Storage of Oxygen in the Bladders of the Seaweed Ascophyllum Nodosum and Their Adaptation to Hydrostatic Pressure

The writer’s interest in the facts described below arose out of an experience in salvage work when a sunken submarine had to be marked by buoys moored so as to float on the surface at low water only and to become submerged as the tide rose. When submerged they were naturally exposed to an external pressure corresponding to the hydrostatic head of water above them, and this was often too great for the strength of lightly constructed steel buoys which, in consequence, crumpled up, sank to the bottom and were lost. The difficulty was overcome by charging the buoys with air under a pressure of 10 lb. per sq. in. which, while insufficient to burst them when at the surface, afforded adequate counter-pressure to support their shell plating when submerged. As the floats or bladders of many seaweeds are subjected to similar alternations of hydrostatic pressure as the tide rises and falls it seemed interesting to enquire whether they have evolved any special defence and it was soon noticed that the bladders of Ascophyllum nodosum usually contained gas at a pressure of some 2 – 4 lb. per sq. in. above atmospheric pressure. This seaweed consists of narrow branching fronds which, at intervals of a few inches, are inflated into bladders which are, roughly speaking, of the same shape and size as acorns though often much larger. It grows so densely on rocks that neighbouring individuals must be in competition for sunlight, and an obvious function of the bladders is to float up the plants so as to obtain as much of it as possible. The habitat is between tide-marks, so that for one part of the day the plants are submerged, for another floating, and for the third and generally longest portion stranded high and dry. At high tide (which we will suppose to occur a.m.) the fronds stick up like standing corn (Text-fig. 1A) and what sunlight reaches them through the water, often very muddy, is mainly captured by the outer ranks. At half-tide, when direct sunlight can reach them, most of the fronds are arranged in a thick horizontal layer (Text-fig. 1B) with the bladders of the uppermost tier half out of water and basking in the sun while those of the underlying fronds are heavily shaded. At this phase the efficiency of the bladders becomes of prime importance for, under the sifting action of the wavelets, those with the highest buoyancy shoulder themselves to the top of the floating raft of vegetation and, unless the sea is very rough, will remain there until the whole mass is stranded by the falling tide (Text-fig. 1C), when the upper layer can look forward to some 6 hours of daylight while the undermost, buried under a mat of seaweed some inches thick, is in darkness. This jostling interlude is the critical period of the plant’s tidal day.

It will be shown later that hydrostatic pressure may cause such loss of gas from some bladders as to render them flabby and less efficient as floats than their better adapted neighbours which, in consequence, will override them at every tide and allow them small chance of recovery. It will be further shown that the plants protect themselves from this misfortune, physiologically, by storing a reserve of gas under pressure and, anatomically, by a specific thickening of the bladder walls.

On palpating the bladders of a healthy plant they feel tense as though the contained gas were under pressure, and this impression is confirmed if one is punctured under water when gas bubbles rush out in a characteristic way. The pressure can be exactly measured by the device illustrated in Text-fig. 2.

A hypodermic needle (A) is fixed with sealing wax into the end of a narrow glass tube (C) connected through rubber tubing (D) with a levelling reservoir (E) and the whole system filled with mercury up to the base of the needle. A glass cup (B) is fitted to the top of (C) and is filled with water till the top of the hypodermic needle (which projects into it) is under water. A bladder (A) is cut from the seaweed so that a few millimetres of frond are left projecting from each end, if cut off too closely the bladder will leak. The bladder is then pushed down on to the point of the needle and impaled in such a way that the needle pierces along the axis of the frond till it enters the cavity of the bladder when, if the contained gas is under pressure, some of it will pass through the hollow needle and force down the mercury in the glass tube. The reservoir is now raised till the mercury returns to its former level at the base of the needle, when the height of the top of the mercury in the reservoir above this point will give the original gas pressure inside the bladder in millimetres of mercury. The object of the water in the glass cup is to demonstrate any leakage of gas along the needle track, and if bubbles are seen to escape at the moment of impalement the operation has failed and another attempt must be made with a fresh bladder. To prevent the hypodermic needle cutting out a plug of tissue in the manner of a cork borer and so blocking itself I find it desirable to plug the piercing tip with hard wax or lead and to file a hole for admission of gas in the side of the needle just behind the plug.

It is convenient to express the gas pressure inside the bladder in terms of water head so that it may readily be compared with the external hydrostatic pressure which it helps the bladder to resist, and this has been done on the basis that I ft. of salt water head is equivalent to 23 mm. of mercury.

The pressure found in A. nodosum bladders varies seasonally and from day to day, as it depends on the amount of sunlight caught by the plant under observation ; Table I is a synopsis of the mean pressures found in batches of ten bladders collected at random from the sea wall at East Cowes at different times of year.

From these and other observations it appears that for about 9 months of the year the bladders contain gas under appreciable pressure, the maximum being reached in April when the plant’s reproductive activity is at its height. In the dark months of October, November and December the pressure is zero or slightly negative, a number of the bladders become somewhat flattened, and the whole plant appears to be in a dormant condition. Now the tidal range at Cowes and the distribution in depth of A. nodosum growing there are such that the average hydrostatic pressure on the bladders at high tide is about 4 ft. of water head and the maximum 8 ft., so that for most of the year the gas pressure within the bladders would afford fair counter-pressure and support. It will be shown later that the bladder gas consists, roughly speaking, of air enriched by additional oxygen secreted by the plant, and that its pressure is due to the latter component which constitutes a reserve of gas which can be drawn on for metabolic purposes and also serves to maintain buoyancy at a maximum, in spite of the occasional losses of bladder gas which occur from causes set out in a subsequent section.

Zeller & Neikerk (1915) have summarized the literature on the gaseous contents of seaweed bladders; Willie (1889) found that various species contained from 20 to 37 per cent of oxygen, but maintained that CO₂ was always completely absent. Lucas, working on Australian seaweeds, also failed to detect CO₂ and concluded that the high percentage of oxygen was the result of the oxygen dissolved in the sea “osmosing” into the bladders; Zeller & Neikerk (1915), however, found CO₂ in Nereocystis from Puget Sound in quantities ranging from 2·5 per cent by night to 0-29 per cent by day, and concluded that the pneumatocyst is “not only a float but a reservoir in the gas exchange of the metabolic process”. Langdon (1916), also working on Nereocystis, disagreed with some of Zeller & Neikerk’s findings and made the surprising discovery that the floats of this weed, having a capacity running up to 4 litres, contain carbon monoxide in sufficient concentration to kill a canary in 15 sec. and a guinea-pig in 10 min. In twelve analyses he found CO ranging from 1·1 to 5 per cent, oxygen from 16 to 23 per cent and a little CO₂ but, as it appears that the gas was collected over water, the determination of the last would not be very accurate.

In the case of A. nodosum when the CO₂ content was needed I have collected the gas from a number of bladders over mercury and pooled it so as to provide enough gas for duplicate analyses with the Haldane apparatus, but for analysing the gas in a single bladder have used the Krogh micro-apparatus which though giving the oxygen within 1 per cent is not capable of determining the small amount of CO₂ present.

On a summer day at 9.0 a.m. the pooled contents of a number of bladders gave oxygen 26·8 per cent and CO₂ 0·2 per cent. The tide then rose and covered the plants. At 4.30 p.m., when they were again accessible, fresh samples were taken which showed that the oxygen had risen to 29·8 per cent, the CO₂ remaining at 0·2 per cent at 8.30 p.m., when the rising tide was once more about to reach the weed on which the evening sun had been shining as it hung moist on the sea wall, the oxygen had mounted to 31·9 per cent with CO₂ 0·2 per cent. During the night the oxygen fell so that bladders gathered at 6.30 next morning contained oxygen 27·2 per cent and CO₂ 0·25 per cent. From these and similar observations it can be said that the percentage of oxygen rises during sunlight and falls during darkness, and that secretion of oxygen can take place both when the plant is submerged at high tide and when it is stranded at low tide. In view of Langdon’s discovery a careful search was made for carbon monoxide, but no trace of this or any other combustible gas could be detected and the residual gas is taken to be nitrogen. In October, November and December when, as already stated, the bladders maintain no internal gas pressure they contain what is practically air, the oxygen ranging from 19 to 22 per cent. If artificially cut off from all daylight the plants use up all the oxygen in their bladders and the C0₂ content may rise to 5 per cent, but in Nature I have rarely found the oxygen below 19 per cent, and the highest figure found has been 37 per cent, which is fairly common. The oxygen content of different bladders on the same plant may differ widely according to variations of sunlight and shadow, and in long fronds the distal bladders contain more than the proximal because they are the first to reach daylight as the tide falls and the last to be submerged as it rises. Also while submerged, being uppermost, they get most illumination. This is illustrated in Text-fig. 3 A, showing the gradient of oxygen found in the bladders from top to bottom of a frond about 3 ft. long. To establish that this gradient depends on the relative positions of the bladders and not to the distal ones being younger or more efficient than the proximal, a similar frond was cut off and tied down by the tip to its own stump so that it floated upside down. On analysis a few days later the original gradient was found to have been reversed, the older bladders, now uppermost, containing more oxygen than the younger (Text-fig-3B)

It is found that when the percentage of oxygen in the bladders corresponds with that in ordinary air, so also does the internal gas pressure correspond with that of the atmosphere, and no excess pressure can be detected by the manometer but that when bladders contain a higher percentage of oxygen than does air they also have a correspondingly higher gas pressure. The following examples are taken from a series in which the total gas pressure and oxygen percentage were measured in individual bladders freshly uncovered by the tide at different times of year. If gathered after long exposure to the air or allowed to dry in the laboratory, shrinkage of the bladder wall may raise the pressure anomalously.

The third column in the table is important as showing that the positive gas pressure is derived from secreted oxygen; it was calculated as follows. Taking the last example in the table we see that the positive gas pressure was 7·8 ft. (head of salt water). Now 33 ft. head is equivalent to 1 atmosphere of pressure, so the absolute pressure in the bladder was 40·8 ft. The gas contained 35 per cent of oxygen and (disregarding the trace of CO₂ likely to be present) we may say that there was 65 per cent of nitrogen. The partial pressure exerted by this nitrogen was therefore 65 per cent of 40·8 ft., which is 26·5 ft., which is 80 per cent of 33 ft. or 80 per cent of 1 atmosphere of pressure. The other bladders in the table give similar results showing that, whatever the total pressure within a bladder, the nitrogen component is only exerting the same partial pressure as the nitrogen in the atmosphere or that naturally dissolved in the sea water bathing the plant and presumably is a passive element which has entered the bladder by diffusion.

The bladders do not stretch appreciably so that, once they are full of gas, secretion of additional gas while raising the internal pressure does not increase their volume or add to their efficiency as floats; on the other hand, so long as some internal pressure remains, gas may be lost from the bladder without diminishing its buoyancy. Of course if so much gas is lost as to bring down the internal gas pressure to atmospheric any further loss will diminish the volume of the bladder and the support it gives to the frond.

During darkness the plant draws on the reserve of oxygen stored in its bladders and may use up so much as to reduce the internal pressure to zero, but such physiological loss is easily made up during the day by any plant which gets its fair share of light. A more serious loss of gas may arise if, through an abnormally high tide, the plant is exposed to a greater hydrostatic pressure than that to which it is adapted. This can be imitated by weighting a bunch of the weed and lowering it into deep water for a few hours when, on pulling it to the surface again, the bladders will be found to have become flattened or deeply dimpled (Pl. I, fig. 2) owing to loss of the gas which has diffused out through their somewhat permeable walls. When the tide rises over a bladder or, as in this case, it is lowered into the sea, the increasing hydrostatic pressure is at first countered by the internal gas pressure, but, with increasing depth, the two opposed pressures become equal and the bladder is then in equilibrium with no strain on its walls. Beyond this point the hydrostatic pressure can still increase by several feet without distorting the bladders whose tough walls and smooth oval shape enable them to resist considerable external pressure and to shield their contained gas from it. This gas therefore remains at its original pressure but, with still deeper submersion, a point will be reached where the bladder collapses with formation of a dimple. When this occurs the contained gas will be compressed by the deformation and will take up the pressure corresponding to the hydrostatic head above the bladder. For instance, if a bladder becomes submerged to a depth of 33 ft. (corresponding to a pressure of 2 atmospheres absolute) and fails to resist, the gases in it will assume that pressure. For the sake of simplicity we may disregard the exact proportions of nitrogen and oxygen and call the mixture air. Now the air dissolved in the surrounding sea water is normally at a pressure of about 1 atmosphere absolute, thus the bladder gas (which resembles air in composition) will tend to diffuse out into solution in the sea under a pressure of about 1 atmosphere. In this way losses of gas take place which will increase the dimpling and be more than the plant is able to replace with the amount of sunlight it can catch in its crippled condition, with the result that succeeding high tides squeeze out more and more gas till the bladders are emptied. Obviously the plants which have stored a large reserve of gas are least liable to this disaster but, unfortunately, those which grow at the greatest depth and so are exposed to the highest hydrostatic pressure are the very plants which, from the nature of things, get least sunlight and least opportunity of making and storing oxygen. We have seen (Text-fig. 3) how the oxygen decreases from bladder to bladder going downwards on the same plant and similarly the gas pressures found in plants growing low in the tidal zone are less than those in their neighbours at a higher level. In spite of this disharmony the seaweed seems to protect itself pretty well at places with a moderate tidal range, and at Cowes (with a ⁠. tidal range) one rarely finds a dimpled specimen but it seemed desirable to find out how it fared in spots with a much greater rise and fall of tide.

Commander J. Whitla Gracey, R.N.R., Haven Master of Bristol, was able to tell me that AscophyUum nodosum grew abundantly at Portishead (with a 40 ft. tidal range), and most kindly procured samples for me, while I am indebted to his department for the levels and hydrographic data in Text-fig. 4, which illustrates the conditions at a steep bluff of rock named Battery Point. Seaweeds of all kinds overlap there, but the zone of A. nodosum is quite distinct. The axis of the diagram is the “halftide level” ascertained by meaning the heights of all the high waters and low waters for the year. Similarly the “mean high water” is the mean of the year’s high tides, and “high-water equinoctial springs” is the highest to which the tide ever rises except under the influence of great storms. We see that at the average high water the plants living at the upper margin of their zone are only submerged to a depth of 5-1 ft., but those at the lower margin to a depth of 20 ft., which corresponds to a hydrostatic pressure of 9 lb. per sq. in., while at equinoctial tides they may have to resist 12 lb. per sq. in. At every low tide all the plants are high and dry.

Measurement of the gas pressure in different plants showed that it was in no wise adapted to counter the hydrostatic pressures they had to resist, for plants from the upper margin had an internal pressure corresponding to 11 ft. head of water (more than would ever be above them) while plants from the lower margin had only 3·7 ft. of pressure which could not give them much support against 20 ft. of hydrostatic pressure; but further examination showed that these seaweeds are protected in another way which is graded to suit the depth at which they grow, for when bladders of plants from the lower margin of the zone are slit open their walls are found to be nearly three times as thick as those from plants of the upper margin.

The standard method of measuring thickness for constructing Table III was to pass a small cork borer right through each specimen at its widest point in a direction normal to the plane of the frond so as to cut out a disc from each side, both discs were gauged in a suitable micrometer and, if they differed at all in thickness, the mean was taken.

The thickening or “armouring” shown in Table III enables the bladders of plants in the lower part of the zone to protect their contained gas from the hydrostatic pressure which would otherwise cause it to diffuse away as described on p. 204. The armoured bladders are naturally less buoyant than the thin-walled bladders of the upper part of the zone and consequently afford less support to the frond which, however, compensates by forming them at closer intervals, as may be seen on comparing the two types of plants shown in Pl. I, fig. 1. The necessity for this armouring can be demonstrated by transplanting a bunch of the weed from the upper to the lower margin of the zone, a distance of 15 ft. vertically; the plants may be cut from their attachment and secured in their new position with string, for their roots are merely organs of attachment. In such experiments loss of gas from the unadapted plants can be observed after one tide, and by the third day about half of their bladders will be deeply dimpled. I have not been in a position to watch the process continuously, but have found after 3 weeks every bladder had collapsed into a concave form like the bowl of a spoon (Pl. I, fig. 2), was practically empty of gas and devoid of buoyancy. Even among adapted plants the margin of safety seems to be very small, for, searching along the lowest level of growth, one finds numerous stunted or dying individuals whose bladders have been crushed flat, presumably by some unusually high tide. It seems likely indeed that at Portishead hydrostatic pressure is the limiting factor which determines the downward spread of the plant, for, if one compares the zone of growth as it exists there with the zone at Cowes (Text-fig. 5) it is apparent that, relative to the tidal range, the upper margin occupies the same position at both places but that the lower margin is shifted upwards at Portishead as though to avoid pressure.

The simplicity and directness of this case of adaptation are noteworthy. In a belt of seaweed a few yards wide one finds the adverse environmental factor of hydrostatic pressure increasing from a trifle at the upper margin to a limiting value at the lower, and the plants protecting themselves by thickening their bladder walls in corresponding degree according to the level at which they grow. Those plants which need protection have it, the others do not. At present there is little evidence upon which to decide whether the thickening is a direct response which every plant is capable of making if exposed to severe hydrostatic pressure at the beginning of its life or whether, as seems more likely, it is a manifestation of different constitutions in the young plants which natural selection sifts into the upper or lower part of the zone according to the thickness of the bladders they are capable of developing. In this connexion it may be mentioned that the bladders of plants from the middle of the zone while, on the average, intermediate in thickness vary among themselves in this character more than do their neighbours of the two margins, not only from plant to plant but even from side to side of the same bladder.

I desire to thank Surgeon-Commander F. R. Mann, R.N. for the drawing from which Pl. I, fig. 2 was made.