OBSERVATIONS ON THE PHYSIOLOGY OF THESWIM BLADDER IN CYPRINOID FISHES

BY H. M. EVANS AND G. C. C. DAMANT.

(Received 2.6th February 1928.)

(With Five Text-figures.)

THE swim bladder of fishes is primarily a hydrostatic organ and with rare exceptionscontains or tends to contain the exact quantity of gas which is necessary to make thespecific gravity of the whole fish equal to that of the water in which it is swimming,so that it can rest in mid-water tending neither to rise nor sink: this normal con-dition is called neutral buoyancy. Since gas is compressible and water is not, anyincrease of external or atmospheric pressure by acting through the non-rigid bodywalls will reduce the volume of gas in the swim bladder and cause the fish to sinkin the water (condition of negative buoyancy). This condition can also be producedby aspirating some of the gas from the swim bladder or by attaching a small weightto the fish.

When in a state of negative buoyancy produced by any of these treatments(provided that the interference has not been excessive), the fish will compensate(i.e. restore its neutral buoyancy) by introducing additional gas into its swimbladder. In fish with closed swim bladders (Physoclisti) it has long been known thatthis additional gas is mainly oxygen and is secreted into the swim bladder byorgans known as red bodies or gas glands. Such organs are absent in many fisheswhose swim bladders are furnished with ducts communicating with the exterior, andexperiments which have been published on the method by which such fish compen-sate are inconclusive.

Working with such Cyprinoids as Carp, Roach and Goldfish, which have a longpneumatic duct leading from the oesophagus to the posterior sac of the swim bladder,we find that when a condition of negative buoyancy is induced artificially they combatit by rising to the surface, taking air into the mouth and passing it thence to theswim bladder. If they are prevented from doing this an alternative remains, for wefind that in spite of the absence of red bodies they can slowly secrete a gas rich inO2 into their swim bladders. The following experiments demonstrate these facts.

When fish are observed in glass sided aquaria it is possible to judge with greatexactness whether they are in neutral buoyancy and if not how far compensation hasproceeded, provided that incautious movements or changes of illumination arenot allowed to scare them into rapid excited swimming.

Two small goldfish were placed in a corked glass bottle containing water(Fig. 1). The cork was pierced by a tube at which the experimenter sucked with his

Physiology of the Swim Bladder in Cyprinoid Fishes 43

mouth till the reduction of atmospheric, pressure (by about 25 cm. Hg) was seen tocause the escape of a bubble or two of air from the swim bladder via the pneumaticduct and mouth. They were then quickly transferred to a glass aquarium (Fig. 2)where one was imprisoned under a bell-jar completely filled with water and so cutoff from access to the surface while the other was left free to promenade and "takethe air." The fish were much agitated and showed marked negative buoyancy,dropping "like stones" to the bottom at each temporary cessation of activeswimming. After five minutes both were trying to reach the surface; the prisoner

J)

1

Fig. 1.

could only knock its head against the glass dome, but the free fish came to the sur-face and took gulps of air then dropping back to the bottom where it spat out a fewbubbles, repeating the performance at short intervals. Some of the air remainedinside for the negative buoyancy of this fish steadily diminished and had disappearedin i i hours. Meanwhile the prisoner retained its negative buoyancy, though after24 hours it was thought to be somewhat less heavy than at the start; after 48 hourscompensation had progressed further but the negative buoyancy was still so greatthat the fish rested on the bottom and fell there (though less heavily than at first)after each swimming effort had ceased.

Air was now blown up under the bell-jar so as to form a large bubble at the top.

44 H. M. EVANS arid G. C, C. DAMANT

The prisoner immediately swam up, gulped air in the same way as the free fish haddone and, in less than two hours it had fully compensated and was resting com-fortably in mid-water (neutral buoyancy).

In another type of experiment negative buoyancy was produced by puncturingthe swim bladder and removing a certain quantity of gas. Roach were placed in alarge tank in which there was a constant flow of water. A vertical wire netting par-tition divided the tank into two equal compartments one of which was provided witha wire netting cover resting about one inch below the surface of the water. The fishconfined in this half are called non-access fish because they cannot reach the surfaceor gulp air while the fish in the other half are called free-access fish.

Fig. 2.

The puncture was made in all the experiments at a point 2 scales above the lateralline at a distance of 7 lateral line organs posterior to the head. This corresponds withthe centre of the anterior sac. They were punctured with the needle, of a hypodermicsyringe and approximately 2 c.c. of gas removed from the swim bladder of each.Eight of them were then placed in the free-access and seven in the non-access com-partment of the tank. All showed marked negative buoyancy. After two hoursthree of the free-access fish were partially compensated and were observed to beswimming on the surface at frequent intervals with \ inch of the head exposed; theygulped air and spat out froth. After seven hours all the free-access fish were poisedfreely in the tank and swam without effort in a condition of neutral buoyancy whilethe non-access fish were observed to be swimming up to the wire netting and thencoming down tail firsts still showing marked negative buoyancy and resting hori-zontally on the bottom or at an acute angle with their tails touching it.

After 24 hours three non-access and three free-access fish were killed and the

Physiology of the Swim Bladder in Cyprinoid Fishes 45

gas from their swim bladders collected over water and carefully measured when itwas found that each of the free-access fish contained more gas per gramme of bodyweight than did any of the non-access fish, the average difference being between 3and 4 cc. in the case of 100 gm. fish.

The subsequent history of the remaining fish was as follows. After 29 hours thefour non-access fish were still in negative buoyancy and trying to reach the surface;at one moment all four were swimming upwards in an almost vertical attitude. Thefish with free access naturally remained in neutral buoyancy.

After 48 hours two of the four non-access fish appeared to be nearly, if not quite,compensated; all four could move more freely and no longer swam obliquely withthe head pointing upwards. After 72 hours two were in neutral buoyancy but twostill tended to rest on the bottom. After 112 hours all four had neutral buoyancy.Summing up, we see that of four fish without access to the surface two requiredbetween two and three days to compensate and two required more than four days,while all eight of those with access to the surface compensated within seven hours.

Further experiments showed that the period of compensation in non-accessfish is variable and may be much longer; we have cases in which neutral buoyancywas not attained till the 16th, 28th and even the 48th day.

The following experiment was devised to study more closely the feeble andvariable power of compensation which the foregoing had shown us was possessedby Cyprinoids prevented from gulping air. In it negative buoyancy was produced bya more natural method than aspiration and means were provided for measuring theprogress of compensation with some accuracy. Three goldfish were confined in alarge aspirating bottle supplied at its base with running water from a tap. Afterpassing through the bottle the water escaped through a discharge pipe which couldbe raised or lowered so as to vary the hydrostatic pressure within the bottle betweennine feet of water as a maximum and six inches which was the depth of the con-tainer.

With no access to air {i.e. when the aspirating bottle was completely filled withwater (Fig. 3, A) the fish under pressures of a few feet of water showed negativebuoyancy and uneasiness with the usual signs of wanting to gulp air. After halfan hour the discharge tube was lowered and the pressure reduced to normal,whereupon two of the fish started to float upwards with positive buoyancy showingthat they had already partly compensated towards the increased pressure. Bysimilar experiments it is found that compensation goes on slowly and in about sixhours fish can compensate to five feet of water pressure, but in two experimentswhere the fish were left under a head of eight feet of water for 24 and 48 hoursrespectively they remained in negative buoyancy, though by lowering the pressureit was found that they had compensated to five feet but apparently could carry theprocess no further in the time. (It will be understood that the fish are kept under thehigh pressure continuously except for the intervals of about five minutes' durationwhen, by lowering the discharge tube till the fish showed signs of positive buoyancy,the degree of compensation was gauged.) On varying the experiment so that a largebubble of air was trapped over the running water in the aspirating bottle (Fig. 3, j?)

46 H. M. EVANS and G. C. C. DAMANT

the fish were seen to make use of it. Almost directly the pressure was raised theycame to the surface, gulped air in the usual way and compensated to eight feetpressure in the space of five minutes or so.

Discharge whichcan be raised

or lowered

9 Feet of rubbertube

9feet of rubbertube

t

Fig. 3-

It might be suggested that fish which compensate by swallowing air store it inthe stomach or intestines and not in the swim bladder, but careful examinationsmade to decide this point have always revealed a total absence of gas in the digestivetract. For instance, a Roach weighing one pound was punctured in the anteriorsac of the swim bladder and between 3 and 4 c.c. of gas aspirated. The fish was

Physiology of the Swim Bladder in Cyprinoid Fishes 47

observed to regain neutral buoyancy by gulping air in the course of three quartersof an hour. It was then killed and dissected under water. On opening the ab-dominal cavity two or three tiny bubbles of gas escaped from the peritoneal cavity;no doubt this gas had escaped there from the punctured swim bladder. The in-testines and stomach were then removed without interfering with the pneumaticduct. No gas escaped as they were removed and no gas was apparent as they weredivided with scissors throughout their length.

COMPOSITION OF THE SWIM BLADDER GAS IN CYPRINIDAEAND OTHER FRESHWATER FISH.

We have analysed the gases from-normal Roach and Bream; the body of thefish was completely immersed under water while the sample was collected throughan aspirating needle over mercury. The analysis was done in a Haldane portableapparatus. About half the samples were examined for combustible gases but as notrace of such was ever found the residual gas is assumed to be nitrogen in all cases.

Table I.Analyses of gases from the swim bladders of Cyprinoid fish.

V^g V /O/

/io-5

Roach ,

8-36-8

io-83'33'93"O

\ 3*5

CO2(%)4-2

2'54*44*32"32"O2*32" I

Bream

Average 6-o- 2*8 N2 (by diff.) 91*2%.

The Bream was caught on a line and its gases collected in a floating laboratorywithin a few minutes of capture. The gases from Roach were removed a few daysor hours after capture, the fish having been meanwhile confined in a large tank withconstant supply of fresh water.

F. G. Hall (1924) to whom we are indebted for the most recent work on swimbladder gases, gives the results of analyses of 12 samples from (American) Carp.The average was

O2 5*7 per cent. CO2 3*7 per cent.,

which is strikingly similar to our own determination.His series included three samples with the O2 between 7 and 8 per cent., one

over 12 per cent, and the remainder below 5 per cent, which corresponds with therange of variation found by us.

In the American Perch, which has a closed swim bladder and cannot swallowair, he found the gas to contain

O2 19-9 per cent. CO2 0*63 per cent,(average of 28 samples).

H. M. EVANS and G. C, C. DAMANT

He then proceeded to investigate the effect of removal of gas by taking a secondsample from each fish some hours after the first and found that in Perch the oxygenpercentage had risen to nearly double its original volume while in Carp there waslittle change. He remarks "The Carp has an open duct leading from the swimbladder while the Perch has a closed swim bladder. This is believed to be the ex-planation of the difference in response of the two types to the withdrawal of air fromthe swim bladder" and with this comment leaves the matter.

With Roach, allowed free access to the surface after being aspirated, we getresults closely resembling those of Hall with Carp, but if the fish are prevented fromgulping air the difference is striking.

Approximately equal quantities of gas were removed from the swim bladders ofseven Roach by aspiration with a hypodermic syringe. The fish were then placed inthe experimental tank described on p. 44, four of them being cut off from the surfaceby wire netting and the other three, as controls, being able to reach it and gulp air.When the process of compensation was seen to be complete, which in the case ofthe non-access fish was after some days, they were killed and the gases drawn offand analysed.

Table II .

Comparison of gases found in the swim bladders of Roach after compensating with andwithout free access to the surface.

Average

Non-access fishO2%23-824-826-827-5

25-7

co22-33-23-84-4

3-4

Free-access fish7-87-6

462-32-S

Average 8-9 3-1

These results show that Roach with negative buoyancy requiring gas to inflatetheir swim bladders are able, if prevented from reaching the surface and gulping it,to secrete oxygen to about the same extent as do Perch which are Physoclisti withprominent red glands. In Roach no gland or modified epithelium has been de-scribed on the inner coat of the swim bladder. The process is however very slow ascompared with fish possessing gas glands, for instance, Woodland (1911) found thatPollack of about the same weight as our Roach could secrete 5 c.c. of gas in lessthan 12 hours.

Physiology of the Swim Bladder in Cyprinoid Fishes 49

THE PRESSURE OF GASES IN THE SWIM BLADDER OF CYPRINIDAE.

If a Cyprinoid such as a Minnow or Roach which has been kept for weeks orperhaps its whole life in water no more than six inches deep be killed and opened, itwill be noticed that the swim bladder is tense and feels quite hard, because the gasescontained in it are at a pressure considerably above that of the atmosphere plus thesix inches hydrostatic pressure which is the maximum external pressure of thefish's recent environment.

That such a pressure exists during life becomes evident when gas is drawn offfrom the swim bladder as in the experiments described on p. 44. In none of thefish was it necessary to suck with the syringe, for as soon as its needle entered thebladder the piston was driven to the top by the pressure of the contained gases.

We have found no trace of this phenomenon in non-cyprinoid fish such asPerch whose (closed) swim bladders in the same circumstances are quite soft to thetouch; the contained gases being at atmospheric pressure.

The excess pressures in the swim bladders of Roach have been measured byintroducing a hollow needle connected with rubber tubing (filled with liquidparaffin) to a "Tyco" pressure gauge. The fish were rapidly killed, generally bydestroying the brain, and the needle thrust into the swim bladder, either more orless blindly through the flank of the entire fish or carefully after opening the bodyand exposing the swim bladder. In the first case time is saved, but readings of

Table III.

Gas pressures in swim bladders of Normal Roach.

Method of killing

Pithed5>

>>

>>

>>

>>

>>

>>

»>

>>

>>

>J

J>

»>

>>

>>

>>

Etherised

H.C.N.

Body opened or not

Yes>>j>

>>

5>

>>

3>

»>

>>

»>

>>

J>

J>

>>

M

Noj>

*9

yy

>>

»>

Yes

Pressure in mm. of Hg

Anterior sac

706790428070506040707250542038

44

83"

58536264

Average 59

1

Posterior sac.

€80

12080

70904080

4054203840

84

58

64

67. IS

62

BJEB-Vli

50 H. M. EVANS and G. C. C. DAMANT

pressure may be too low through gas escaping from the bladder into the peritonealcavity at the side of the needle puncture. In the second case leakage is prevented bydoing the operation under water and carefully inserting the needle through a thickmuscular part of the bladder wall; on the other hand the effect of the necessarymanipulation on the contractile bladder walls may cause some alteration of pressure.

Table III gives the results of a number of experiments, the method by which thefish was killed is indicated and also whether the body was opened before measuringthe pressure or not. In most cases the pressure in the two sacs into which the bladderis divided was taken separately. Generally they were much the same within thelimits of possible error but in some cases there was a difference showing that com-munication between the two sacs had been cut off by closure of the sphincter aspreviously described by Evans.

We find then that when Roach are kept in water less than a foot deep the averagepressure in their swim bladders will be 60 mm. Hg or 2§ ft. of water pressure.A reduction of atmospheric pressure by about 60 mm. Hg which is equivalent toraising the bladder pressure by a further 60 mm. Hg will generally cause the escapeof a bubble of air from the bladder via the pneumatic duct and mouth, so thatspeaking figuratively the safety valve in the average Roach is loaded to about120 mm. Hg. It would be interesting to know whether when such fish are livingat a greater depth, say four feet of water, they still maintain a bladder pressure inexcess of that due to hydrostatic head, but we have no satisfactory means of de-termining the point. Undoubtedly the pressure is partly due to tone in the musclesof the bladder wall which have been described by Evans, but it is difficult to decidein what way it is advantageous to the fish. Apparently it is incompatible with theexistence of such structures as the red glands of the Physoclisti, for with a bladderpressure in excess of the aortic blood pressure it is difficult to see how bloodcould reach them. The Roach's bladder wall is so impermeable to gases thatwhen dissected out and left on the laboratory table in a dish with a little salinesolution to keep it moist, the pressure within does not appreciably fall till putre-faction sets in after two or three days. Ability to compress the contents of thebladder as required would obviously enable a fish to counteract the effect of changesof hydrostatic pressure on its buoyancy within a limited range: thus a Roach livingin a pond three feet deep might keep its swim bladder contents at a constantpressure of 67 mm. Hg (and therefore at a constant volume), maintaining this bymuscular exertion when at the surface and allowing hydrostatic pressure to takeover part of the work at greater depths till at the bottom the muscles could be fullyrelaxed. The result would be that the fish's buoyancy would be the same at alldepths in that pond.

We do not believe this to be the right explanation. Goldfish with fairly highbladder pressure when confined in the apparatus shown in Fig. 3 and suddenlyexposed to an increase of pressure of no more than 18 inches of water show signsof negative buoyancy and discomfort which they try to remedy by swallowing airin a way that certainly does not suggest that the muscular mechanism indicatedabove is available. Again if one attaches small pieces of cork or lead to fish so as to

Physiology of the Swim Bladder in Cyprinoid Fishes 51

upset their neutral buoyancy, they will compensate within limits by altering thevolume of gas in their swim bladders, and measurements of the pressure shouldshow if the bladder muscles were used to assist the process. The following experi-ment was done to investigate the point.

A batch of Roach of average weight 100 gm. were divided into two groups ofsix one lot being kept as controls, while to each fish of the other a small cork floatwas attached. After five hours all the fish were killed and the bladder pressuresdetermined.

The pressures in the corked fish, being much below the normal, demonstrateclearly that compensation was effected by expulsion or absorption of gas and not bycompression of the original quantity of gas into a smaller volume. Finally the gaseswere squeezed out of the bladders of all the fish and measured, when it was foundthat the corked fish only had 6-5 c.c. per 100 gm. of body weight and the controls9-1 c.c. per 100 gm. of body weight. We obtained a similar decisive answer fromfish to which small lead weights had been attached. In these the pressure rosemuch above normal.

Table IV.

Pressure of gases in swim bladders of " Corked" fish and Controls.

" Corked " fish Controls(mm. Hg) (mm. Hg)

25 7033 8022 4420 9025 6340 50

Average 27 66

A chance observation is worthy of record as illustrating how the buoyancy canbe affected by the bladder pressure. A number of Roach that had been kept in atank with about one foot depth of water for nearly three months became infectedby fungus disease and died: soon after death some were seen to be floating on thesurface while others lay on the bottom. The average pressure in seven fish foundfloating was 30 mm. Hg while the average pressure in three which were lying deadat the bottom of the tank was 90 mm. Hg. Apparently the fish floated or sankaccording to whether their bladder muscles contracted or relaxed about the time ofdeath.

Since the bladder pressure does not seem in life to be a means of regulatingbuoyancy we can only indicate one other way in which it might be useful. Since asfar as we know the high pressures described above are confined to the Cyprinidaewhich are one of the group of fishes possessing the Weberian mechanism, it isnatural to suppose that the two are connected, and we suggest provisionally thatthe proper working of the Weberian ossicles demands a certain tension in the bladderwalls which is normally maintained by the reaction of the contained gases againstthe compressive force applied by the bladder muscles.

4-2

52 H. M. EVANS and G. C. C. DAMANT

ANATOMY OF THE PARTS CONCERNED IN INFLATION OFTHE SWIM BLADDER.

Evans (1925) has described and figured the muscular bands and sphincters andnerve supply of the swim bladder in Cyprinidae, including the sphincter at the exitof the pneumatic duct from the posterior air sac and the nerve plexus at its junctionwith the oesophagus. One of the objects of this paper is to describe the bulbousenlargement of the pneumatic duct which occurs at this point and appears to con-stitute a definite organ concerned with the inflation of the swim bladder. A similarstructure occurs in the Gymnotidae and John Hunter (1861), after describing thetwo separate "air bags" of Gymnotus electricus, mentions that their ducts unite toform a common passage which becomes cellular and then opens into the oeso-phagus. Sections of this cellular portion of the common duct show a structure similarto that found in the bulb of the pneumatic duct in Cyprinidae.

We propose to call this organ the pneumatic bulb. Guyenot (1909), describingit, writes "the pneumatic duct for 1 to 1-5 cm. presents a fleshy enlargement withlongitudinal and circular striated muscle continuous with that of the oesophagus....In the lumen of this enlargement are a number of blind diverticula separated intocompartments and lined by villi like those of the oesophagus." He then proceedsto state that "the small diameter of the pneumatic duct and the abundance ofmucous secretion contributes, with the muscular sphincter, to form an apparatusnot only opposing the penetration of external air but also, to a certain extent, theexit of air from the swim bladder, and that the muscles of the pneumatic sphincterand particularly the striated muscles have a tonic contraction which prevents egressof air and makes the penetration of air impossible at least in Cyprinidae."

He finds that "the minimum pressure required to produce exit of air is 65 cm.of water, and for the entry of air 195 cm. of water." We believe that the foregoingexperiments prove that the entry of air is not only possible but is the most rapid andusual method adopted by these fish to inflate the swim bladder. The anatomicalstructure of the pneumatic bulb is not only consistent with this function but ispresumably evolved for the special purpose of enabling swallowed air to be pumpedinto the swim bladder.

The organ has been examined by one of us (H.M.E.) in Roach, Tench, Bream,Minnow and Carp and a similar type of structure is present in all of these. Thespecimen now described was taken from a Roach weighing six ounces. The pneu-matic duct in its passage forward from the posterior sac of the swim bladder be-comes gradually thicker at a point about 4-8 mm. before its junction with the wallof the oesophagus. Fig. 4 is a diagrammatic longitudinal section of these parts andwill enable the reader to follow the structure of the duct as it passes through thewall of the oesophagus.

Starting at the narrow entrance of the duct into the oesophagus, it is seen todilate and form a considerable cavity occupying three quarters of the thickness oflie oesophageal wall. Next follows a narrower portion into which several diverticulapen: these diverticula run parallel to the long axis of the duct. The total length of

Physiology of the Swim Bladder in Cyprinoid Fishes 53

the duct lying within the wall is 4 7 mm. so that the total length of the pneumaticbulb is approximately 1 cm. About 0-3 mm. before joining the gullet and for adistance of about 07 mm. within the oesophageal wall, the duct breaks up into aspongy portion the total length of which is therefore about i-o mm.

The opening into the gullet is surrounded by a sphincter muscle and the ductthroughout its course is surrounded by a thick investment of striated muscle con-tinuous with the striated muscle of the oesophageal wall. The mucous membrane ofthe oesophagus is continued throughout the anterior portion of the duct for about4-0mm. and presents numerous "taste buds" surrounding the orifice, but the last0-7 to O'S mm. of the duct within the oesophageal wall presents an entirely differentstructure as described above, it has here a spongy structure and is now lined withcolumnar epithelium.

T IS

LUMEN OF (ESOPHAGUS

Fig. 4. Longitudinal section of wall of gullet through oesophageal bulb (diagrammatic).

Fig. 5 is a detailed drawing under a quarter-inch objective of the pneumaticbulb of the Tench before its junction with the oesophagus. The lumen of the mainduct is shown and one large diverticulum surrounded by circular muscle fibres,external to which are longitudinal muscle bundles.

Apparently in the pneumatic bulb we have an air pump with its entranceguarded by "taste buds" and a sphincter,and its main cavity lined with mucousepithelium and provided with powerful striated muscular walls. Between this partand the duct proper is interposed a spongy labyrinth which evidently might act asa filter.

It is interesting to compare the situation of the taste buds in the above fisheswith that in mammals. In man they are found at the base of the tongue on thesides of the papillae vallatae and also on the fungiform papillae. According toSchafer (1894) t n e y are specially numerous over a small area just in front of theanterior pillar of the fauces, also on the anterior surface of the soft palate and theposterior surface of the epiglottis.

In the process of swallowing the post-nasal space and the entrance into the

54 H. M. EVANS and G. C. C. DAMANT

larynx have to be cut off so that food can pass only into the oesophagus. The tastebuds appear in just those positions where one would expect sense organs to beposted as sentinels to protect the air cavities. In Cyprinoid fish we find taste budsat the orifice of the pneumatic bulb, and conclude that they are there as part of themechanism for preventing food from entering it.

We desire to express our gratitude to Prof. A. E. Boycott for allowing us to workin his laboratory at University College Hospital and for much valuable help andmany suggestions.

Fig. 5. Pneumatic bulb of Tench before its junction with oesophagus, showing duct and one dir-verticulum and circular and longitudinal muscle bundles. J in. obj.

SUMMARY.

1. Experimental proof is given that Cyprinoid fish can inflate their swim bladderseither by swallowing air or, far more slowly, by secreting a gas rich in oxygen,although they have no gas glands in the ordinary sense of the term,

2. Attention is directed to the high gas pressure which such fish maintain intheir bladder and its probable use is discussed.

3. The anatomy of the pneumatic bulb at the oesophageal end of the pneumaticduct in Cyprinoid fishes is described.

Physiology of the Swin Bladder in Cyprinoid Fishes 55

REFERENCES.

EVANS, H. M. (1925). Proc. Roy. Soc. B, 97, 545-576.GUYENOT, E\ (1909). Bull. Sci. Nat. de France et Belgique, 43, 203—296.HUNTER, J. (1861). Essays and Observations, ed. by R. OWEN, p. 419.HALL, F. G. (1924). Biol. Bull. Woods Hole Mass. XLVII, pp. 79-115.SCHAFER, E. A. (1894). Quain's Anatomy, vol. in, pt. 3. Tenth ed. p. 149.WOODLAND, W. N. F. (191 I ) . Anatamischer Anzeiger, 40, 225-242.