The Action of Potassium on Echinoderm, Molluscan and Crustacean Muscle

As far as the writer is aware, no kymograph records of a rhythmic preparation from an echinoderm have yet been published. Crozier (1916) described the actions of various ions on the cloacal complex of a large Bermudan holothurian, but did not use the kymographic method. The writer finds that a fairly satisfactory preparation can be made by excising the cloacal complex of Cucumaria elongata (i.e. the gut w′all and the hinder end of the body-wall musculature, the whole being slit up longitudinally to allow ready penetration of the bathing fluid) and suspending it, as a longitudinal strip, in a bath of suitable design (e.g. Wells, 1937, Fig. 4).

Papers on the potassium relations of these hearts are already available (see Heymans (1923) for Aplysia, and Lovatt Evans (1912), Hogben (1925) and Cardot (1922) for Helix). In no case, however, have the experiments been conducted in such a way as to allow ready comparison with those made by the writer on other species, and he has therefore reinvestigated the question in order to make such comparison possible. The hearts were perfused by means of a constant-pressure cannula (Hogben, 1925,-Fig. 2) inserted into the auricle. In the case of Helix, there exists a difference of opinion between Lovatt Evans, who states that the heart is peculiarly insensitive to potassium, and Hogben and Cardot, who deny this. The writer’s experiments lead to the latter conclusion. Lovatt Evans does not explicitly state in his paper that the solutions of abnormal potassium content were applied to the interior surfaces of the heart. If they were not, then insensitivity would be expected.

Descriptions of the ion relations of various decapod hearts are already available (Hogben, 1925; Wells, 1928 ; Zoond & Slome, 1929). The different species vary, as described below, in their reaction to moderate potassium excess, and it was felt that further data were desirable. The heart of the shore crab has not yet been studied from this standpoint, and was therefore chosen for the present work. It was perfused by means of a constant-pressure cannula inserted into the sternal artery.

Bathing and perfusion fluids were made by mixing isotonic solutions of the single salts—0·6 M NaCl and KC1, and 0·4 M CaCl, and MgCl₂, for the marine species, whose body fluids are normally isotonic with sea water, and 0·125 Af NaCl and KC1, and 0·083 M CaCl₃ for Helix, whose blood is roughly isotonic with frog Ringer.

In each experiment two fluids were employed. One consisted of NaCl, to which appropriate amounts of the divalent metal chlorides had been added, and the other of KC1, with identical additions. A trace of buffer was also included, and the pH was adjusted to 7·4 for Aplysia, or 8·0 for the other three species. These two fluids were mixed in various proportions, and the mixtures were applied to the tissue. In this way, the ratio of Na to K was varied at will, while the osmotic pressure, pH, and calcium and magnesium concentrations remained constant. The details of these concentrations are given in the figure legends; for Cucumaria, the calcium and magnesium were present in the same amount as in sea water, while the various hearts were studied in magnesium-free mixtures.

Ideally, each experiment should start in a ‘normal’ mixture, whose potassium content is equal to that of the body fluids. In the case of many invertebrates, however, the potassium content of the blood is not known. As most preparations will remain active for some time in the absence of potassium, strictly comparable results can be got by taking a potassium-free mixture as the starting point in each case, and studying the effects of changing from this to a series of solutions with different potassium contents, returning to the potassium-free fluid after each exposure. This was the method adopted in the experiments here described.

The general picture given by a rhythmic muscle in response to sudden change from a potassium-free fluid to one containing a small amount of potassium (say Na:K = 100; 1 or 2) is as follows:

1. Immediately after the change there is a sharp drop in tone.

2. The change is followed by a more or less well-defined period of inhibition (‘potassium paradox’ of Libbrecht (1921)).

3. After the paradox, the beat is more vigorous, more regular, and above all more lasting than before.

On changing back to potassium-free, the most striking result is a sharp tone rise. There is no paradox.

The tone drop caused by low potassium concentrations is shown by all the preparations employed in this research (Figs. 1–4). It appears in a very wide range of other rhythmic preparations, and perhaps in all. It is shown, for instance, by the gastropod and insect crops, by the earthworm gut, and by vertebrate plain and cardiac muscle. The chief differences in appearance between records from different muscles result from variation in the relative amplitudes of the tone and rhythm contractions. In hearts, and especially in crustacean and vertebrate hearts, the tone changes are relatively small, while in gut preparations they are generally very great.

The tone rise seen on returning to a potassium-free fluid is simply the reciprocal of this drop, and is also of general occurrence.

The potassium paradox is usually shown by the Cucumaria preparation, but was not clearly seen in the records of the molluscan hearts, and only occasionally appeared in the case of Carcinus. It is, however, a widely distributed phenomenon. It has been reported as occurring in the Helix and Dytiscus crops (Wells, 1928; Hobson, 1928), in the Limulus heart (Chao, 1934), in the Maia and Homarus hearts (Wells, 1928), and in vertebrate plain and cardiac muscle (Libbrecht, 1921; Jendrassik, 1924). The diastolic standstill produced by small amounts of potassium in the hearts of Palinurus and Octopus (Zoond & Slome, 1929) is apparently a par-ticularly striking case of potassium paradox. The conditions governing the extent of the paradox are sometimes rather tricky (Chao, 1934). In the Aplysia crop, appearance of the paradox is favoured by the presence of magnesium or by a high calcium content (Wells, 1928). The phenomenon could probably be demonstrated in the gastropod heart in suitably designed experiments, e.g. in the presence of magnesium.

On changing from a potassium-free fluid to one containing a moderately high potassium concentration (say Na: K= 100 : 10 or 20), the following responses are seen:

1. In most cases, the preparation passes into contracture.

2. The rhythmic contractions are inhibited.

On returning to potassium-free, the contracture passes off rapidly, while recovery of the beat is less prompt.

Contracture is clearly shown by the Cucumaria preparation, and by the snail and sea-hare hearts (Figs. 1–3). It is, in fact, exceedingly widespread among rhythmic muscles, and appears to be the most general response to potassium excess. There exists, however, a group of preparations in which the reaction to moderate excess is somewhat different. After a more or less noticeable transient contracture, they reach a level intermediate between the original diastolic and systolic levels, and usually nearer to the former. After this the lever traces a horizontal or slowly rising line. This type of response is shown by the frog heart, and by most crustacean hearts (Fig. 4). An intermediate condition appears in the earthworm gut, where contracture is evoked, but passes off fairly rapidly (Wu, 1939). The writer does not believe that this variation in reaction pattern indicates a fundamental difference in the physiological make-up of the preparations. In the Crustacea, at least, the heart can be brought to full contracture by sufficiently increasing the potassium concentration (Fig. 4), and in the spider crab some individuals show prolonged contracture, and others a response like that in the middle record of Fig. 4, to moderate potassium excess (Wells, 1928). A somewhat similar situation is encountered in vertebrate skeletal muscle. Even in different muscles of the same animal, the type of response to potassium excess varies greatly. Thus the frog’s ileofibularis consists of two parts, one of which shows great and prolonged potassium contracture, while the other responds to identical potassium concentrations with a slight, transient, fibrillating contraction (Sonunerkamp, 1928). The records show a rather striking resemblance to those got from rhythmic muscles—the ‘Tonus-bündel’-of the ileofibularis to a snail heart, and the other portion to a lobster heart—and it is at least possible that in rhythmic muscles we are concerned with a physiological differentiation of the same kind.

Inhibition of the rhythm is always seen with sufficient potassium excess.¹ Many authors (e.g. Heilbrunn, 1937; Fenn, 1940) refer to a stoppage of the type seen in the lower parts of Figs. 2 and 3 as systolic, and to one like the centre part of Fig. 4 as diastolic. It will, however, be obvious, from an inspection of the accompanying records, that the heights at which arrest occurs have nothing to do with the normal systolic and diastolic heights. Indeed, the figures suggest that the stoppage is always diastolic, as regards the rhythm, but superposed on a tonic contraction of variable pattern. The fact that there are two types of contraction, rhythmic and tonic, is clearly seen in the records of Tow potassium’ effects. Occasionally, one encounters a heart whose rhythmic mechanism is out of action ; such a preparation will trace a horizontal line in potassium-free with a sharp little drop on changing to low potassium, and a rise on returning. In a normal heart, these effects appear as variations of the diastolic level. In describing the reactions to high potassium, it is best to restrict the terms systolic and diastolic to the rhythmic type of contraction. In this sense stoppage is probably always diastolic.

Comparison of the Aplysia and Helix hearts is interesting. The osmotic pressure of the perfusion fluid is nearly five times greater in the former case than in the latter. The general picture of their potassium relations is, however, identical, provided that the potassium concentrations are given relatively to the concentrations of other ions. Broadly speaking, any mixture which exerts a given effect on the Aplysia heart will exert the same effect on Helix, if it is diluted five times. The factor which determines whether any given concentration of potassium will evoke the Tow’ or ‘high’ type of response is therefore not the absolute value of. that concentration, but the ratio which it bears to some other variable—either to the other external ion concentrations, or to some constituent of the cells; This is of course only a special case of a general rule. Most, or all, rhythmic preparations have essentially similar potassium responses, when the amount of potassium is given in proportion to the other ionic constituents of the medium, whether they are taken from marine, fresh-water or terrestrial animals.

A number of experiments were made on the Helix heart, to find out whether calcium : potassium antagonism would yield an explanation of this rule. The Helix heart is tolerant of a wide range of calcium concentrations. The action of various potassium concentrations was tested, in some experiments with 2, and in others with 15, c.c. of CaCl₂ to 100 c.c. of NaCl (or KCI). It was found that the calcium concentration has a marked influence on the amount of potassium required to produce the ‘high potassium’ type of effect (Fig. 5). In the presence of 2 c.c. CaCl₂ contracture is produced occasionally by 7·5 and always by 10 c.c. KCI to 100 c.c. NaCl. With 15 c.c. Cad, on the other hand, the effectiveness of potassium seems to be roughly halved, for the ‘high’ effect is given occasionally by 15 and always by 20 c.c. KCI to 100 c.c. NaCl. It will be noticed, in Fig. 5, that this antagonistic action of calcium extends to both the tone effect and the rhythm effect of potassium.

Although these experiments indicate that calcium:potassium antagonism is a powerful factor under these particular conditions, they also suggest that other factors contribute to the difference between the two molluscan species, for increasing the calcium concentration seven and a half times has only halved the effectiveness of potassium.

In the Aplysia crop, the ammonium ion closely parallels the potassium ion in its physiological actions. Starting from a potassium-free fluid, a small amount of ammonium chloride produces relaxation of tone and improvement in the rhythm, while a large amount causes contracture (Wells, 1928). Physiological parallelism between the two ions is to be expected, for they are very similar in their physical properties. On the other hand, ammonium chloride solutions contain traces of ammonia, even at only weakly alkaline reactions, and this substance, because of its ready entry into the cells, might exert disturbing actions. In the Aplysia crop the amount of ammonium necessary to produce the ‘high potassium’ effect is about five times as great as the corresponding amount of potassium. Comparable experiments were made on the Helix heart. The ‘low potassium’ effect is easy to produce with ammonium instead of potassium (Fig. 6). The ‘high potassium’ effect, on the other hand, is not evoked, even when half the sodium is replaced by ammonium. The effect of this substitution is to produce a Tow potassium’ response, followed by the gradual onset of a contracture; the latter differs from potassium contracture in its slowness, and in the fact that it endures long after return to ammonium-free fluid. The parallelism between the two ions is therefore less perfect in the Helix heart than in the Aplysia crop.

1. The effects of various potassium concentrations on the cloacal complex of Cucumaria elongata, and on the hearts of Aplysia punctata, Helix pomatia and Car emus maenas are described.

2. It is pointed out that there exists a fundamental similarity between many, and perhaps all, types of rhythmic muscle, as regards their responses to changes in potassium concentration.

3. Data on calcium : potassium antagonism and ammonium : potassium parallelism in the Helix heart are presented.