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On a peculiarity of planarian digestion
Camp. Biochem. Physiol.,1974, Vol. 48A, pp. 601 to 607. Pergamon Press. Printed in Great Britain
ON A PECULIARITY I. M. SHEIMANN
OF PLANARIAN
DIGESTION
and N. JU. SAKHAROVA
Institute of Biophysics, Academy of Sciences of the U.S.S.R., (Received 27 February
Push&noon,
U.S.S.R.
1973)
Abstract-l. Structural changes in the planarian body after feeding were studied. 2. In the initial hours of digestion the cells of intestinal epithelium swelled and gradually filled almost the whole body. 3. The first food particles approached the nervous structures 3 hr after feeding. Their contact continued up to the fourth day. The greater part of the intestinal cells were then destroyed. After the fifth day the structure of the intestine was gradually restored. 4. The contact between the nervous system and the partially digested food particles may be the basis of the specific “memory transfer” phenomenon through planarian cannibalism.
INTRODUCTION IN 1962 AN interesting phenomenon was described: the learning of flatwormsplanarians-was facilitated after feeding with conditioned planarians (McConnell, 1962). Similar results were obtained by other authors (Westerman, 1963 ; John, 1964; McConnell & Shelby, 1970). Further investigations in this direction were performed on rodents and were modified significantly. At present, a large number of papers dealing with “memory transfer” by the use of injections of brain extracts of conditioned animals have been published. The “memory transfer” analysis is facilitated when working on animals with a primitive nervous system. In 1967, we carried out a set of experiments on Dug&u rig&z-cannibal planarians. The sensitization to conditioned stimuli, applied to the planarian donors (that were eaten), has appeared in the planarian recipients (that were fed). This “transfer” phenomenon was of a short duration and it cannot therefore be considered as a “memory transfer” per se. At the same time, short-term sensitization has a specific character and corresponds to predetermined stimuli. In a special set of experiments the different groups of planarian donors were stimulated with a strong light, vibration or electrical shock. After eating such planarians the worm recipients show a corresponding increase of sensitivity to light, to vibration or to both light and vibration (Sheimann, 1970). Thus when studying the “memory transfer” phenomenon we have seen the transmission of the factor responsible for the specificity of excitation under both conditioned and unconditioned stimuli from donors. According to our results this factor acts in the recipients for a limited period of time.
601
602
I. M.
SHEIMANN AND N. Ju. SAKHAROVA
In analysing the “memory transfer” phenomenon on planarians, we studied first of all the structural changes in the body of planarians in the course of digestion and nutrient propagation. MATERIALS
AND
METHODS
After a week of starvation, planarians, Dugesiu tigrina, were fed with tubifex (Chironomus sp.) and were then fixed in Zenker liquid for 1-6 hr and 1-7 and 9 days. Sections of 7 pm were stained with azur-eosin at pH 4.6 (Lilli, 1954). Starving animals served as controls. RESULTS
The main volume of the starving planarian body is engaged largely with parenchyma which fills the space between the gut and the other organs. It consists of large vacuolized eosinophilic cells. Among them there are eosinophilic granular formations of different shapes and sizes and cells of an embryonic type-neoblasts stained with azur (7-8 pm) and large polymorphic cells (Xl-30 pm) (Sakharova, 1972). The gut walls of planaria D. tigrina are formed by single-layer epithelium. In starving worms it is represented by low and smooth cells which contain eosinophilic cytoplasm and lipid vacuoles. After alcohol treatment these vacuoles lose their contents and appear to be empty (Willier et al., 1925). The nuclei of the gut cells are arranged near the outlined basal membrane which separates the gut from the parenchyma. Among the columnar cells, spherical or goblet cells, filled with eosinophilic granules, can be seen (Fig. 1). After feeding, the structure of the gut changes. One hr after feeding, the intestinal lumen is significantly extended and filled with food. Some gastrodermal cells are enlarged and have finger-like projections directed into the lumen which phagocytose the food particles. After 2 hr, similar projections appear in all the gastrodermal cells. In their cytoplasm the food vacuoles stained with eosin can be seen (Fig. 2a). After 3 hr, further enlargement of phagocytosing cells leads to a thickening of the intestinal wall and to a decrease in size of the lumen. The surface of the lumen is not smooth. In the cells of the anterior parts of the gut, eosinophilic vacuoles and granules predominate, but in the posterior parts, azurophilic ones are predominant. The histochemical investigation of these vacuoles shows that their contents consist of protein at various stages of splitting (Willier et al., 1925). The lipid vacuoles are met only in the basal parts of cells. After 4-5 hr, in all sections of the intestine, the phagocytosing cells are large and contain many vacuoles and granules. The distinction in their colour, in different parts of the body, is preserved. After 6 hr all the gastrodermal cells are elongated and filled with vacuoles and granules stained differently. The lipid vacuoles are disposed in the bases of the cells. In the head part, the surface of the gut is smoother than that in the tail part. The spherical cells of the gut disappear (Fig. 2b). One day after feeding, the intestinal lumen decreases compared with preliminary periods, and the gastrodermal cells enlarge further. They are filled with
FIG. 1, Structure of the body of hungry planaria D. Ggrka. gc, The gut cells; Iv, the lipid vacuoles; n, the nuclei; L, the gut lumen; P, the parenchyma; N, the neuropil region; nc, the ganglion cells; m, the basal membrane.
ON
A PECULIARITY
OF PLANARIAN
DIGESTION
603
granules and vacuoles which also differ in colour. Often, the cell boundaries are difficult to distinguish, or are completely absent. In some sites, the basal membrane is broken. In the head part of the gut there are also broken apical membranes of the cells. Lipid vacuoles are not seen. It is possible that they are used up as energy sources for phagocytosis (Willier et uZ., 1925). After 2 days, the intestinal lumen disappears. The cell boundaries and the basal membrane are visible only at some sites. Along a large space, the gut wall is represented as a symplast. In its apical parts the vacuoles and granules are stained with eosin but, approaching the bases, the inclusions stained with azur predominate. Sometimes, near the apical boundary, one can observe empty vacuoles. After 3-4 days the intestine is represented by a large and spread-out symplast in which there is a small lumen (Fig. 2~). After 5 days, in the gut, the symplast and the individual cells exist simultaneously. In the head section there are small cells with smooth apical borders. At their bases lie the nuclei, surrounded with basophilic cytoplasm. There are large eosinophilic vacuoles at the apical ends of the cells. In the middle part of the body, the intestinal cells are higher and their apical ends are not smooth. The borders between cells are not distinguishable everywhere. Sometimes, on the apical surface, there are ruptures and eosinophilic vacuoles with picnotic contents. In the tail part, the intestinal cells have large vacuoles with picnotic contents, broken cell membranes and damaged sites of basal membrane. On the bases of the gut wall lie many nuclei surrounded with basophilic cytoplasm (Fig. 2d). After 7 days the intestine shows almost no damage. In the head part, the gut wall contains large cells with many vacuoles. In the tail part the gut cells are small and their vacuoles are empty, or filled with an eosinophilic substance. The nuclei are on the basal layer of the cytoplasm stained with azur. After 9 days, the gut structure is similar to that seen initially. The intestinal lumen is visible and becomes empty. It is surrounded with columnar and spherical cells. At the apical ends of the columnar cells there are empty vacuoles and in the spherical cells, dense eosinophilic granules. In the bases of cells lie the nuclei surrounded with basophilic cytoplasm. Simultaneously, with the changes in the gut, reorganization of the parenchyma occurs. As a result, new space relations arise between the gut and nervous structures. One hour after eating, there is no change in the structure of the parenchyma. After 2 hr, the parenchimal cells become flat and the volume of parenchyma as a whole decreases, especially in the tail section. After 3 hr, the space occupied by the parenchyma continues to decrease. The walls of the gut, consisting of the large cells filled with food vacuoles and granules, lie close to the lower parts of the nervous ganglion and near the nerve cords. From the intestinal cells the projections, with granular eosinophilic contents, are extended to the parenchyma. At this time similar formations are visible between the nerve cells of the ganglion and near the nerve cords (Fig. 3a).
604
I. M.
SHEIMANN
AND
N. Ju. SAKHAROVA
After 4-5 hr, a further decrease of the parenchymal stratum occurs. The eosinophilic granular formations are distributed at the site of the ganglion and the nerve cords. After 6 hr, the parenchymal tissue remains chiefly under the external epithelium but, in the intergut folds, it is represented by narrow strips of contracted cells. Near the nervous structures large accumulations of eosinophilic granular formations are visible. After 1 day the parenchymal stratum becomes insignificant. In the region of the ganglion lie the stretched cells with granular inclusions. In the gut wall there are cells with projections stretched in the direction of the ganglion and filled with eosinophilic granules. The gut walls are joined to the nerve cords (Fig. 3b). After 2 days there are thin strips of parenchyma between the intestinal loops. The expanded intestinal walls are intimately joined to the ganglion, and almost completely surround the nerve cords. After 3 days the parenchymal stratum remains under the external epithelium and between intestinal loops in the form of thin strips of contracted cells. In the head part there are numerous eosinophilic granular elongated formations. After 4-5 days, the volume of parenchymal tissue increases. The ganglion and intestine are separated from one another. The nerve cords are also separated from the intestine. In the parenchymal stratum which divides the intestine from the nervous system there are granular formations and cells with eosinophilic granules. After 7-9 days a further increase of the parenchyma size occurs. It is filled with different granular formations. At the same time the distribution and quantity of neoblasts in the parenchyma also changes. During the first hours after feeding most of them migrate under the external epithelium where they remain up to the fourth or fifth day. Later on, they become redistributed throughout the body. DISCUSSION
Our data show that in the course of digestion a significant histological rebuilding occurs in the body of the planaria, D. tigrina. The captured food enters the sections of the intestine irregularly but it is usually distributed primarily through the anterior section of the gut. Probably, an asynchronous manner of histological changes in various parts of the intestine is related to this fact. The initial changes are represented by the formation of the projections of gut cells towards the lumen and the food is captured by them. Such projections have been described as early as 1925 (Willier et al., 1925). Further reorganizations of the body are connected with digestion. Almost all the processes of splitting of the food particles occur in the cells of the gut (Willier et al., 1925 ; Jennings, 1957, 1962). The particles of food are split under the action of various enzymes while the food is passing from the apical end to the bases of the cells. The medium of the cells changes from acidic to slightly alkaline. The gradual changes of chromatlinity of food particles from eosinophilia to basophilia and back seem to be due to the changes described above. A similar change of chromaflinity was also observed in other species of planarians (Willier et al., 1925; Horne & Darlington, 1967). During the first 3 days
ON
A PECULIARITY
OF PLANARIAN
DIGESTION
605
after feeding, the gut walls became thicker, When the phagocytosis of the food was completed, the apical surface of the cells gradually became smooth. As the gastrodermal cells enlarge, the intestinal lumen decreases and, on the third day, disappears. Simultaneously the basal membrane which separates the gut from the parenchyma is destroyed. It is difficult to distinguish the cell boundaries. The intestinal walls become symplastic. Five days after feeding, individual cells with different features of destruction are visible again. Most probably, the organism gets rid of wasted cells and in their place new cells, differentiated from the neoblasts, appear. Up to the ninth day restoration of the initial structure of the intestine occurs. When digestion is finished, the food resources are kept in the cells of the gut (Willier et al., 1925; Jennings, 1957). According to our data the whole cycle of digestion in D. tigrina lasts for 7-9 days. The first period-the phagocytosis-lasts for 6 hr and the digestion for 3-4 days. The destruction and the following restitution of the intestinal structure occur from the fifth to the ninth day. The intestine of planarians has two functions: digestion and distribution of nutrients. Organs are supplied with nutrients by means of heavy ramifications of the intestine and by the instability of its cellular structure (Beclemishev, 1964). According to our data this function is also conditioned by the state of the parenchyma. Between the third and the sixth hours a compression of the fixed cells of the parenchyma occurs and the neoblasts are shifted towards the external epithelium. This results in a sharp decrease in the volume of the parenchyma and in an approach of the intestine to the other organs. Some authors consider that the parenchymal cells take part in the transfer of nutrients directly (Jennings, 1957; Rosenbaum & Rolon, 1960; Berg & Berg, 1968). According to other data (Willier et al., 1925) only the gastrodermal cells fulfil this distributive function. The method used by us for staining allows us to distinguish between individual tissues and to observe their changes. The observations show the food particles to be always arranged in the gastrodermal cells independentIy of their state. Three hr after feeding, the projections with eosinophilic granules directed to the parenchyma arise. Perhaps the nutrient is distributed firstly through them. After 34 hr similar granular formations are visible near the ganglion and the nerve cords. Further distribution of nutrients is provided with the deformation of the intestine. On the second to fourth days after feeding, the enlarged intestinal walls come close to the ganglion and surround the nerve cords. In the course of the enlargement and the deformation of the gastrodermal cells and also of the destruction of their membrane, a direct contact is set up between the nervous system and the intestinal contents. According to these data one can say that the transfer of nutrients from the gut to the nervous system begins during the first hours after capture of the food. In the beginning these substances move along the projections and from the second to the fourth day the ganglion and the nerve cords seem to be immersed in the mass of the intestine.
606
1. M. SHEIMANNANDN. Ju. SAKHAROVA
Five days after feeding the parenchyma is restored and the direct contact between the nervous system and the intestine is interrupted. The number of granular formations in the parenchyma increases. They seem to be converted phagocytes (Berg 2% Berg, 1968) or noncellular formations of a different shape which are connected neither with the intestine nor with the nervous system. Probably nutrients are maintained mainly in them. Comparison of the morphological changes during digestion and of the physiological “memory transfer” phenomenon shows a correlation between them. The physiological phenomenon, expressed as an increase of specific excitability for several hours after food intake (5-6 hr), lasts for 3 days, then decreases and disappears on the fifth day. Analysing these data one may suppose that during cannibalism the body of the planaria recipient obtains a substance from the planaria donor and this substance acts specifically on the nervous system of the former. The “transfer” phenomenon is manifested just while there is direct contact between the nervous structures and the intestinal contents. At this time the food particles from the gastrodermal cells are significantly basophilic. Cessation of the “transfer” phenomenon coincides with the destruction of the intestinal cells. Perhaps the enzymes acting at the beginning of digestion only partially break up the food, and the specific properties of the food particles are maintained. Later, during cell destruction, a final disintegration of nutrients occurs and its specific action ceases. It is transformed into a passive nourishing resource.
REFERENCES BECLEMISHEVV. V. (1964) Bases of the Comparative Anatomy of the Invertebrates, Vol. 2. Nauka, Moscow. BERG G. G. & BERG 0. A. (1968) Presence of trimetaphosphatase in the Turbellarian Dugesia tigrina and the relation of enzyme distribution to food uptake. Trans. Am. microsc. Sot. 87, 335-341. HORNE M. K. & DARLINGTONJ. T. (1967) Uptake and intracellular digestion of ferritin in the Planarian Phagocata gracilis. Trans. Am. microsc. Sot. 86, 268-273. JENNINGSJ. B. (1957) Studies on feeding, digestion and food storage in free-living flatworms. Biol. Bull. mar. biol. Lab., Woods Hole 112, 63-80. JENNINGSJ. B. (1962) Further studies on feeding and digestion of triclad Turbellaria. Biol. Bull., mar. biol. Lab., Woods Hole 123, 571-581. JOHN E. R. (1964) Studies on learning and retention in Planaria. In Brain Function (Edited by BRAZIERM. A.), Vol. 2, pp. 161-182. University of California Press. LILLIE R. D. (1954) Histopathologic Technic and Practical Histochemistry, 2nd Edition, pp. 118-119. McGraw-Hill, New York. MCCONNELL J. V. (1962) Memory transfer through cannibalism in planarians. J. Neuropsych&. 3 (Suppl. I), 4248. MCCONNELLJ. V. & SHELBY J. M. (1970) Memory transfer experiments in invertebrates. In Molecular Mechanisms in Memory and Learning (Edited by UNGARG.), pp. 71-101. Plenum Press, New York. ROSENBAUMR. M. & ROLON C. J. (1960) Intracellular digestion and hydrolytic enzymes in the phagocytes of planarians. Biol. Bull., mar. biol. Lab., Woods Hole 118, 315-323. SAKHAROVA N. Ju. (1972) On sources of regeneration in planarians. Ontogenez 3, 95-100.
ON A PECULIARITY OF PLANARIAN DIGESION
607
SHEIMANNI. M. (1970) On the complicated structure of the memory trace (in a primitive nervous system). In Memory and Trace Processes (Edited by LIVANOVM. N.), pp. 210-214. Materials of the 2nd conference on the problem of memory and trace processes, Pushchino. WE~TEEMANR. A. (1963) Somatic inheritance of habituation of responses to light in planarians. Science, Wash. 140, 676-677. WILLIER B. H., HYMAN 2. H. & RIFENBURCHS. A. (1925) A histochemical study of intracellular digestion in triclad flatworms. J. Morph. Physiol. 40, 299-340. Key Word Index-Planarians; “memory symplast; projections of the intestinal cells.
transfer”;
digestion;
intestine
structure;
ON A PECULIARITY I. M. SHEIMANN
OF PLANARIAN
DIGESTION
and N. JU. SAKHAROVA
Institute of Biophysics, Academy of Sciences of the U.S.S.R., (Received 27 February
Push&noon,
U.S.S.R.
1973)
Abstract-l. Structural changes in the planarian body after feeding were studied. 2. In the initial hours of digestion the cells of intestinal epithelium swelled and gradually filled almost the whole body. 3. The first food particles approached the nervous structures 3 hr after feeding. Their contact continued up to the fourth day. The greater part of the intestinal cells were then destroyed. After the fifth day the structure of the intestine was gradually restored. 4. The contact between the nervous system and the partially digested food particles may be the basis of the specific “memory transfer” phenomenon through planarian cannibalism.
INTRODUCTION IN 1962 AN interesting phenomenon was described: the learning of flatwormsplanarians-was facilitated after feeding with conditioned planarians (McConnell, 1962). Similar results were obtained by other authors (Westerman, 1963 ; John, 1964; McConnell & Shelby, 1970). Further investigations in this direction were performed on rodents and were modified significantly. At present, a large number of papers dealing with “memory transfer” by the use of injections of brain extracts of conditioned animals have been published. The “memory transfer” analysis is facilitated when working on animals with a primitive nervous system. In 1967, we carried out a set of experiments on Dug&u rig&z-cannibal planarians. The sensitization to conditioned stimuli, applied to the planarian donors (that were eaten), has appeared in the planarian recipients (that were fed). This “transfer” phenomenon was of a short duration and it cannot therefore be considered as a “memory transfer” per se. At the same time, short-term sensitization has a specific character and corresponds to predetermined stimuli. In a special set of experiments the different groups of planarian donors were stimulated with a strong light, vibration or electrical shock. After eating such planarians the worm recipients show a corresponding increase of sensitivity to light, to vibration or to both light and vibration (Sheimann, 1970). Thus when studying the “memory transfer” phenomenon we have seen the transmission of the factor responsible for the specificity of excitation under both conditioned and unconditioned stimuli from donors. According to our results this factor acts in the recipients for a limited period of time.
601
602
I. M.
SHEIMANN AND N. Ju. SAKHAROVA
In analysing the “memory transfer” phenomenon on planarians, we studied first of all the structural changes in the body of planarians in the course of digestion and nutrient propagation. MATERIALS
AND
METHODS
After a week of starvation, planarians, Dugesiu tigrina, were fed with tubifex (Chironomus sp.) and were then fixed in Zenker liquid for 1-6 hr and 1-7 and 9 days. Sections of 7 pm were stained with azur-eosin at pH 4.6 (Lilli, 1954). Starving animals served as controls. RESULTS
The main volume of the starving planarian body is engaged largely with parenchyma which fills the space between the gut and the other organs. It consists of large vacuolized eosinophilic cells. Among them there are eosinophilic granular formations of different shapes and sizes and cells of an embryonic type-neoblasts stained with azur (7-8 pm) and large polymorphic cells (Xl-30 pm) (Sakharova, 1972). The gut walls of planaria D. tigrina are formed by single-layer epithelium. In starving worms it is represented by low and smooth cells which contain eosinophilic cytoplasm and lipid vacuoles. After alcohol treatment these vacuoles lose their contents and appear to be empty (Willier et al., 1925). The nuclei of the gut cells are arranged near the outlined basal membrane which separates the gut from the parenchyma. Among the columnar cells, spherical or goblet cells, filled with eosinophilic granules, can be seen (Fig. 1). After feeding, the structure of the gut changes. One hr after feeding, the intestinal lumen is significantly extended and filled with food. Some gastrodermal cells are enlarged and have finger-like projections directed into the lumen which phagocytose the food particles. After 2 hr, similar projections appear in all the gastrodermal cells. In their cytoplasm the food vacuoles stained with eosin can be seen (Fig. 2a). After 3 hr, further enlargement of phagocytosing cells leads to a thickening of the intestinal wall and to a decrease in size of the lumen. The surface of the lumen is not smooth. In the cells of the anterior parts of the gut, eosinophilic vacuoles and granules predominate, but in the posterior parts, azurophilic ones are predominant. The histochemical investigation of these vacuoles shows that their contents consist of protein at various stages of splitting (Willier et al., 1925). The lipid vacuoles are met only in the basal parts of cells. After 4-5 hr, in all sections of the intestine, the phagocytosing cells are large and contain many vacuoles and granules. The distinction in their colour, in different parts of the body, is preserved. After 6 hr all the gastrodermal cells are elongated and filled with vacuoles and granules stained differently. The lipid vacuoles are disposed in the bases of the cells. In the head part, the surface of the gut is smoother than that in the tail part. The spherical cells of the gut disappear (Fig. 2b). One day after feeding, the intestinal lumen decreases compared with preliminary periods, and the gastrodermal cells enlarge further. They are filled with
FIG. 1, Structure of the body of hungry planaria D. Ggrka. gc, The gut cells; Iv, the lipid vacuoles; n, the nuclei; L, the gut lumen; P, the parenchyma; N, the neuropil region; nc, the ganglion cells; m, the basal membrane.
ON
A PECULIARITY
OF PLANARIAN
DIGESTION
603
granules and vacuoles which also differ in colour. Often, the cell boundaries are difficult to distinguish, or are completely absent. In some sites, the basal membrane is broken. In the head part of the gut there are also broken apical membranes of the cells. Lipid vacuoles are not seen. It is possible that they are used up as energy sources for phagocytosis (Willier et uZ., 1925). After 2 days, the intestinal lumen disappears. The cell boundaries and the basal membrane are visible only at some sites. Along a large space, the gut wall is represented as a symplast. In its apical parts the vacuoles and granules are stained with eosin but, approaching the bases, the inclusions stained with azur predominate. Sometimes, near the apical boundary, one can observe empty vacuoles. After 3-4 days the intestine is represented by a large and spread-out symplast in which there is a small lumen (Fig. 2~). After 5 days, in the gut, the symplast and the individual cells exist simultaneously. In the head section there are small cells with smooth apical borders. At their bases lie the nuclei, surrounded with basophilic cytoplasm. There are large eosinophilic vacuoles at the apical ends of the cells. In the middle part of the body, the intestinal cells are higher and their apical ends are not smooth. The borders between cells are not distinguishable everywhere. Sometimes, on the apical surface, there are ruptures and eosinophilic vacuoles with picnotic contents. In the tail part, the intestinal cells have large vacuoles with picnotic contents, broken cell membranes and damaged sites of basal membrane. On the bases of the gut wall lie many nuclei surrounded with basophilic cytoplasm (Fig. 2d). After 7 days the intestine shows almost no damage. In the head part, the gut wall contains large cells with many vacuoles. In the tail part the gut cells are small and their vacuoles are empty, or filled with an eosinophilic substance. The nuclei are on the basal layer of the cytoplasm stained with azur. After 9 days, the gut structure is similar to that seen initially. The intestinal lumen is visible and becomes empty. It is surrounded with columnar and spherical cells. At the apical ends of the columnar cells there are empty vacuoles and in the spherical cells, dense eosinophilic granules. In the bases of cells lie the nuclei surrounded with basophilic cytoplasm. Simultaneously, with the changes in the gut, reorganization of the parenchyma occurs. As a result, new space relations arise between the gut and nervous structures. One hour after eating, there is no change in the structure of the parenchyma. After 2 hr, the parenchimal cells become flat and the volume of parenchyma as a whole decreases, especially in the tail section. After 3 hr, the space occupied by the parenchyma continues to decrease. The walls of the gut, consisting of the large cells filled with food vacuoles and granules, lie close to the lower parts of the nervous ganglion and near the nerve cords. From the intestinal cells the projections, with granular eosinophilic contents, are extended to the parenchyma. At this time similar formations are visible between the nerve cells of the ganglion and near the nerve cords (Fig. 3a).
604
I. M.
SHEIMANN
AND
N. Ju. SAKHAROVA
After 4-5 hr, a further decrease of the parenchymal stratum occurs. The eosinophilic granular formations are distributed at the site of the ganglion and the nerve cords. After 6 hr, the parenchymal tissue remains chiefly under the external epithelium but, in the intergut folds, it is represented by narrow strips of contracted cells. Near the nervous structures large accumulations of eosinophilic granular formations are visible. After 1 day the parenchymal stratum becomes insignificant. In the region of the ganglion lie the stretched cells with granular inclusions. In the gut wall there are cells with projections stretched in the direction of the ganglion and filled with eosinophilic granules. The gut walls are joined to the nerve cords (Fig. 3b). After 2 days there are thin strips of parenchyma between the intestinal loops. The expanded intestinal walls are intimately joined to the ganglion, and almost completely surround the nerve cords. After 3 days the parenchymal stratum remains under the external epithelium and between intestinal loops in the form of thin strips of contracted cells. In the head part there are numerous eosinophilic granular elongated formations. After 4-5 days, the volume of parenchymal tissue increases. The ganglion and intestine are separated from one another. The nerve cords are also separated from the intestine. In the parenchymal stratum which divides the intestine from the nervous system there are granular formations and cells with eosinophilic granules. After 7-9 days a further increase of the parenchyma size occurs. It is filled with different granular formations. At the same time the distribution and quantity of neoblasts in the parenchyma also changes. During the first hours after feeding most of them migrate under the external epithelium where they remain up to the fourth or fifth day. Later on, they become redistributed throughout the body. DISCUSSION
Our data show that in the course of digestion a significant histological rebuilding occurs in the body of the planaria, D. tigrina. The captured food enters the sections of the intestine irregularly but it is usually distributed primarily through the anterior section of the gut. Probably, an asynchronous manner of histological changes in various parts of the intestine is related to this fact. The initial changes are represented by the formation of the projections of gut cells towards the lumen and the food is captured by them. Such projections have been described as early as 1925 (Willier et al., 1925). Further reorganizations of the body are connected with digestion. Almost all the processes of splitting of the food particles occur in the cells of the gut (Willier et al., 1925 ; Jennings, 1957, 1962). The particles of food are split under the action of various enzymes while the food is passing from the apical end to the bases of the cells. The medium of the cells changes from acidic to slightly alkaline. The gradual changes of chromatlinity of food particles from eosinophilia to basophilia and back seem to be due to the changes described above. A similar change of chromaflinity was also observed in other species of planarians (Willier et al., 1925; Horne & Darlington, 1967). During the first 3 days
ON
A PECULIARITY
OF PLANARIAN
DIGESTION
605
after feeding, the gut walls became thicker, When the phagocytosis of the food was completed, the apical surface of the cells gradually became smooth. As the gastrodermal cells enlarge, the intestinal lumen decreases and, on the third day, disappears. Simultaneously the basal membrane which separates the gut from the parenchyma is destroyed. It is difficult to distinguish the cell boundaries. The intestinal walls become symplastic. Five days after feeding, individual cells with different features of destruction are visible again. Most probably, the organism gets rid of wasted cells and in their place new cells, differentiated from the neoblasts, appear. Up to the ninth day restoration of the initial structure of the intestine occurs. When digestion is finished, the food resources are kept in the cells of the gut (Willier et al., 1925; Jennings, 1957). According to our data the whole cycle of digestion in D. tigrina lasts for 7-9 days. The first period-the phagocytosis-lasts for 6 hr and the digestion for 3-4 days. The destruction and the following restitution of the intestinal structure occur from the fifth to the ninth day. The intestine of planarians has two functions: digestion and distribution of nutrients. Organs are supplied with nutrients by means of heavy ramifications of the intestine and by the instability of its cellular structure (Beclemishev, 1964). According to our data this function is also conditioned by the state of the parenchyma. Between the third and the sixth hours a compression of the fixed cells of the parenchyma occurs and the neoblasts are shifted towards the external epithelium. This results in a sharp decrease in the volume of the parenchyma and in an approach of the intestine to the other organs. Some authors consider that the parenchymal cells take part in the transfer of nutrients directly (Jennings, 1957; Rosenbaum & Rolon, 1960; Berg & Berg, 1968). According to other data (Willier et al., 1925) only the gastrodermal cells fulfil this distributive function. The method used by us for staining allows us to distinguish between individual tissues and to observe their changes. The observations show the food particles to be always arranged in the gastrodermal cells independentIy of their state. Three hr after feeding, the projections with eosinophilic granules directed to the parenchyma arise. Perhaps the nutrient is distributed firstly through them. After 34 hr similar granular formations are visible near the ganglion and the nerve cords. Further distribution of nutrients is provided with the deformation of the intestine. On the second to fourth days after feeding, the enlarged intestinal walls come close to the ganglion and surround the nerve cords. In the course of the enlargement and the deformation of the gastrodermal cells and also of the destruction of their membrane, a direct contact is set up between the nervous system and the intestinal contents. According to these data one can say that the transfer of nutrients from the gut to the nervous system begins during the first hours after capture of the food. In the beginning these substances move along the projections and from the second to the fourth day the ganglion and the nerve cords seem to be immersed in the mass of the intestine.
606
1. M. SHEIMANNANDN. Ju. SAKHAROVA
Five days after feeding the parenchyma is restored and the direct contact between the nervous system and the intestine is interrupted. The number of granular formations in the parenchyma increases. They seem to be converted phagocytes (Berg 2% Berg, 1968) or noncellular formations of a different shape which are connected neither with the intestine nor with the nervous system. Probably nutrients are maintained mainly in them. Comparison of the morphological changes during digestion and of the physiological “memory transfer” phenomenon shows a correlation between them. The physiological phenomenon, expressed as an increase of specific excitability for several hours after food intake (5-6 hr), lasts for 3 days, then decreases and disappears on the fifth day. Analysing these data one may suppose that during cannibalism the body of the planaria recipient obtains a substance from the planaria donor and this substance acts specifically on the nervous system of the former. The “transfer” phenomenon is manifested just while there is direct contact between the nervous structures and the intestinal contents. At this time the food particles from the gastrodermal cells are significantly basophilic. Cessation of the “transfer” phenomenon coincides with the destruction of the intestinal cells. Perhaps the enzymes acting at the beginning of digestion only partially break up the food, and the specific properties of the food particles are maintained. Later, during cell destruction, a final disintegration of nutrients occurs and its specific action ceases. It is transformed into a passive nourishing resource.
REFERENCES BECLEMISHEVV. V. (1964) Bases of the Comparative Anatomy of the Invertebrates, Vol. 2. Nauka, Moscow. BERG G. G. & BERG 0. A. (1968) Presence of trimetaphosphatase in the Turbellarian Dugesia tigrina and the relation of enzyme distribution to food uptake. Trans. Am. microsc. Sot. 87, 335-341. HORNE M. K. & DARLINGTONJ. T. (1967) Uptake and intracellular digestion of ferritin in the Planarian Phagocata gracilis. Trans. Am. microsc. Sot. 86, 268-273. JENNINGSJ. B. (1957) Studies on feeding, digestion and food storage in free-living flatworms. Biol. Bull. mar. biol. Lab., Woods Hole 112, 63-80. JENNINGSJ. B. (1962) Further studies on feeding and digestion of triclad Turbellaria. Biol. Bull., mar. biol. Lab., Woods Hole 123, 571-581. JOHN E. R. (1964) Studies on learning and retention in Planaria. In Brain Function (Edited by BRAZIERM. A.), Vol. 2, pp. 161-182. University of California Press. LILLIE R. D. (1954) Histopathologic Technic and Practical Histochemistry, 2nd Edition, pp. 118-119. McGraw-Hill, New York. MCCONNELL J. V. (1962) Memory transfer through cannibalism in planarians. J. Neuropsych&. 3 (Suppl. I), 4248. MCCONNELLJ. V. & SHELBY J. M. (1970) Memory transfer experiments in invertebrates. In Molecular Mechanisms in Memory and Learning (Edited by UNGARG.), pp. 71-101. Plenum Press, New York. ROSENBAUMR. M. & ROLON C. J. (1960) Intracellular digestion and hydrolytic enzymes in the phagocytes of planarians. Biol. Bull., mar. biol. Lab., Woods Hole 118, 315-323. SAKHAROVA N. Ju. (1972) On sources of regeneration in planarians. Ontogenez 3, 95-100.
ON A PECULIARITY OF PLANARIAN DIGESION
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SHEIMANNI. M. (1970) On the complicated structure of the memory trace (in a primitive nervous system). In Memory and Trace Processes (Edited by LIVANOVM. N.), pp. 210-214. Materials of the 2nd conference on the problem of memory and trace processes, Pushchino. WE~TEEMANR. A. (1963) Somatic inheritance of habituation of responses to light in planarians. Science, Wash. 140, 676-677. WILLIER B. H., HYMAN 2. H. & RIFENBURCHS. A. (1925) A histochemical study of intracellular digestion in triclad flatworms. J. Morph. Physiol. 40, 299-340. Key Word Index-Planarians; “memory symplast; projections of the intestinal cells.
transfer”;
digestion;
intestine
structure;