The use of alizarin red S to detect and localize calcium in gametophyte cells of ferns

An aqueous solution of alizarin red S containing chloral hydrate both clears intact chlorophyllous gemma cells of Vittaria graminifolia and stains for protoplasmic calcium. Verification that the stain was protoplasmic rather than in the cell wall was shown by a positive reaction in extruded protoplasm. Similar staining was found in extruded protoplasm of Onoclea sensibilis spores. Differentiating gemma cells show localized protoplasmic accumulations of Ca2+ at sites where asymmetric cell divisions initiate the formation of rhizoids, antheridia or vegetative cells. The staining properties of the dye depend on careful control of pH and the addition of appropriate amounts of KCl to the mixture. Treatment of Onoclea spores and Vittaria gemmae with 100 mM EGTA for 30 min nearly abolishes staining of their extruded protoplasts and also of intact cells of gemmae. The use of alizarin red S with and without chloral hydrate demonstrates different pools of protoplasmic Ca2+. When Onoclea spores are ruptured to extrude the protoplasm, both dye mixtures stain a peripheral, granular protoplasmic component. However, the chloral hydrate-containing dye also reveals Ca2+ associated with small particulate protoplasmic components. Extruded protoplasm of gemma cells stains intensely with alizarin-chloral hydrate, but does not stain with alizarin lacking chloral hydrate.     

The role of water in protoplasmic permeability and in antagonism

The behavior of the cell depends to a large extent on the permeability of the outer non-aqueous surface layer of the protoplasm. This layer is immiscible with water but may be quite permeable to it. It seems possible that a reversible increase or decrease in permeability may be due to a corresponding increase or decrease in the water content of the non-aqueous surface layer. Irreversible increase in permeability need not be due primarily to increase in the water content of the surface layer but may be caused chiefly by changes in the protoplasm on which the surface layer rests. It may include desiccation, precipitation, and other alterations. An artificial cell is described in which the outer protoplasmic surface layer is represented by a layer of guaiacol on one side of which is a solution of KOH + KCl representing the external medium and on the other side is a solution of CO₂ representing the protoplasm. The K⁺ unites with guaiacol and diffuses across to the artificial protoplasm where its concentration becomes higher than in the external solution. The guaiacol molecule thus acts as a carrier molecule which transports K⁺ from the external medium across the protoplasmic surface. The outer part of the protoplasm may contain relatively few potassium ions so that the outwardly directed potential at the outer protoplasmic surface may be small but the inner part of the protoplasm may contain more potassium ions. This may happen when potassium enters in combination with carrier molecules which do not completely dissociate until they reach the vacuole. Injury and recovery from injury may be studied by measuring the movements of water into and out of the cell. Metabolism by producing CO₂ and other acids may lower the pH and cause local shrinkage of the protoplasm which may lead to protoplasmic motion. Antagonism between Na⁺ and Ca⁺⁺ appears to be due to the fact that in solutions of NaCl the surface layer takes up an excessive amount of water and this may be prevented by the addition of suitable amounts of CaCl₂. In Nitella the outer non-aqueous surface layer may be rendered irreversibly permeable by sharply bending the cell without permanent damage to the inner non-aqueous surface layer surrounding the vacuole. The formation of contractile vacuoles may be imitated in non-living systems. An extract of the sperm of the marine worm Nereis which contains a highly surface-active substance can cause the egg to divide. It seems possible that this substance may affect the surface layer of the egg and cause it to take up water. A surface-active substance has been found in all the seminal fluids examined including those of trout, rooster, bull, and man. Duponol which is highly surface-active causes the protoplasm of Spirogyra to take up water and finally dissolve but it can be restored to the gel state by treatment with Lugol solution (KI + I). The transition from gel to sol and back again can be repeated many times in succession. The behavior of water in the surface layer of the protoplasm presents important problems which deserve careful examination.     

The Use of Alizarin Red S To Detect and Localize Calcium in Gametophyte Cells of Ferns

An aqueous solution of alizarin red S containing chloral hydrate both clears intact chlorophyllous gemma cells of Vittaria graminifolia and stains for protoplasmic calcium. Verification that the stain was protoplasmic rather than in the cell wall was shown by a positive reaction in extruded protoplasm. Similar staining was found in extruded protoplasm of Onoclea sensibilis spores. Differentiating gemma cells show localized protoplasmic accumulations of Ca²⁺ at sites where asymmetric cell divisions initiate the formation of rhizoids, antheridia or vegetative cells. The staining properties of the dye depend on careful control of pH and the addition of appropriate amounts of KC1 to the mixture. Treatment of Onoclea spores and Vittaria gemmae with 100 mM EGTA for 30 min nearly abolishes staining of their extruded protoplasts and also of intact cells of gemmae. The use of alizarin red S with and without chloral hydrate demonstrates different pools of protoplasmic Ca²⁺. When Onoclea spores are ruptured to extrude the protoplasm, both dye mixtures stain a peripheral, granular protoplasmic component. However, the chloral hydrate-containing dye also reveals Ca²⁺ associated with small particulate protoplasmic components. Extruded protoplasm of gemma cells stains intensely with alizarin-chloral hydrate, but does not stain with alizarin lacking chloral hydrate.     

NATURE OF THE ACTION CURRENT IN NITELLA : III. SOME ADDITIONAL FEATURES

Several forms of the action curve are described which might be accounted for on the ground that the outer protoplasmic surface shows no rapid electrical change. This may be due to the fact that the longitudinal flow of the outgoing current of action is in the protoplasm instead of in the cellulose wall. Hence the action curve has a short period with a single peak which does not reach zero. On this basis we can estimate the P.D. across the inner and outer protoplasmic surfaces separately. These P.D.''s can vary independently. In many cases there are successive action currents with incomplete recovery (with an increase or decrease or no change of magnitude). Some of the records resemble those obtained with nerve (including bursts of action currents and after-positivity).     

Cell Physiological Studies on the Avena Coleoptile Treated with Ribonuclease

It was revealed with excised Avena coleoptile that the growth promoting effect of indole-3-acetic acid was inhibited by pretreatment with ribonuclease (Masuda 1959a, b). This effect of ribonuclease was presumed to involve its digestive action on the ribonucleic acid at the protoplasmic surface (Masuda 1959b). Ribonuclease treatment decreases the cation binding capacity of the ribonucleic acid at the protoplasmic surface (Masuda 1959a).
On the other hand, it has been confirmed that indole-3-acetic acid bas a remarkable effect on the physico-chemical properties of protoplasmic surface such as permeability (Masuda 1955) and adhesiveness of protoplasm to the cell wall (Masuda 1957, Masuda and Takada 1957).
The purpose of the present study is to see the effect of ribonuclease on some protoplasmic properties of cells of Avena coleoptile and substantiate the authors view on the participation of ribonucleic acid in the cell elongation.     

AN ULTRASTRUCTURAL STUDY OF THE RELATIONSHIP BETWEEN MICROTUBULES AND MICROFIBRILS IN COTTON (GOSSYPIUM HIRSUTUM L.) CELL WALL REVERSALS

The arrangement of cellulosic fibrils in the cell walls of cotton fibers is very unusual; rather than exhibiting a continuous spiraling in one direction, they intermittantly reverse their gyre. Microtubules that line the periphery of the protoplasm, subjacent to the plasmalemma, tend to parallel the deployment of the cell wall microfibrils. It was not known whether this parallelism persisted through the reversal. By studying tangential sections of the cell wall/protoplasmic interfaces at the reversals, we show that congruity continues even through the reversals. Colchicine treatment did not appear to inhibit cellulose synthesis but it did abolish microtubules in the cotton fiber cells and deranged normal cell wall microfibrillar orientation. Previously, cotton fibers have been shown to possess most of the familiar organelles, but we found two new features not reported heretofore. They are microfilaments and peculiar “polygonal structures” that appear to be associated with the plasma membrane.     

THE KINETICS OF PENETRATION : XV. THE RESTRICTION OF THE CELLULOSE WALL

When Valonia cells are impaled on capillaries, it is in some ways equivalent to removing the comparatively inelastic cellulose wall. Under these conditions sap can migrate into a free space and it is found that on the average the rate of increase of volume of the sap is 15 times what it is in intact cells kept under comparable conditions. The rate of increase of volume is a little faster during the first few hours of the experiment, but it soon becomes approximately linear and remains so as long as the experiment is continued. The slightly faster rate at first may mean that the osmotic pressure of the sap is approaching that of the sea water (in the intact cell the sap osmotic pressure is always slightly above that of the sea water). This might result from a more rapid entrance of water than of electrolyte, as would be expected when the restriction of the cellulose wall was removed. During the linear part of the curve the osmotic concentration and the composition of the sap suffer no change, so that entrance of electrolyte must be 15 times as fast in the impaled cells as it is in the intact cells. The explanation which best accords with the facts is that in the intact cell the entrance of electrolyte tends to increase the osmotic pressure. As a consequence the protoplasm is partially dehydrated temporarily and it cannot take up more water until the cellulose wall grows so that it can enclose more volume. The dehydration of the protoplasm may have the effect of making the non-aqueous protoplasm less permeable to electrolytes by reducing the diffusion and partition coefficients on which the rate of entrance depends. In this way the cell is protected against great fluctuations in the osmotic concentration of the sap.     

Intercellular movement of protoplasm in vivo in developing endosperm of wheat caryopses

Summary Earlier work in our laboratory indicated that protoplasmic constituents can migrate from one cell to another in certain tissues of higher plants. Further investigations have been conducted using garlic bulbs and wheat nucellus for microscopic observation of intercellular protoplasmic movement in vivo. These gave preliminary indications of the dynamic characteristics of migrating nuclei and cytoplasm. The present paper gives recent results providing new evidence for intercellular protoplasmic movement that is neither hindered by the presence of cell wall nor the narrowness of channels of intercellular connection. By careful manipulation, intact endosperm sacs could be taken from developing caryopses (6–8 days after fertilization) without apparent injury to constituent cells. Shortly after the living specimen is mounted on the microscopic stage, asynchronous intercellular protoplasmic movement can be observed here and there. It can be seen that protoplasm extrudes in rapid but intermittent movements from one cell to the next by vigorous contraction. Although various cell constituents may move together, they can also be quite independent of each other. The moving units, though undergoing violent deformation, resume their normal shape and structure following intercellular migration. Evidently this kind of movement is a naturally occurring and active phenomenon closely related to the physiological state of the tissue. Electron microscopic studies reveal that a limited number of plasmodesmatal channels undergo modification and enlarge to 100–400 nm, through which the protoplasmic constituents pass.     

Cytomorphogenesis in cenocytic green algae. V. Segregative cell division and cortical microtubules in Dictyosphaeria cavernosa (Siphonocladales,Chlorophyceae)*

Disassembly and reassembly of cortical microtubules (MT) during and after segregative cell division (SCO) in Dictyosphaeria cavernosa (Forssk.) Børgesen were observed using fluorescence microscopy. Parallel cortical MT in a mother cell were intact just after the initiation of SCD, but soon circular, MT-free patches appeared. Protoplasmic contraction enlarged the patches, and in these areas, the protoplasm eventually became perforated. Long and undulating cortical MT were arranged densely in the reticulate protoplasm. During further protoplasmic contraction, cortical MT appeared to be random and decreased in density. Finally, short and random cortical MT were present in the segregated protoplasts. Parallel cortical MT reassembled in the expanding daughter cells. After the daughter cells came in contact with one another, a radial system of cortical MT was constructed at the side that faced the inside of the mother cell wall. A microtubule inhibitor (amiprophos methyl, APM) had no effect on SCO. Segregative cell division was not induced directly by mechanical wounding. A comparison between SCO and wound-induced protoplasmic contraction was made.     

Electron Microscope Observations on Spore Formation in the True Slime Mold Didymium nigripes

SYNOPSIS. Developing and mature sporangia of the true slime mold Didymium nigripes were studied with the electron microscope to follow the course of spore formation. The sporangium forms from the plasmodium as a protoplasmic bleb which differentiates into a stalk and an apical sphere containing a mass of protoplasm. Nuclei within this protoplasmic mass undergo synchronous division (presumably meiosis). The division spindle forms within the nuclear membrane which is retained intact throughout the division; centrioles have not been observed at the spindle poles. At the same time the nuclei are dividing, the protoplasm cleaves to give ultimately uninucleate spheres—the incipient spores. Capillitial threads come to lie in the furrows created by the cleaving protoplasm. A wall consisting of an inner thick component and an outer thin component forms about each sphere. Cyto-chemical tests suggest that the inner wall of the spore is cellulose-containing and that the outer component might contain chitin.     

MECHANICAL RESTORATION OF IRRITABILITY AND OF THE POTASSIUM EFFECT

Treatment of Nitella with distilled water apparently removes from the cell something which is responsible for the normal irritability and the potassium effect, (i.e. the large P.D. between a spot in contact with 0.01 M KCl and one in contact with 0.01 M NaCl). Presumably this substance (called R) is partially removed from the protoplasm by the distilled water. When this has happened a pinch which forces sap out into the protoplasm can restore its normal behavior. The treatment with distilled water which removes the potassium effect from the outer protoplasmic surface does not seem to affect the inner protoplasmic surface in the same way since the latter retains the outwardly directed potential which is apparently due to the potassium in the sap. But the inner surface appears to be affected in such fashion as to prevent the increase in its permeability which is necessary for the production of an action current. The pinch restores its normal behavior, presumably by forcing R from the sap into the protoplasm.     

SOME PROPERTIES OF PROTOPLASMIC GELS : I. TENSION IN THE CHLOROPLAST OF SPIROGYRA

The chloroplast of Spirogyra is a long, spirally coiled ribbon which may contract to form a short, nearly straight rod. This happens under natural conditions and it can also be produced by a variety of inorganic salts and by some organic substances. It also occurs when the chloroplast is freed by centrifugal force from the clear peripheral protoplasm which is in contact with the cellulose wall. It would therefore seem that the chloroplast may be passively stretched by the action of the clear protoplasm and hence it contracts as soon as it is set free. This contraction happens in dead as well as in living cells. It would be of much interest to know how the protoplasm brings about the coiling of the chloroplast and how the chloroplast is set free by various reagents. Presumably they must penetrate the living protoplasm to produce the effects described. In one species partial contraction without detachment from the peripheral protoplasm can be brought about by lead acetate. This is reversible. Lead nitrate does not produce this result. The attack upon the problem is greatly facilitated by the study of dead cells. Thereby we reduce the number of variables but the chloroplast continues to react to certain chemical and physical agents in much the same manner as in the living cell and the solution surrounding it can be controlled as is not possible in the living cell. We must await further investigation to learn what plant and animal cells contain gels under tension and what functions they perform.     

Ion transport in Nitellopsis obtusa

The distribution and rates of exchange of the ions sodium, potassium, and chloride in single internodal cells of the ecorticate characean, Nitellopsis obtusa, have been studied. In tracer experiments three kinetic compartments were found, the outermost "free space" of the cell, a compartment we have called "protoplasmic non-free space", and the cell sap. The concentrations in the vacuole were 54 mM Na(+), 113 mM K(+), and 206 mM Cl(-). The steady state fluxes across the vacuolar membrane were 0.4 pmole Na(+)/cm.(2) sec., 0.25 pmole K(+)/cm.(2) sec., and 0.5 pmole Cl(-)/cm.(2) sec. The protoplasmic Na/K ratio is equal to that in the vacuole but protoplasmic chloride is relatively much lower. Osmotic considerations suggest a layer 4 to 6 micro thick with sodium and potassium concentrations close to those in the vacuole. The fluxes between protoplasm and external solution were of the order of 8 pmoles Na(+)/cm.(2) sec. and 4 pmoles K(+)/cm.(2) sec. We suggest that the protoplasm is separated from the cell wall by an outer protoplasmic membrane at which an outward sodium transport maintains the high K/Na ratio of the cell interior, and from the vacuole by the tonoplast at which an inward chloride transport maintains the high vacuolar chloride. The tonoplast appears to be the site of the principal diffusion resistance of the cell, but the outer protoplasmic membrane probably of the main part of the potential.     

Organized Protoplasmic Units of the Plant Cell: I. THEIR OCCURRENCE, ORIGIN, AND STRUCTURE

The development of the placental tissue of the tomato fruithas been investigated and it has been found that the protoplasmof cells of this tissue separates into organized units of protoplasmas the fruit matures and ripens. Similar organized protoplasmicunits have been detected in other fruits at a comparable stageof development. A study has been made of the structure and stability of theseorganized units, and it has been deduced that these organizedunits of protoplasm are in fact compartments of the protoplasmof the cell which have become separated from one another. Eachorganized protoplasmic unit is able to exist independently ofthe cell for several days, and is apparently surrounded by amembrane of the endoplasmic reticulum. The possible significance of these new findings is discussedin relation to the structural and biochemical organization ofthe plant cell.     

DISSIMILARITY OF INNER AND OUTER PROTOPLASMIC SURFACES IN VALONIA. II

In measurements of P.D. across the protoplasm in single cells, the presence of parallel circuits along the cell wall may cause serious difficulty. This is particularly the case with marine algae, such as Valonia, where the cell wall is imbibed with a highly conducting solution (sea water), and hence has low electrical resistance. In potential measurements on such material, it is undesirable to use methods in which the surface of the cell is brought in contact with more than one solution at a time. The effect of a second solution wetting a part of the cell surface is discussed, and demonstrated by experiment. From further measurements with improved technique, we find that the value previously reported for the P.D. of the chain Valonia sap | Valonia protoplasm | Valonia sap is too low, and also that the P.D. undergoes characteristic changes during experiments lasting several hours. The maximum P.D. observed is usually between 25 and 35 mv., but occasionally higher values (up to 82 mv.) are found. The appearance of the cells several days after the experiment, and the P.D.''s which they give with sea water, indicate that no permanent injury has been received as a result of exposure to artificial sap. If such cells are used in a second measurement with artificial sap, however, the form of the P.D.-time curve indicates that the cells have undergone an alteration which persists for a long time. On the basis of the theory of protoplasmic layers, an attempt has been made to explain the observed changes in P.D. with time, assuming that these changes are due to penetration of KCl into the main body of the protoplasm.     

THE COMPOSITION OF THE CELL SAP OF THE PLANT IN RELATION TO THE ABSORPTION OF IONS

1. Chemical examination of the cell sap of Nitella showed that the concentrations of all the principal inorganic elements, K, SO₄, Ca, Mg, PO₄, Cl, and Na, were very much higher than in the water in which the plants were growing. 2. Conductivity measurements and other considerations lead to the conclusion that all or nearly all of the inorganic elements present in the cell sap exist in ionic state. 3. The insoluble or combined elements found in the cell wall or protoplasm included Ca, Mg, S, Si, Fe, and Al. No potassium was present in insoluble form. Calcium was predominant. 4. The hydrogen ion concentration of healthy cells was found to be approximately constant, at pH 5.2. This value was not changed even when the outside solution varied from pH 5.0 to 9.0. 5. The penetration of NO₃ ion into the cell sap from dilute solutions was definitely influenced by the hydrogen ion concentration of the solution. Penetration was much more rapid from a slightly acid solution than from an alkaline one. It is possible that the NO₃ forms a combination with some constituent of the cell wall or of the protoplasm. 6. The exosmosis of chlorine from Nitella cells was found to be a delicate test for injury or altered permeability. 7. Dilute solutions of ammonium salts caused the reaction of the cell sap to increase its pH value. This change was accompanied by injury and exosmosis of chlorine. 8. Apparently the penetration of ions into the cell may take place from a solution of low concentration into a solution of higher concentration. 9. Various comparisons with higher plants are drawn, with reference to buffer systems, solubility of potassium, removal of nitrate from solution, etc.     

CHANGES OF APPARENT IONIC MOBILITIES IN PROTOPLASM : II. THE ACTION OF GUAIACOL AS AFFECTED BY pH

The normal P.D. across the protoplasm of Valonia macrophysa is about 10 mv. negative (inwardly directed). On adding 0.01 M guaiacol to the sea water the P.D. becomes positive and then slowly returns approximately to the normal value. In many cases this behavior is not much affected by raising the pH and so increasing the concentration of the guaiacol ion but in other cases such an increase makes the P.D. somewhat more negative. But if we wait until the exposure to guaiacol has lasted 5 minutes (and the P.D. has returned to its normal value) before we raise the pH, the result is very different. The cell then behaves as though it had been sensitized to the action of the guaiacol ion which appears to be far more effective than undissociated guaiacol in making the P.D. more positive. This may be due in part to the high apparent mobility of the guaiacol ion and in part to alterations which it produces in the protoplasm (such alterations increase the P.D. across the protoplasm whereas ordinary injury would be expected to lower it and the cells live on after this treatment and show no signs of injury). This action of the guaiacol ion is in marked contrast to the behavior of other anions whose effect resembles that of Cl^-.     

The mechanism of accumulation in living cells

When a compound enters a living cell until its activity becomes greater inside than outside, it may be said to accumulate. Since it moves from a region where its activity is relatively low to a region where its activity is relatively high, it is evident that work must be done to bring this about. The following explanation is suggested to account for accumulation. The protoplasmic surface is covered with a non-aqueous layer which is permeable to molecules but almost impermeable to ions. Hence free ions cannot enter except in very small numbers. The experiments indicate that ions combine at the outer surface with organic molecules (carrier molecules) and are thus able to enter freely. If upon reaching the aqueous protoplasm these molecules are decomposed or altered so as to set the ions free, the ions must be trapped since they cannot pass out except in very small numbers. If we adopt this point of view we can suggest answers to some important questions. Among these are the following: 1. Why accumulation is confined to electrolytes. This is evident since only ions will be trapped. 2. Why ions appear to penetrate against a gradient. Actually there is no such penetration since the ions enter in combination with molecules. The energy needed to raise the activity of entering compounds is furnished by the reactions involved in the process of accumulation. 3. Why, in absence of injury, ions do not come out when the cell is placed in distilled water. Presumably the outgoing ions will combine at the outer surface with carrier molecules and then move inward in the same way as ions coming from without. 4. Why the relative rate of penetration falls off as the external concentration increases. This is because the entrance of ions is limited by the number of carrier molecules but no such limitation exists when ions move outward since they can do so without combining with carrier molecules. 5. Why accumulation is promoted by constructive metabolism which is needed to build up the organic molecules and by destructive metabolism which brings about their decomposition. 6. Why measuring the mobilities of ions in the outer protoplasmic surface does not enable us to predict the relative rate of entrance of ions. We find for example in Nitella that K⁺ has a much higher mobility than Na⁺ but the accumulation of these ions does not differ greatly. This is to be expected if they enter by combining with molecules at the surface. Only if K⁺ is able to combine preferentially will it accumulate preferentially. 7. Why ions may come out in anoxia and at low temperatures. If these conditions depress the formation of carrier molecules and their decomposition in the protoplasm, the balance between intake and outgo of ions will be disturbed and relatively more may come out. 8. Why the excess of internal over external osmotic pressure is less in sea water than in fresh water. As the external concentration of ions increases the rate of intake does not increase in direct proportion since the number of carrier molecules does not increase and this slows down the relative rate of intake of ions. But it does not slow down the rate of exit of ions since they need not combine with carrier molecules in order to pass out. Hence the excess of ions inside will be relatively less as the concentration of external ions increases. 9. How water is pumped from solutions of higher to solutions of lower osmotic pressure. If metabolism and consequently accumulation is higher at one end of a cell than at the other, the internal osmotic pressure will be higher at the more active end and this makes it possible for the cell to pump water from solutions of higher osmotic pressure at the more active end to solutions of lower osmotic pressure at the less active, as shown experimentally for Nitella. This might help to explain the action of kidney cells and the production of root pressure in plants.     

A MICRO INJECTION STUDY ON THE PERMEABILITY OF THE STARFISH EGG

The experiments with the NH₄Cl are similar to, and corroborate micro injection experiments performed in connection with some work on mustard gas in which the writer collaborated. Eggs immersed in sea water containing decomposed mustard gas, at a certain low concentration are not affected. If, however, the solution be injected, the egg quickly cytolyzes owing to the free HCl present. A similar impermeability of the protoplasmic surface film to certain substances was also encountered in injection work on Amœba. Amœbœ immersed in an aqueous solution of eosin will not take the stain till after death. On the other hand, the eosin, when injected into the Amœba, quickly permeates the protoplasm, to be arrested only at the surface. The semipermeability of a living cell appears primarily to be a function of its surface film. It is immaterial whether this film be that of the original cortex of the cell, a film newly formed over a cut surface, or a film that surrounds an artificially induced vacuole within the cell. As long as such a surface film exists neither the acid group of the NH₄Cl nor the alkaline group of the NaHCO₃ can, within certain concentration limits, penetrate the protoplasm. These solutions, if injected beneath the surface film, however, will produce their characteristic effects upon the protoplasm.     

CYTOMORPHOLOGICAL ASPECTS OF WOUND HEALING IN SELECTED SIPHONOCLADALES (CHLOROPHYCEAE)1

Wound healing in most of the Siphonocladales investigated differs markedly from healing processes known in other coenocytic green algae. No wound plug is evident during healing in Ernodesmis verticillata (Kützing) Børgesen, Boergesenia forbesii (Harvey) Feldmann, Cladophoropsis membranacea (C. Agardh) Børgesen, Siphonocladus tropicus (Crouan) J. Agardh, Struvea elegans Børgesen, Struvea sp. and Valonia spp. The wound response in these genera involves substantial protoplasmic motility which includes retraction of the cell contents from the wound site. The protoplasm then either closes around the central vacuole (e.g. Ernodesmis) or breaks up into numerous spherical protoplasts (e.g. Valonia ventricosa J. Agardh). An intermediate pattern of healing is present in Cladophoropsis where the protoplasm usually separates into a small number of portions that individually heal in a manner similar to Ernodesmis. In all cases, the protoplasts resulting from wounding are viable. Struvea anastomosans (Harvey) Piccone, Chamaedoris sp. and Boodlea sp. are unusual in producing a distinct wound plug during healing, thereby resembling the mode of healing reported in various codialean and dasycladalean genera. It is hypothesized, that wound-induced protoplasmic motility in the Siphonocladales involves the same cellular mechanisms that segregate protoplasm during cell, division.