar to that produced by decame- 2 3 4 6 7 8 Fig. 4. Hare, 3 kg. Simultaneous records of supramaximal motor nerve twitches of tibialis (upper record) and soleus (lower record). At (1) intravenous injection of 0 5 mg, followed at (2) by 1 mg decamethonium iodide. At (3) and (4) tetanic stimulus to motor nerve, 50/sec. 30 min later at (5) decamethonium iodide, 1 mg i.v. At (6) 1 mg neostigmine methylsulphate. At (7) 0-2 mg D-tubocurarine chloride and at (8) 1 mg neostigmine methylsulphate. NEUROMUSCULAR BLOCK AND SPECIES DIFFERENCES 243 thonium, that is to say, a block differing in many ways from one due to long- lasting depolarization only. 1 2 3 4 5 Fig. 5. Monkey 8 kg. Simultaneous records of supramaximal nerve twitches of tibialis (upper record) and soleus (lower record). At (1) and (2), 10 mg. and, 10 min later, at (3) and (4) 30 mg succinylcholine dibromide. At (5) tetanic stimulus to motor nerve, 50/sec. kg I 1.0 .-1 0-5 _05 04 03 02 01 03 _ 0.2 _ox1 I _ Fig. 6. Dog, 10 kg. Simultaneous records of supramaximal nerve twitches of tibialis (upper record) and flexor digitorum sublimis (lower record). At (1) and 20 min later at (2), intra- venous injection of 1 mg of succinylcholine dibromide. Result of interposition of a dose of D-tubocurarine between doses of decamethonium The impression resulting from the consideration of these facts is that the picture obtained from the monkey, dog and hare is similar to that obtained in the cat when an injection of D-tubocurarine is interposed between the doses of decamethonium, as all substances raising the end-plate threshold to acetyl- choline diminiish the activity of any substance producing a neuromuscular block by long-lasting depolarization.