1. Isolated marginal sphincters of Calliactis and Metridium show little spontaneous activity, but over a limited frequency range give quick facilitated contractions to not less than two stimuli, and slow contractions having latencies of many seconds to multiple stimuli over a wider frequency range.

  2. Earlier observations on the effects of ions on whole animals are confirmed and extended. Excess K+ causes spontaneous quick contractions and responses to single stimuli ; excess Ca2+ has no direct excitatory effects, but causes an immense enhancement of the quick response without affecting the slow response to stimulation ; excess Mg2+ abolishes both quick and slow contractions.

  3. Most drugs, including acetylcholine and its associates, histamine, and 5-hydroxytryptamine, have no effects on either quick or slow contractions. Important effects occur only with tyramine, tryptamine and adrenaline at concentrations of 1 × 10−5 and 1 × 10−4. No inhibitory drug was discovered.

  4. Tyramine has no direct excitatory effects but it greatly enhances the quick facilitated responses without enhancing (and in some circumstances depressing) the slow responses to stimuli. It also causes small responses to single stimuli.

  5. Tryptamine causes direct quick contractions of the sphincter, gives quick responses to single stimuli and somewhat enhances both quick and slow responses to stimuli.

  6. Adrenaline has no effect on quick facilitated responses but causes direct slow responses and some enhancement of slow responses to stimuli.

  7. The effects of ions and drugs are obtained only when they are applied directly to the sphincter region. They do not occur when they are applied only to an attached strip of column.

  8. It is noted that the effects of excess K+ and tryptamine are identical, and that the effects of excess Ca2+ and tyramine are also identical; adrenaline stands alone as a ‘slow-excitor’.

  9. The physiological significance of the results is discussed in relation to different components of the responding system and to the mechanisms of quick and slow contraction in the sphincters.

Ever since Pantin (1935) showed facilitation to be the outstanding physiological feature of the quick closing response of the sea anemone Calliactis parasitica there has been much interest in this response. It is unique in that a single stimulus has no mechanical effect; responses occur only to the second and subsequent stimuli within a frequency range whose lower limit is about one stimulus per 3.0 sec. (17° C.) where the response finally vanishes. There is also a remarkably regular relationship between frequency of stimuli and the size of the individual steps in the ‘staircase’. Yet the mechanism remains mysterious; in spite of some indirect evidence that facilitation is a chemical process at the junctions, direct proof of this is lacking. This indirect evidence comes from studying the effects of temperature on Metridium (Hall & Pantin, 1937), of ions on Calliactis (Ross & Pantin, 1940), and of drugs and extracts on Calliactis and Metridium (Ross, 1945 a, b, 1952). The general conclusion was that neuromuscular transmission is a two-process system, in which the first process, facilitation, is both a prerequisite and a modifier of the second process, excitation of the muscle.

The work described in the present paper repeats and extends the earlier studies using not whole animals but preparations containing muscles that in life close up the anemone, the marginal sphincters of Calliactis and Metridium. Although some results merely confirm what was seen in the whole animals, a number of new effects have been observed. The new information chiefly relates to the slow movements of sphincter preparations (Batham & Pantin, 1954; Ross, 1957a) which are overshadowed in the whole animal by the quick response.

The methods employed were the same as those described in the preceding paper (Ross, 1960). Most of the experiments have been conducted on ring preparations of the margins of Calliactis and Metridium in which the main muscular element is the marginal sphincter. Movements have been recorded on simple spring levers since these muscles, under constant load, tend to stretch indefinitely. These levers permit some shortening but they mainly record changes in tension.

With the sphincter preparations it was possible to use an improved method to test whether substances were having general neurosensory effects or local neuromuscular effects. A twochambered bath was constructed with a diaphragm of tambour rubber separating an inner from an outer compartment (Fig. 1). Sphincters with a full-length strip of column attached were placed in the inner chamber and when the strip of column had been allowed to relax, it was drawn through a tiny slit in the rubber diaphragm. The rubber closed around the joint between the strip and the sphincter and, in most cases, the fit was tight enough to prevent mixing of the fluids in the two chambers. By maintaining a higher level in the inner chamber, it was possible to see whether any leakage occurred. Electrodes were hooked into the end of the strip so that the preparation could be stimulated, as from the base of the column. The sphincter loop was then attached to a lever so that its activity and responses could be recorded.

By this technique it was possible to treat the sphincter region alone by introducing substances into the inner bath and also to record its responses to indirect stimulation. Alternatively, by introducing substances into the outer chamber, the column alone could be treated. By comparing the results in the two situations it could be discovered whether treatments were exerting their effects by acting on or near the sphincter, or whether they were acting on sense organs and/or the conducting nerve net and only affecting the sphincter indirectly.

The properties of the Calliactis sphincter preparation have been fully described in an earlier paper (Ross, 1957a). In the absence of stimulation the preparation, like skeletal muscle, is quiescent. To electrical stimuli at higher frequencies these preparations give admirable quick faciliated contractions to every stimulus after the first. They also give slow responses, which may surmount the quick responses, but which are best seen after a number of stimuli at frequencies too low to cause quick contractions. Examples of these responses are seen in many of the subsequent figures.

The results are presented in a series of tables, each column indicating the effects of the treatments on one property of the sphincter preparation. As before, an attempt has been made to express these effects in a roughly quantitative way. Direct contractions can be easily identified as normally the preparation is quite inactive. The estimate of the number of direct contractions is based on an arbitrary rating in which ‘+’ indicates up to 10 per hour, ‘+ +’, from 11 to 20 per hour, and ‘+ + +’ more than 20 per hour. Tonus is reckoned as ‘+’ if tension is increased by up to 10% of full contraction, as ‘+ +’ for an increase from 11 to 20% and ‘+ + +’ for an increase in tension of more than 20%. If responses to single stimuli did occur, a 20% frequency of such responses was rated ‘+’, 21-40% as ‘+ +’ and more than 40% as ‘+ + +’. Changes in responses to stimulation were rated as ‘+’ or ‘−’ for a 20 % increase or decrease in magnitude, ‘+ +’ or ‘− −’ for a 21-40% change and ‘+ + +’ or ‘− − −’for changes greater than 40% in either direction.

The action of ions

The effects of various ion treatments are summarized in Table 1. There is no need to describe them fully, since in most respects they confirm the earlier results on whole animals (Ross & Pantin, 1940).

K+ causes frequent direct contractions and some responses to single stimuli (Fig. 2 a, b), but the most consistent effect is an early enhancement of the quick response to stimulation (Fig. 2b). There is no sign, however, that K+ causes direct slow contractions or affects the size of the slow response. The tendency for K+ to increase tonus seems to be due largely to its effect in prolonging contractions and inhibiting relaxation. Within a short time (15 min. with K+X4) all responses become depressed and eventually the preparation becomes inexcitable.

By contrast, Ca2+ has no direct contractile or tonic effects on the Calliactis sphincter preparation; however, it causes a huge enhancement of the response to stimulation but without changing the shape of the response (Fig. 3 a). One effect of Ca2+ not observed with whole animals is seen in Fig. 3b, viz. a tendency to respond with a small movement to every stimulus, including the first of a series. These small movements make it difficult to see whether Ca2+ is having any effects on the slow response. The records in Fig. 3 c show slow responses recorded on a very slow drum to twenty-five stimuli separated by intervals of 5-5 sec. at 22° C. Before treatment the latency was standard at about 150 sec., but with Ca2+x4 after 24 min. the response began on the 5th stimulus, i.e. with a latency of about 25 sec. and the final contraction was vastly bigger. Close examination showed, however, that this effect was due to the summation of tiny contractions on each stimulus and was not the typical smooth slow contraction at all, although it developed into the usual type of slow contraction towards the end. There were a number of tests in which Ca2+ produced some shortening of the latency of the slow responses, but it is clear that Ca2+, like K+ exerts its main effect on the quick response and has only a slight effect on the slow response to stimulation.

The direct effects of K+-free sea water on the sphincter preparation are very much like those of excess K+, but with many contractions, and with a maintenance of tension for long periods yet without enhancing or depressing the responses to stimulation very much (Fig. 2c). Ca2+-free sea water, however, is an active depressant; the muscle loses tone and within 5-10 min. ceases to respond at all. As with the column preparations, the whole effect is very similar to that of excess Mg2+. The excitant effects with Mg2+-free sea water on the column were reproduced on the sphincter. Uninterrupted quick contractions made it difficult to test responses to stimuli and to detect genuine responses to single stimuli (Fig. 3d).

Various combinations of excess and lack of two or more ions were also tested and showed the same relations here as with the column preparations, viz. that Mg2+ effects, both excess and lack, cannot be counteracted by increasing or decreasing Ca2+ or K+. Isotonic NaCl (lacking K+, Ca2+ and Mg2+), however, was a powerful excitant, producing big but infrequent quick contractions, and eventually a sustained full contraction.

All these treatments have been tested on the preparations with an attached strip of column in the two-chambered bath described above. The direct effects with excess K+ and Mg2+-free sea water, and the enhancement with excess K+ and excess Ca2+, were only obtained when the sphincter loop in the inner bath was exposed to the treatment. When the strip of column in the outer bath was treated, no direct effects, no enhancement and no responses to single stimuli were observed.

The action of drugs

Most of the drugs used on whole animals and on column preparations have been used again on Calliactis sphincter. A selection of the results is set out in Tables 2 and 3. The experimental procedure was the same as that described for testing drugs on the column preparations.

Many results may be dismissed at once without comment since they are clear negatives. Thus acetylcholine and its associates, eserine, atropine, curare, etc., had no interesting effects. With Whole animals these substances had no effects on facilitation and quick responses (Ross, 1945 a), and we see now that they do not affect the slow responses either. Some other substances of widespread activity and occurrence, for example, histamine, у-aminobutyric acid, tetramethylammonium iodide, are equally ineffective. In fact, only three substances had important effects : adrenaline, tyramine and tryptamine. These require a full description, although their main features may be seen from the tables.

Tyramine

Tyramine produces a remarkable enhancement of the responses of whole Calliactis (Ross, 1945 a) and this appears also in the sphincter preparations (Fig. 4a). This enhancement, however, does not extend to the slow response. Indeed, the tyramine enhancement of the quick response is often accompanied by a depression of the slow response, where the latter appears in controls as a slow delayed movement surmounting an earlier quick contraction (Fig. 4b). It also depresses slow responses at lower frequencies, though at such frequencies small contractions to single stimuli generally appear (Fig. 4 c). These experiments failed to confirm my earlier observation that tyramine causes big responses to single stimuli. Perhaps this was a mistake, or it may be that some factor outside the sphincter was involved. The tyramine effect is unique; 3-hydroxytyramine has no effects at all. In passing, it can be noted that the tyramine effect is very similar to the effect of excess Ca2+, a point which will need to be discussed.

Tryptamine

A note has already been published (Ross, 19576) on the action of tryptamine on both column and sphincter preparations of Calliactis and Metridium. Additional data now permit a more exact description of the effects of tryptamine on the activity and responses of the sphincter.

It has been confirmed that the outstanding feature of the tryptamine effect is a tendency for direct quick contractions to occur. Fig. 5 a shows a section of a continuous record from 15 to 90 min. after introducing tryptamine (hydrochloride— Roche) at a concentration of 1 × 10-4, in which contractions are occurring at the rate of about one per min. The appearance of the effect, both the size and the number of the contractions, is rather unpredictable but the effect is always of this same general type.

These direct effects of tryptamine coincide with a tendency to respond to single stimuli. Fig. 5 b shows a record on a slow drum in which responses to single stimuli were obtained every time the preparation was tested.

The indirect effects of tryptamine on the size and latency of quick and slow contractions are not easy to determine. The tendency to respond to single stimuli makes it virtually impossible to test the effects of low-frequency stimulation and complicates the responses to stimuli at frequencies which give quick facilitated responses. Working with lower concentrations, at which direct effects are not so much in evidence (1 × 10-5), it has been possible to see that tryptamine enhances the quick responses to stimuli. Occasionally with higher concentrations it is possible to get an impressive enhancement, but it never approaches the size of the tyramine enhancement at similar concentrations. The effects of tryptamine on the slow responses of the sphincter of Calliactis are less clear. Many further experiments have thrown doubt on the earlier published statement (Ross, 1957 b) that tryptamine enhances the slow responses.

Other indolealkylamines have been tested on the Calliactis sphincter preparation (Table 3). As reported earlier (Ross, 1957b), 5-hydroxytryptamine (5-HT) is inactive compared with tryptamine; direct quick contractions were seldom seen and only moderate enhancement of the quick response to stimuli occurs at concentrations of 1 × 10-4. On the other hand, 5-methoxytryptamine (5-MT) caused direct quick contractions exactly like tryptamine, though the preparation seemed to be less sensitive to it than to the parent substance. The other derivative available, 5-benzyloxytryptamine (5-BT), like 5-HT, was ineffective. The drugs lysergic acid diethylamide and reserpine, inhibitor and releaser of 5-HT, respectively, on certain other systems (Page, 1958), had no action on the sphincter preparation. Thus the pattern already seen with these indolealkylamines in the column preparations is repeated here, in which tryptamine stands as strongly excitatory, 5-M as excitatory, and 5-HT and 5-BT and drugs associated with 5-HT elsewhere, are all without effect.

Adrenaline

In whole animals adrenaline had no effect on responses to stimulation (Ross, 1945 a), but we now know that it has a powerful action on the column musculature of Calliactis, causing direct contractions and the maintenance of high tonus (Ross, 1960).

Experiments on sphincter preparations have confirmed that adrenaline has no effect on the facilitated response and does not evoke direct quick contractions. However, it does cause slow contractions almost immediately after being introduced, at concentrations of 1 × 10-5 to 1 × 10-4. These contractions look like the slow contractions of the sphincter to low-frequency stimuli and two examples are seen in Fig. 6. In Fig. 6a there is a single small quick contraction at about 3 min. In Fig. 6,b, with a stronger dose, tension develops more quickly and irregularly. These irregularities are not due to quick contractions but to slight variations in the rate of slow contraction. Visual observation shows that a preparation giving quick responses, say when treated with excess K+ or tryptamine, twitches visibly with every contraction. During the experiment with adrenaline seen in Fig. 6 b, the preparation shortened steadily and smoothly without any twitching whatsoever.

Adrenaline also has some effect on slow responses to stimuli. The number of stimuli required to elicit a slow contraction at a definite frequency is usually considerably reduced by adrenaline. Fig. 6a shows a virtually unchanged quick response and an enhanced slow response with adrenaline at 1 × 10-5 and Fig. 6b shows an enhanced slow response immediately after adrenaline at 1 × 10-4.

Apparently adrenaline has a selective action on the slow response mechanism of the sphincter. No other treatment with ions or drugs has had an effect of this kind. Fig. 7 shows the contrast, on a very slow drum, between the direct actions of tryptamine and adrenaline on the same preparation. This contrast is more marked if one is able to observe the preparation during the recording. Tryptamine contractions are visible twitches with a sharp beginning, and are followed by rapid relaxation without any maintenance of tension. Adrenaline contractions are gentle movements, barely visible, beginning almost imperceptibly on the record. Relaxation is slow and incomplete, and tension rises steadily. Quick movements may occur (Fig. 7 shows one example) but they only briefly interrupt the typical picture.

Other sympathomimetic substances have been tested on the sphincter preparations, together with some adrenergic blocking agents, and cocaine, which potentiates many adrenaline effects in vertebrates. The results are shown in Table 2. None of these substances, not even noradrenaline and dopamine, have effects like adrenaline and most of them have no effects at all. Exceptions are : isoamylamine, which has moderately excitatory and enhancing effects ; cocaine, which causes some enhancement of both quick and slow contractions; 933 F, which causes some enhancement of both kinds of response. The last substance was tested on whole Calliactis (Ross, 1945 a) and gave greatly enhanced quick responses and some responses to single stimuli. In the isolated preparations the enhancement was less and no responses to single stimuli were seen.

The adrenaline inhibitors, the adrenergic blocking agents and the potentiating drug cocaine have no effect on the responses, or on the sensitivity of the preparations, to adrenaline. Thus, sensitivity to adrenaline is found without the allied pharmacological properties usually associated with adrenaline-sensitive systems. It is also worth noting that the effects of adrenaline and tyramine have little in common in anemones, whereas in vertebrates, although they differ in some respects, the two drugs have many identical effects.

The three most effective substances, tyramine, tryptamine and adrenaline, were all tested on the ‘strip’ preparation in the two-chambered bath. No effects were obtained when the drug was introduced into the outer chamber containing the strip of column (Fig. 1). Only when they were applied in the inner bath, directly in contact with the sphincter, did the characteristic effects appear. This rules out the possibility of general sensory or nervous excitation and indicates that these substances act on the sphincter itself, or on nervous elements in its immediate neighbourhood.

Batham & Pantin (1954) studied the isolated sphincter of Metridium and found that it gives both quick and slow responses to stimuli. The quick response, however, differs from that of Calliactis sphincter. In Calliactis the quick response, if it is to occur at all, begins on the second stimulus of a series. In Metridium a quick response may occur only after several preliminary stimuli. In my experience four or five stimuli are usually required and it is rare to get a quick response from as few as two stimuli. Moreover, the quick response of Metridium sphincter is usually not a visible twitch as it is in Calliactis. To a series of stimuli, tension develops smoothly rather than in a series of discrete steps. In some ways it is better described as an early than as a quick response ; the movement, although faster than the slow contraction, is much slower than the response of Calliactis sphincter and may continue to develop tension long after stimuli cease. Yet in spite of these differences, the frequency requirements of the quick facilitated response in Metridium sphincter are the same as in Calliactis, i.e. stimuli must not be more than 3 sec. apart or the quick response fails to appear at all.

Slow responses of Metridium sphincter preparations occur either as smooth sigmoid movements following the quick contractions after a definite latent period (usually about 12 min.), or as similar movements at longer latency to stimuli more than 3 sec. apart. This is like Calliactis sphincter. Metridium sphincter preparations, however, show a good deal of inherent slow activity which is not seen in Calliactis. This preparation, therefore, combines a number of features of the general column musculature with a partial development of a capacity for quick contraction ; it may be said to occupy an intermediate position. For this reason it seemed desirable to examine the effects upon it of some of the treatments found to be effective on the other preparations.

Before undertaking these tests it was necessary to collect more information about the responses of the Metridium sphincter preparation. Batham & Pantin (1954) gave some data on this and I have compiled more from my experiments. These are given in Table 4.

One point of interest in this table is that, within the frequency range which elicits quick responses, good slow contractions can be produced in this preparation by small numbers of stimuli that elicit no quick responses, or very minute ones. Thus with five stimuli separated by intervals of either 1.1or 2.2 sec., the quick response is only about one-quarter to one-eighth the size of the slow movement coming in about 12 min. later. With two or three stimuli at these frequencies large slow contractions are obtained although the quick contractions are insignificant. But with ten stimuli separated by intervals of 1.1 sec. almost the whole response is of the quick type (Table 4b). It seems clear from this that the two movements are alternative ways of developing tension in the same muscle and not separate muscle systems whose maximal contractions are additive. It is beyond the scope of this paper to discuss these figures but they show the quick and slow contractions sharing the task of responding to stimuli rather differently as compared with Calliactis sphincter. It would seem that the physiology of quick and slow contraction in these animals might be clarified by further studies on the Metridium sphincter preparation.

The results of treating Metridium sphincter preparations with a selection of ions and drugs are shown in Table 5. These reinforce the general picture of ion and drug effects on column preparations and on Calliactis sphincter. Excess K+ has excitant and tonic effects, but after at first enhancing quick responses it induces a general depression. Excess Ca2+ lacks direct effects but produces a spectacular and long-lasting enhancement of the quick response. Adrenaline causes direct contractions and increases tonus without affecting responses to stimuli, though at higher concentrations the direct effects make it difficult to obtain any exact records of the size or latency of responses. Noradrenaline showed signs of some direct action on Metridium sphincter, in contrast to its ineffectiveness on the other preparations, but it would require more work to decide how much importance should be attached to this observation. Tyramine showed no direct effects but again enhanced the quick response without any comparable effects on the slow response, though a slight effect on the latter was detected. Tryptamine, on the other hand, was followed by direct contractions and an increase in tonus; it also caused some enhancement of the response and shortened the latent period. 5-HT was ineffective.

The effects of Ca2+, tyramine and tryptamine in enhancing the responses are all similar in type. The quick response to a given number of stimuli is not only bigger in the treated preparation but it also begins earlier and is evoked by fewer stimuli. This ‘facilitation number’ (number of stimuli required just to elicit a contraction) seldom fell below three. Unlike Batham & Pantin (1954), I have never seen responses to single stimuli in untreated preparations, nor with any of the treatments used in these experiments.

Examples of these effects on Metridium sphincter preparations are seen in Figs. 8 and 9. It is interesting that although the timing of quick and slow contractions in the preparation is rather different from that in Calliactis sphincter, the effects of ions and drugs are roughly the same. The much slower and delayed ‘quick* or ‘early’ contraction of Metridium sphincter apparently uses the same basic mechanism.

When the effects of ions and drugs on both column and sphincter preparations are reviewed a certain pattern appears. Table 6 summarizes the data on several treatments which have marked positive effects. It shows : (1) the effects of K+ and tryptamine coincide closely; (2) the effects of Ca2+ and tyramine are almost identical; (3) the adrenaline effect is unique and without parallel amongst the ions. Some suggestions can be made about the nature of these effects.

In cases (1) and (2) above, it is reasonable to suppose that effects which are so similar must arise from the same basic actions on the system. Excess K+ generally has depolarizing effects on excitable tissues. Therefore, its effects, and those of tryptamine, on anemone preparations are possibly due to an unspecific depolarizing and destabilizing action on the muscle membranes. Excess Ca2+ has no clear-cut general action on nerve and muscle. In anemone preparations excess Ca2+ and tyramine seem to enhance the contractility of the sphincter muscle when given the stimuli appropriate for quick contractions. Adrenaline evokes slow contractions of both types of preparation, and thus fulfils one of the requirements of a specific slow-excitor substance. The almost complete failure of other substances which act at adrenergic endings rules out any immediate suggestion that neuromuscular transmission in the slow response is an adrenergic mechanism. Yet Östlund’ s (1954) unidentified ‘catechol-4* in Metridium may be linked functionally in some way with the adrenaline-sensitive slow contraction mechanism, and his finding should be followed up.

One of the main purposes of the work was to find out in what ways the quick and slow contractile systems resembled each other and in what other ways they differed. Table 6 shows that, allowing for the difference in time scale, they resemble each other in their responses to one treatment which directly excites nerve and muscle by a general depolarizing action, namely, excess K+ (and probably tryptamine also since its effects are so similar). The two contractile systems are affected similarly by treatments which depress and abolish excitability, excess Mg2+ and Ca2+-lack, though the latter may be due to a relative excess of Mg2+ when Ca2+ is withdrawn Ross (1960). Thus it seems likely that the quick and slow contractions are based on essentially similar conditions of membrane excitability, since their excitability is heightened and depressed by the same treatments in both preparations.

The two types of contraction differ profoundly in the effects on them of the treatments Ca2+ and tyramine, which affect the contractility of the muscles involved in quick responses to stimuli. Thus it seems that the main difference between the quick and slow contractions resides in the contractile mechanism, or in the mechanism by which contractions are evoked. The selective action of adrenaline in evoking slow contractions of both sphincter and column preparations reinforces the view that essentially different processes of initiating muscular contraction are involved in the two types of movement.

It is worth noting that the results provide no evidence that facilitation and slow contraction are affected in the same ways by the same treatments. The facilitation mechanism, if it is to be regarded as controlling the size of the quick response, seems to be most influenced by Ca2+ and tyramine, which affect slow contractions slightly if at all. Adrenaline, the only important slow-excitor substance found in this survey, is singularly without effect on facilitation. Therefore, if facilitation is carried out by a ‘facilitator ‘and slow contraction is initiated by a ‘slow-transmitter ‘, these two substances and their modes of action must be quite distinct. Some preliminary work with extracts of Calliactis on both sphincter and column preparations has given evidence of potent substances with direct tonic effects on the slow contractile system. So far no effects of extracts on facilitation, or on slow responses to stimuli, have been detected. Work is in progress to confirm these preliminary results, and if they are confirmed, to find out what kinds of substances in the extracts are responsible for the effects observed in the preparations.

Although this work has shown that certain ions and drugs act selectively on the mechanisms of quick and slow contraction, these mechanisms seem to be linked functionally in some way. At lower frequencies of stimulation, which produce staircases with small steps, slow contractions take the form of superimposed secondary responses which get bigger as the facilitated response gets smaller (Ross, 1957a). In some Calliactis sphincter preparations the quick contractions are very weak even at higher frequencies, and usually these are surmounted by immense slow contractions, suggesting that the failure of the quick contraction is linked with enhancement of the slow responses which follow later. It is also noteworthy that Ca2+ and tyramine, which selectively augment the quick contraction, tend to depress the slow responses (Fig. 4 b). All these facts point to a complementary relationship between the two responses of the sphincters, as if some transmitter, produced when impulses reach the muscle, contributes something to any quick contractions that occur, and the remainder (or its product) evokes the later slow contraction. In subsequent work it will be important to look for factors which might explain this complementary relationship between the two responses of the sphincters.

Some of this work was done at the Marine Biological Association’ s Laboratory at Plymouth and at the Stazione Zoologica, Naples. My thanks are due to the Directors of these two institutions and their staffs for the facilities I enjoyed and the assistance and kindness I received. The visit to Naples was made possible by a grant of £200 from the Browne Fund of the Royal Society, which I gratefully acknowledge. I also thank Dr B. Heath-Brown of Messrs Roche Products, Welwyn Garden City, for the gift of several derivatives of tryptamine, and Dr H. Blaschko, of the Pharmacology Department, Oxford, for samples of a number of sympathomimetic drugs and adrenergic blocking agents not available commercially, and the last named too for his interest and helpful advice on a number of occasions. Mr L. Sutton provided invaluable technical assistance during the later stages of the work. Prof. D. W. Ewer and Dr H. O. Schild read the manuscript and made many helpful criticisms and suggestions.

Batham
,
E. J.
&
Pantin
,
C. F. A.
(
1954
).
Slow contraction and its relation to spontaneous activity in the sea-anemone Metridium senile (L.)
.
F.Exp. Biol
.
31
,
84
103
.
Hall
,
D. M.
&
Pantin
,
C. F. A.
(
1937
).
The nerve net of the Actinozoa. V. Temperature and facilitation in Metridium senile
.
F. Exp. Biol
.
14
,
71
8
.
Östlund
,
E.
(
1954
).
The distribution of catechol amines in lower animals and their effect on the heart
.
Acta physiol, scand
.
31
,
suppl. 112
, p.
67
.
Page
,
I. H.
(
1958
).
Serotonin (5-hydroxytryptamine); the last four years
.
Physiol. Rev
.
38
,
277
335
.
Pantin
,
C. F. A.
(
1935
).
The nerve net of the Actinozoa. I. Facilitation
.
F. Exp. Biol
.
12
,
119
38
.
Pantin
,
C. F. A.
(
1938
).
Discussion meeting on the transmission of excitation in living material
.
Proc. Roy. Soc. B
,
123
,
397
9
.
Ross
,
D. M.
(
1945a
).
Facilitation in sea anemones. I. The action of drugs
.
F. Exp. Biol
.
22
,
21
31
.
Ross
,
D. M.
(
1945b
).
Facilitation in sea anemones. II. Tests on extracts
.
F. Exp. Biol
.
22
,
32
6
.
Ross
,
D. M.
(
1952
).
Facilitation in sea anemones. III. Quick responses to single stimuli in Metridium senile
.
F. Exp. Biol
.
29
,
235
54
.
Ross
,
D. M.
(
1957a
).
Quick and slow contractions in the isolated sphincter of the sea anemone Calliactis parasitica
.
J. Exp. Biol
.
34
,
11
28
.
Ross
,
D. M.
(
1957b
).
The action of tryptamine and 5-hydroxytryptamine on musclesof sea anemones
.
Experientia
,
13
,
192
.
Ross
,
D. M.
(
1960
).
The effects of ions and drugs on neuromuscular preparations of sea anemones. I. On column preparations of Calliactis and Metridium
.
F. Exp. Biol
.
37
,
732
52
.
Ross
,
D. M.
&
Pantin
,
C. F. A.
(
1940
).
Factors influencing facilitation in Actinozoa. The action of certain ions
.
J. Exp. Biol
.
17
,
61
73
.