The Physiology of the Ascent of Sap

Note the first record exhibiting the minimum due to similar phase in the two contacts. Phase-difference increased by- passage of current as indicated by the enhanced response in the second part of the record (see text) . intensity the speed of the propagated wave is enhanced against the direction of the current, whilst it is retarded in the same direction as the current} We take a stem of Canna and make a longitudinal slit which extends nearly to the top : the two halves are separated from each other by the insertion of a piece of mica, and the separated ends of

the cut stem are wrapped with pieces of moist cloth. The electrodes are placed on two points, A and B, which are in the same horizontal line and are in the same phase before the passage of the current, as is independently demonstrated by the horizontal line shown in the record (fig. 84) indicating an electric minimum. A constant current is now sent into the lower end of the stem at e e^, ascending sdong the left half and descending by the right half. The hydraulic wave is retarded on one side and accelerated on the other : hence a difference of phase is induced at a and b, as shown in the resulting pulsating record. The passage of the current produced a displacement of the base-line (not shown in the figure) ; but this static displacement does not explain the pulsations, which are due to the induced phase-difference. On the stoppage of the constant current, the induced phase- difference disappeared and the record became once more horizontal.

The experiments described show that there is a definite layer in the stem which by its pulsatory activity maintains the ascent of sap : this layer has been localised by the Electric Probe. In dicotyledonous plants it is the inner- most layer of the cortex abutting on the vascular tissue. The dead xylem exhibits no pulsation. The vascular tissue of the xylem is, however, injected with sap by the pulsating activity of the cortex, the intervening distance being very small.

For the vmi-directional propulsion of sap, the further condition of sequence of pulsation or phase-difference is necessary. This has been demonstrated by the method of electric exploration by which the points of electric maxima and minima have been determined. The distance between successive points of electric minima or maxima is half the wave-length of the hydraulic wave. The length of the wave is found to be increased by a rise of temperature, which also enhances the velocity of the ascent of sap.

The cellular pulsations cause a pumping action ; and the sequence of pulsation from cell to cell brings about the unidirectioned flow of the sap. The sap expelled during the contraction of one cell is absorbed by the cell higher up during its phase of expansion. There is a propaga- tion of a wave of contraction, preceded by one of expansion ; in consequence of this the sap is, as it were, squeezed forward. A succession of such waves maintains the continuous ascent of sap.

Initiation of pulsation — Effect of stimulus — Efiect of differential hydro- static pressure — Effect of constant electric current — Effect of variation of temperature on pulsation — Effect of anaesthetics — Effect of dimin- ished internal pressure — Summary. In the endeavour to obtain a clearer general conception of the ascent of sap, as effected by the pulsating cellular mechanism discussed in previous chapters, it may now be enquired, how is the pulsation initiated ?

It has been demonstrated that certain agents enhance the rate of ascent, with consequent increase of turgor of the tissue ; whilst other agents induce a depression or arrest of ascent with a resulting diminution of turgor. These diverse effects must ultimately be due to induced variations in the pulsation* of the active cells. The final analysis would be reached if we could record the waxing and waning of the pulse-throbs of an individual cell under varying changes in the environment.

The pulsating cell has been compared to a pump with alternate expansive up-stroke and contractile down-stroke. The rate of propulsion of water will thus be increased by enhanced frequency or increased amplitude of pulsation ; diminished frequency or amplitude will, on the other hand, cause a diminished rate of propulsion. A more complex effect will be produced when the up- and down-strokes are unequal. It is the up-stroke that sucks in water : hence with a relatively enhanced up- stroke an accumulation of sap will occur in the cell, which will become distended and more turgid. If, on the other hand, the up-stroke is reduced and the down-stroke increased, the result will be a diminution of turgor.

With these preliminary considerations, we proceed to study the effects of external agents on the pulsation of the individual cell, and thus to obtain an insight into the mechanism by which the rate of propulsion of sap is en- hanced or depressed. Nothing, indeed, could be more impressive than the responding movement of the galvano- meter spot of light, revealing the working of the invisible cellular machinery. Under a stimulating agent, for example, the responding spot of light is violently thrown in one direction (beyond the recording plate), and the heightened activity is manifested by the quickened rate or enhanced amplitude of pulsation. Unfortunately, the recording spot of light, on account of its extreme rapidity or great range of movement, leaves little or no trace on the photographic plate. It is only after the first violent outburst has abated a little that the impression made by the moving spot of light can be found on the plate. The record therefore ex?iibits the character of the change, though not its full extent.

The effects of the following external agencies on cellular pulsation have been studied : i. The effect of stimulus in initiation or enhancement of cellular pulsation. iv. The effect of variation of temperature. v. The effect of anjesthetics. A clue to the initiation of pulsation in the ascent of sap ma}' be found in the cell-to-cell propagation of pulsa- tion in cardiac muscular tissue which has become quiescent after isolation. On applying the mechanical stimulus of a prick, the irritation causes an excitatory impulse which is propagated from cell to cell onwards. This is apparently

what happens in the pulsating layer in the plant ; and we will now endeavour to determine what is the stimulus that initiates the multiple activity of the cells. The pulsating activity of the root-cells may be produced in two ways : first, by an increase of internal pressure ; and secondl}', by the continued action of an external stimulus. The increase of turgor of the root-cells by ab- sorption of water from the soil is partially due to osmotic action ; but the mere increase of turgor by absorption will not suffice to ensure the continuous maintenance of pulsation. For we found that the ascent of sap is arrested when the plant, with its root in water, is kept in darkness : under these unfavourable conditions, the cells pass from the active to the inactive state. We also found that the application of a stimulus renews their activity, with the concomitant renewal of the ascent of sap (p. 57).

The root, under normal conditions, is shielded from the stimulus of light, so that source of stimulation is eliminated. It may be stimulated by chemical substances present in the soil; but of this there is no evidence. The remaining possible source of stimulation, which would appear to be the most important, is the mechanical ; the root and rootlets, in boring their way through the soil, are subjected to the constant stimulus of friction. The total surface stimulated is thus very large. Timiriazeff ^ found from calculation that the total length of the root- hairs of a wheat-plant grown in a flower-pot was twenty kilometres (twelve and a half miles). The stimulation over this enormous surface must be considerable, and capable of initiating and maintaining the activity of the root-cells.

This explanation is based upon the results already obtained concerning the effects of stimulus upon rhythmic activity. It has been shown that, while a strong stimulus applied to a highly excitable tissue inhibits its activity, a stimulus of moderate intensity applied to a sub-tonic tissue initiates and maintains its activity. Further experi- ments on the effect of strong stimulus on cellular pulsation were carried out in the following manner. Thermal or mechanical stimulus was applied to a lateral leaf by applica- tion of a hot wire, or by the section of the leaf. There was no immediate effect on normal pulsa- tion, but after an interval of about a minute, required for the excitation to reach the pulsating cells, a very violent contractile response occurred, exhibited by the down-stroke, which went off the recording plate (fig. 85). The recovery to the normal is seen to

Fig. 86. The Effect of Electric Stimulus on Cellular Pulsation of a Sub-tonic Specimen Note the very marked enhancement of ac- tivity after applica- tion of stimulus at arrow. have taken place after a certain interval of time. It will now be clear that the cellular activity in the interior of the plant may be affected by the stimulation of the leaf by light, or by such mechanical stimulation as that caused by the wind, and that the effect of such a stimulation may affect distant organs through an induced variation in the ascent of sap.

The converse process is that of the depletion of stored energy causing a stoppage of cellular pulsation, and of the restoration of activity by the application of stimulus. The experimental specimen was kept inside a dark room for several hours ; the cellular pulsations were now found to have become very much enfeebled. In order to ascertain that this was caused by the lack of stimulus, I subjected the specimen to the electric stimulation of an induction shock of moderate intensity, which was passed along its length. The galvanometer was disconnected during the process, the connection being re-made a minute after the cessation of stimulus. The record (fig. 86) shows the very great enhance- ment in the cellular activity which persisted for a consider- able length of time. The experiment demonstrates once more that it is physiological activity which effects the ascent of sap, and that this activity is maintained by the action of stimulus.

The second factor in the initiation of rhythmic activity is an increased internal hydrostatic pressure. Diminution of pressure inhibits the pulsatory activity of the Desmodium leaflet, and also of growth in growing organs : increased pressure, on the other hand, enhances the activity. Similar effects are observed in the cellular pulsation which main- tains the ascent of sap. In normal conditions, the root- cells absorb water from the soil, \\ith resulting increase of turgor and of internal hydrostatic pressure. The top of the stem is, however, in a state of diminished turgor and internal pressure, due to the transpiration from the leaves. The cellular activity is therefore greater at the root than at the top of the shoot. In cut stems placed in water, a similar difference of activity exists between the lower and upper ends of the stem. This difference is a contributor}^ factor in the determination of the direction of the pro- pulsion of sap, from a place of greater to a place of lesser

activity. As regards the underlying cellular pulsation, it will presently be shown how the diminution of internal pressure causes a depression culminating in an arrest of pulsation. It has been shown how a quiescent rhythmic tissue is roused to activity by the application of stimulus. The passage of a constant current often acts as a stimulating agent ; a very striking illustration of this I found in the initiation of rhythmic activity of the leaflets of Biophytum, originally at standstill, by the passage of a constant electric cur- rent along the petiole.^

The cellular pulsation which causes the ascent of sap is sometimes found to be greatly enfeebled or even arrested. The ex- periment demonstrating the renewal or enhance- ment of pulsation by the passage of a constant current was carried out in the following manner. The lamina of a lateral leaf of Impatiens was made the indifferent point for the second electric contact. The first contact was made on the stem, at exactly the same level as the lateral leaf, so that the passage of a constant current through the stem should not give rise to a difference of potential between the two contacts. In practice there was a very

small difference which caused the displacement of the base-line. A constant electric current was then led through the length of the plant, one electrode being applied at the tip of the shoot, and the other to the soil in which the root is buried. The current is turned on and off by means of a key. A second reversing key enables us to change the direction of the current. The record given in fig. 87 shows the stimulating effect of the passage of a constant current on cellular pulsation. The normal pulsation was very feeble, but passage of the current is seen to have induced a marked enhancement. The starting of the current is independently exhibited by a displacement of the base-line ; the stoppage of the current is followed by the cessation of enhanced pulsation. It is thus seen how it is possible not only to record the elementary pulses, but also put them under external control, the natural condition being resumed immediately on the cessation of the stimulating current. It may be stated here that the stimulating effect of a constant current is modified b}' the direction and intensity of the current.^

The enhanced rate of ascent of sap under rising temper- ature has already been demonstrated by the increased rate of suction and quickened rate of the Erectile Response ; the converse effect of cold has also been shown in the depression or arrest of the Suctional and the Erectile Response (p. 63). Temperature variation has also a very marked effect on the amplitude or the frequency of pulsation. I have been able to diminish or enhance the amplitude by alternate appHcation of cold and warmth. In fig. 88 it is seen that

1 Bose, ' The Influence of Homodromous and Heterodromous Electric Current on Transmission of Excitation in Plant and Animal,' Proc. R. S., B. Vol. 88, 1915. the amplitude of pulsation is very greatly enhanced by a rise of temperature, the activity of the cellular pump being thereby increased. A rise of temperature is often found to produce also an enhancement of the frequency of pulsation. Thus from the record of the Einthoven the Amphtude of Pulsation Normal pulsation before application of warmth is seen to the left.

galvanometer, the period of a single pulsation in a particular specimen at ordinary temperature was found to be twenty- five seconds. After raising the temperature through 5° C. the period was found to be quickened to fourteen seconds. The very great enhancement of the rate of ascent of sap by the application of dilute chloroform has already been described (p. 70), The following experiments were carried out to determine the responsive variation of pulsation by the appHcation of chloroform to the root.

In the first experiment of the series, a small dose of chloroform was applied. The normal pulsations were feeble ; but the stimulating effect of a small dose of the anaesthetic was so great that the record went off the plate in an upward direction, that is, towards expansion and enhanced suction. Fig. 89. Effect of Chloroform, applied at Arrow, on Cellular Pulsation Note preliminary enhancement of pulsation, with prolonged up- stroke ; pulsation arrested on continued application.

The up-stroke was very much longer than the down-stroke, with the result that the base-line was displaced upwards beyond the plate : consequently, the propulsion of water upwards became very rapid. In the second experiment, I applied a larger dose of chloroform. The preliminary effect of the anaesthetic was stimulatory and the amplitude of pulsation became greatly enhanced ; the up-stroke was longer than the down- stroke, and the base-line was raised towards increased positivity, indicative of enhanced turgor. Continued action of chloroform, however, caused a depression, which cul- minated in the final arrest of pulsation (fig. 89).

It has been shown (p. 54) that the effect of diminished internal pressure, due to the application of a plasmolytic solution to the root, is to induce retardation or arrest of the ascent of sap. I will now describe the effect of diminished pressure on cellular pulsation. In the experi- ment to determine the effect of diminished pressure induced by a plasmolysing- agent, the first part of the record (fig. 90) shows that the normal pulsation, though feeble, was uniform. On application of a dilute KNO3 solution to the root, at the point marked with an arrow, we ob- serve a responsive varia- tioninthepulsation which is very characteristic. The down-stroke, which represents contraction, becomes greatly increased, while the up-stroke, which represents suction, is relatively decreased ; the base-line thus declines downwards, indicating a persistent diminution of turgor. The final result is an arrest of pulsation and a throttling of the channel for propulsion, with the consequent arrest of the ascent of sap.

Fig. 90. Effect of Diminished Internal Pressure on the Cellular Pulsations Note down-stroke being longer than the up-stroke. -Showing the Dependence of the Ascent of Sap on Cellular Activity tissues Chloroform (small dose) Chloroform (large dose) . Plasmolytic solution Enhancement Diminution or arrest Renewal or enhance- ment Enhancement Depression or arrest Arrest Enhancement Depression or arrest Renewal or enhance- ment Enhancement Depression or arrest Arrest

Table XXVIII. on the previous page shows at a glance how the ascent of sap is dependent on the pulsation of active cells in the tissues of the plant. Cellular pulsation is initiated and maintained under the action of stimulus of moderate intensity. It is probable that pulsation is initiated in the root, which is stimulated by mechanical friction against the soil. The direction of the excitatory impulse is from the root upwards, giving rise to the sequence of pulsation from cell to cell.

Cellular activity is enhanced by increased hydrostatic pressure. On account of the loss of water by transpiration, a differential hydrostatic pressure exists between the cells of the root and those at the top of the shoot. This is a co-operating factor in determining the direction of pro- pulsion of sap from the region of greater cellular activity in the root to that of lesser activity at the top of the shoot. A constant electric current of moderate intensity is found, under certain conditions, to enhance the amplitude of cellular pulsation.

A fall of temperature causes a depression in the cellular pulsation, culminating in arrest and the consequent stoppage of the ascent of sap. Rise of temperature, on the other hand, enhances the frequency or the amplitude of pulsation. The activity of the cellular pump is thus greatly increased, with a resulting enhancement of the rate of ascent. The effect of a small dose of chloroform is to induce a very great enhancement of the amplitude of the cellular pulsation ; in the first stage of its action the up-stroke, which represents suction, is relatively long ; the vertically situated cellular pumps serially take up this enhanced suction, and the propulsion of water upwards becomes very rapid. Continued action of the anaesthetic causes a stoppage of pulsation, with the consequent arrest of the ascent of sap.

A diminution of hydrostatic pressure by application of a plasmolysing solution to the root induces an arrest of cellular activity. The characteristic effect on cellular pulsation is that the down-stroke, which represents con- traction and expulsion, becomes greatly increased ; while the up-stroke, representing expansion and suction, is reduced. The general result is a throttling of the channel for the conduction of water, with the consequent arrest of the ascent of sap.