Spleen studiesII. Microscopic observations of the circulatory system of living traumatized spleens and of dying spleensкод для вставкиСкачать
SPLEEN STUDIES 11, MICROSCOPIC 0BSERVATIOE;S O F THE CIRCULATORY SYSTEM OF LIVING TRAUMATIZED SPLRFIK’B, AND OF DYING SPLEENS NELVIN H. KKISELY Bull Laboratory of Anatomy, The University of Chicago INTRODUCTION The first paper of this series (Knisely, ’36) described the structure and some of the activities of the unstimulated splenic vascular system. The observations are that the unstimulated splenic vascular system of mice, rats and cats consists of preformed, lined, completely interconnected vessels. From the structure alone one could easily conclude that the splenic vascular system is ‘closed;’ but the usual concept of the ‘closed’ circulation is too limited to include the functioning of the system, for the venous sinuses separate blood cells from blood fluid. (Venous sinuses in filtering-filling phase retain blood cells but are permeable to blood fluid. Red cells pass through the walls of sinuses during other phases of their cycle by individual penetration.) In the first paper the question was raised, “Why cannot everyone find the connections of arterial capillaries with venous sinuses B ” The observations on unstimulated spleens did not answer this question. Observations of the reaction of the splenic vascular system to trauma and its agonal changes, however, reveal the difficulty of demonstrating the finer vascular system of the spleen in fixed material. ‘Thia work was aided by a grant from the Rockefeller Foundation t o the Biological Sciences Division of The University of Chicago. The a d s t a n c e and counsel of Dr. R. R. Bensley, Dr. William Bloom, and the other members of the staff of Hull Anatomical Laboratory, have been invaluable in this work. 131 THE ANATOXICAL RECORD, VOL 65. NO. 2 A N D WPPLEMENT MAY. 1936 132 MELVIN H. KXISELY Kolliker in his article in Todd’s “Cyclopedia of Anatomy and Physiology” (1847-1849) pointed out the danger of rupturing the fine splenic vessels by injection. On page 191 he says, How these smaller veins are connected with the very distinct capillary network of the pulp I have not been able to find out ; and I do not believe that either injection or inflation of the vein or a microscopic examination will ever give any definite conclusion hereto. F o r these vessels, often possessing but a few little trabeculae f o r their coats are of such delicate texture that they tear by the slightest inechanical force while by the microscope they cannot be distinguished from the surrounding constituents of the pulp. Janosik (’03) who was able to demonstrate the fine vessels of the spleen and their connections emphasized that the injection pressure must be as low as possible. He says on page 580, Wenn man die Milz von der Arteria lienalis her und zwar bei einem moglichst niedrigen Drucke injiziert, so kann man an zahlreichen Stellen einen direkten Ubergang der kleinen Arterien, welche insgesamt Endarterien sind, in kleine Venen nachweisen. Um die Xalpighischen Korperchen herum findet man fast immer Extravasate. Wenn man sich nur des Rerlinerblaus ohne Zusatz irgend eincr Leimmasse bedient, so sind die Extravasate sparlicher und hei der Katze finde ich so ausgedehnte zusammeilhiingende Netze von wohlbegrenzten und zwar mit Endothel ausgekleideten Bahnen, dass man an ein vollstandig geschlossenes Rlutzirkulationssystem in der Milz im Sinne von Sokoloff denken kann. After describing the reactions of tlie splenic vascular system to trauma and its agonal changes it is possible to discuss the ‘ampulla of Thoma. ’ The observations here reported were made using the same transillumination method as was used to study the unstimulated splenic vascular system. TRAUMATIZED SPLEEXS AND DYING SPLEENS 133 OBSERVATIONS I . T h e reaction of the vascular system of living spleens of mice, rats and cats t o s t i m d i produced by esperimeatal manipulation The splenic vascular system is much more sensitive, and more reactive to the stimuli produced by experimental manipulation than is the circulatory system of any other organ I have yet studied. If the surface of the living spleen is cut slightly, two reactions take place immediately. First, great numbers of the previously described physiological sphincters contract. So many shut off the flow that sometimes one has to look for a long time to find any moving blood in the parts of the organ which are accessible for microscopic study. The sheaths of Schweigger-Seidel are especially reactive and powerful. They shut down more quickly and stay closed longer than the other sphincters. Secondly, great numbers of red cells pass out into the pulp partitions through the walls of those sinuses that are in storage phase. (At times during the passage of the red cells it appears as though the sinus had contracted, increasing the pressure on its contents. However, if there is a change in volume it is so small that i t cannot at present safely be used as a criterion to determine whether or not the sinus contracts, and in the absence of other criteria I cannot now be certain either that it does or does not.) The first red cells to pass through the sinus walls are distorted and then snap forward, showing that the membrane lining the sinus is at first intact. The red cells come through so fast, however, and the tissue around the sinuses is soon so clouded that one can no longer see the sinus wall or study the passage of erythrocytes through it. (This passage of red cells out of sinuses is not at all like the escape of whole blood from a small vessel during a microscopic hemorrhage.) The rapidity of passage and the numbers which leave, seem to indicate that the resistance of the sinus wall to the passage of red cells has been altered. 134 MELVIN H. KNISELY Observation of individual red cells which have passed into pulp partitions following stimulation has as yet failed to show that any of them are phagocytized or cytolyzed. They ultimately return one by one to venous sinuses and pass in through the walls. The last erythrocytes passing inward are distorted and snapped forward just as they are in nnstimulated spleens. After a variable period of time, sometimes within 30 minutes later, the routine activities of the unstimulated spleen begin haltingly to take place again. Usually the system does not fully regain the smoothness and uniformity of action that it had before being traumatized; as a rule, under these experimental conditions, there is still an excessive number of red cells left in the pulp partitions at the end of 12 or 15 hours. The latter seems to indicate that under some conditions red cells can be in the pulp partitions for long periods without being phagocytized. This same set of changes, namely, clamping of sphincters and passage of red cells into pulp partitions, occurs whether the stimulation consists of cutting, pinching, scratching, stretching, or twisting the spleen, the gastrolienal ligament, or the vessels and nerves at the hilus. These reactions occur if the spleen is touched with perspiring finger tips or with metallic instruments which are at room temperature. Similar reactions occur, but more slowly, if the temperature of the Ringer's wash solution varies more than 1.5"C. from the temperature of the abdominal cavity of the animal. Thus it would seem that Barcroft and Stephens' report ( '27) that exteriorized spleens are insensitive to trauma, needs further interpretation. Their criterion was that the animal did not show signs of pain when the exteriorized spleen was traumatized. The observations here reported show that the vascular system of the spleen is very sensitive to tranma. Here the criterion is the observed reaction of the living vascular system. These two sets of observations do not contradict each other, for different criteria are used and, as is well known, there are many visceral reactions which do not affect consciousness. TRAUMATIZED SPLEEXS AND DYING SPLEENS 135 From the variety of stimuli which initiate vascular reactions, one can see why the living organ must be so vigilantly protected by careful manipulation when microscopic studies are to be made. Owing to this sensitivity and reactivity, the first spleens studied in this work were seen in the stimulated condition. Only by looking for source after source of stimulation, and eliminating each when it was recognized, was it possible to begin to study the vascular system of this organ while it was free from such traumatic stimulation. The greatest single obstacle in observjng the living unstimulated spleen microscopically, is the constant movement imparted to it by the diaphragm. Every effort to hold it still, even holding the stomach in a clamp, initiates the reactions to trauma. (Incidentally, those specimens are most favorable for study that have relatively long gastrolienal ligaments.) It is entirely possible that the closing of large numbers of sphincters and the migration of erythrocytes constitute a part of the reactions of the system to unknown stimuli that reach the organ during the normal lifetime of an animal. It is believed, however, that none of the stimuli listed are those to which a spleen is subjected during an animal’s normal life. Yet it is significant that such irritating processes almost invariably constitute a part of the preparation of a spleen for injection, fixation by perfusion, or fixation by puncture or immersion. Such traumatic stimulation accounts for the presence of some of the red cells found in the ‘cords’ of fixed and sectioned spleens. Indeed the clamping of the sphincters (especially of the sheaths of Schweigger-Seidel) throws light upon several kinds of results obtained in splenic experiments. For instance : 1. It accounts for the difliculty of injecting the splenic pulp by way of the arterial system: a) Barcroft’s demonstration ( ’25)that the spleen of a dead animal has contracted to much less than its physiological size, b) Robinson’s (’26) observation of the contraction of excised spleens, and c) Mall% statement ( ’00, p. 36), that “Their muscle walls (meaning the 136 MELVIN H. KNISELY arteries) are also so powerful that it is practically impossible to inject them completely in a contracted spleen.” 2. It is probably the reason why injection masses forced into the splenic veins do not come out of the arteries. The initial clamping of sphincters prevents the immediate intravascular transportation of the material, and the subsequent pressure on the outside of the arteries of fluid that had escaped through the venous sinus walls collapses the arteries, making it still more difficult for fluid to enter them. Nisimaru and Steggerda (’32) kept spleens in a carefully controlled constant temperature bath while injecting them, and bogan their injections at low pressures. It is indeed significant that under these conditions material injected into the veins came out through the arteries. 3. The clamping of sphincters may well account for some, though not all, of the holes seen in the walls of the finer arterial system of certain spleens fixed by injection. For consider the alteration in the pressure relationships in the arterial system which simultaneous clamping of great numbers of sphincters brings about. During the flow of blood in the arterial system of an unstimulated spleen there is a progressive decrease in pressure from the large arteries to the arterial capillaries, due to the friction of the moving blood on the vessel walls and the increase in total vascular cross section area. If one small arteriole contracts tightly enough to obliterate its lumen, its blood flow stops and the pressure in it, afferent to the constriction, rises until it equals that in the artery from which the arteriole branches. In an unstimulated spleen some of the branches of a given stem conduct blood while others are closed. There is a rotation of access to the blood supply so that even during the intermittent flow in terminal vessels, there is a progressive decrease in pressure from large arteries to arterial capillaries. But, when large numbers of sphincters on the latter simultaneouly close, the flow in them is stopped and the pressure in the fine vessels increases, approaching as a limit that in the main splenic arteries. TRAUMATIZED SPLEENS AND DYING SPLEENS 137 It is but natural to assume that if the pressure head at which an injection mass is being forced into an organ is that normal for the large artery being injected, the pressure throughout the arterial system therefore corresponds to the normal pressures at each point in the system. But this is not necessarily (in fact, seldom is) the case. The viscosity of the injection mass and the rate of flow at each point are also factors affecting the pressure throughout the vascular system. If the spleen is injected when large numbers of sphincters are shut, the pressure in the finest vessels can (previous to either rupture of fine vessels or forcing sphincters open) be very nearly equal t o the main pressure head at which the injection is being made. Rupture of the more delicate vessels would permit the injection mass to enter the splenic pulp but it would leave holes in the walls of the finest branches of the arterial system. (Compare Kolliker’s statement of the delicacy of these vessels, and Janosik’s that he demonstrated them when the injection pressure was as low as possible.) TI. Observatio%s om the rapid changes in the spleen durimg t h e death of the animal F o r a description of the changes taking place at death the notes on mouse B8, a young animal having a thin spleen, may be used. The activities proceeded too rapidly to be written by the observer; they were recorded while being made by dictation to a stenographer. Each factor has been observed in rats and cats, as well as in mice. -Mouse B8. Previous to the death of the animal an area at the junction of arterial capillaries and venous sinuses was under observation. A venous sinus was emptying into a small vein. White blood cells were visible in blood moving slowly in the long straight capillaries of the pulp partitions. The capillaries all showed the characteristic refractile lining mentioned in the previous paper. The animal had been opened but a short time and the processes under observation were those regularly seen in unstimulated spleens. Without warning, at 9.02 P.M., the heart stopped beating. Irregular respiratory motions continued. 138 MELVIN H. RNISELY At the moment the circulation stopped erythrocytes began to pass through the visibly intact linings of capillaries and sinuses into contiguous tissue. As each one penetrated the lining it was distorted and snapped forward. Red cells in the marginal zone of the malpighian body and in pulp partitions went in all directions, each by itself. A large clear cell shaped roughly like a prolate spheroid moved twice the diameter of a red cell in 1minute. The red cells moved much faster. Incidentally, at this time the intercellular spaces of the pulp partitions probably were somewhat less than 7 p in width for the red cells did not turn over freely in them. At 9.05, as the animal made its last repiratory movement, I was observing the subdivisions of an arteriole. Using the long focus 40 X water immersion objective, 10 X oculars and a green light filter I could see breaks appearing in the refractile linings of the capillaries. At 9.07 a 15 X ocular was substituted for one of the 10 X ’s so that the field could be observed a t either 400 or 600 diameters. One erythrocyte was watched a few moments a t 600 X magnification. When the field was again surveyed at 400 diameters the number of red cells in the marginal zone and pulp partitions had increased noticeably since 9.02, but each one was traveling more slowly than before. As breaks occurred in the capillary walls many highly refractile clear cells, some rounded, some irregularly starshaped, and all constantly changing form, appeared in the field.2 Whether they became visible by moving into focus, Since those observations were made Foot’s ( ’27) article ‘lOn the endothelium of the venous sinuses of the human spleen” ha5 been brought t o my attention. In addition t o studying fixed tissues he made hanging drop prepmations of sera@ spleen (from a spleen that was still warm) and examined them in a warm chamber under the microscope. His in vitro observations are very pertinent to the description of the dying spleen. Foot says, l‘h study of the living cells of the spleen was not very illuminating, owing to the ditficnlty experienced in identifying them; many large cells were found t o be extraordinarily active, quite as lively as fresh water amebae, but it could not be determined whether they were all mscropha@s or whether some of them were not detached endothelium. Oells showing the rod-like morphology of the latter were found, but they were not actively motile. . No membrane nor fragments thereof muld be found h spite of careful searching.” . . TRAUMATIZED SPLEENS AND DYING SPLEENS 139 o r by alteration of their refractility in situ or by both methods could not be determined. While these cells increased in number the refractile borders of the capillaries disappeared. The breaks in the capillary walls were real, for red cells passing through them did not distort and snap forward; they apparently met no more resistance than in progressing through pulp partition tissue. Then large clear irregular cells began to ingest red cells. As a preliminary to the ingestmion,the red cell turned or was turned, so that one flat side was closely apposed to the surface of the ingesting cell. As many as three erythrocytes were seen on one large cell ; then suddenly the red cells were inside the phagocyte. The actual passage through the surface was too fast to see. Phagocytic cells moved through the pulp partition tissue and acquired two, three or five red cells. One was seen with seven in it. Those in this phagocyte then began to turn darker and fragmented, until only two were left. The cytoplasm of the phagocyte meanwhile acquired a dark reddish-grey color. The processes of ingesting and breaking up erythrocytes proceeded simultaneously so that any one phagocyte seldom had more than from three to five red cells visible in it, even though it could be seen to ingest two or three times that number. By this time no vestige of capillary walls was visible and even the end portions of the penicilli branches were invisible for short stretches. By the time the last capillary linings had disappeared, the number of phagocytes visible at any one time had reached its maximum. The red cells in partly filled venous sinuses became arranged in rouleaux. (In 4 years of watching circulation in many organs of small laboratory animals, I have not yet seen rouleaux formation in the living fEowi.ng blood, except under known pathological conditions such as ether anesthesia, pressure or tension on the tissue, trauma, o r preceding the death of an animal.) From 9.11 on the rate of the agonal activities rapidly decreased to a few dow movements of phagocytes. All the visible changes in the structure of the vascular system, the 140 MELVIN H. KNISELY movement of red cells from vessels to tissue, and the intense phagocytic activity took place in about 10 minutes ; most of it much sooner. Red cells pass through the lining of the sinuses and through the positions previously occupied by capillary walls without being distorted and snapped forward. At the end of the agonal changes the sinus walls usually have a ragged appearance. Because the clouds of red cells surrounding the sinuses make the changes in their walls more difficult to see than those of capillary walls, they have been less frequently observed. So the conclusion cannot here be drawn that they occur more rarely. At the end of the death period there is less variation in the diameters of the several venous sinuses of an area than there is in the living. Few are as greatly distended as in an unstimulated spleen, nor are the ones which were last in conducting phase as slender as they had been in the living organ, and the corresponding pulp partitions are therefore more nearly equal in thickness. DISCUSSION OF OBSERVATIONS Some of the agonal changes are more pronounced in one spleen, some are more obvious in another. Every change is usually seen in some degree. In these experiments the beginning of these processes has invariably been proof-positive that the death of the animal was imminent. These agonal changes are probably the source of many of the conflicts in the reports of the structure of the finer splenic vascular system. Consideration of the fact that the agonal changes can be well on their way in 5 minutes and can have gone to completion in 10 minutes, shows the probability that in many cases of fixation by immersion the agonal reactions have begun or gone to completion before the fixative has reached the tissue. Tellyesniczky (’26 edition of Krause’s Enzyklopodie der mikroskopischen Technik, p. 757) measured the rate of diffusion of fixatives into pieces of immersed liver. The slowest rate was 4 to 1mm. in 4 hours. The most rapid TRAUMATIZED ST’LEEFS A 6 D DYIFQ SPLEENS 141 was 1 to 1.25 mm. in 1 hour. X y own measurements (using Tellyesniczky ’s method) show that Romeis’ fixative penetrates mouse spleen a little less than 3 mm. in the first half hour. Mall (’00) makes a statement pertinent t o the question of the adequacy of fixation by injection. He says, ‘‘If artificial circulation is carried on through a spleen in which the muscle is alive and contracted it is found that the spleen must be distended considerably before any fluid comes out the vein. This usually takes a number of minutes after the artificial cjrculation has started.” I n fixation by injection (via artery or vein) the physiological sphincters, traumatically stimulated by the presence of the fixative in the larger vessels or by handling of the organ, could easily stop the flow of fixative to the finer vessels in the agonally reacting area, so that the fixative must either 1) diffuse under pressure through the vessel walls (which takes time), 2) force the sphincters open, or 3) rupture fine vessels. If the vessels are ruptured by pressure or if the agonal reactions have set in before fixation occurs, the finer vascular system of the living unstimulated spleen is no longer complete. Perfusion with saline previous to fixation does not necessarily eliminate the danger of rupturing the vessels. Thus, beginning with the structure of the living unstimulated spleen, different combinations of variations of 1) the number and positions of sphincters which contracted shut, 2 ) the tightness with which they closed, 3) the amount of rupturing caused by injections, 4) the rates of progress of the agonal changes in different spleens, and 5) the time the agonal changes were in progress previous to the fixation of this reactive tissue easily account for the ‘closed’ and all the ‘open’ and compromise circulatory system patterns that have been described from sections of fixed spleens This accounts for the fact that from one part of a fixed spleen a ‘closed’ pattern may be described and from another part ‘open’ patterns, as in Weidenreich’s (’01) classical study, for there is no reason to believe that each of these variables operates equally in all parts of a single spleen. 142 MELVIN H. KNISELY The presence in fixed spleens of holes in the walls of capillaries, slits in venous sinus walls, open-ended capillaries, and whole and fragmented red cells in phagocytes are not evidence that the same features existed under the immediately previous physiological conditions unless it is absolutely certain that the fkation procedure employed does not itself cause changes in the organ, and is rapid enough to prevent the initiation of the agonal changes. It seems probable that the freezing-drying technique (Gersh, ’32) by virtue of the possibility of freezing a whole live mouse or an exposed functioning spleen instantaneously, will prove suitable for studying spleens that have not undergone traumatic and agonal changes. It may well be that some or all the phenomena seen in dying spleens take place as reactions to stimuli which reach the organ during an animal’s normal life. I feel that at present we are in complete ignorance on this point because such features have not yet been seen in living spleens and clearly we cannot be certain that the fixation of the spleens which have previously shown these features was adequate to prevent the traumatic and agonal reactions. Since it has been taught for so many years that the spleen is “the graveyard of the red cells,” we must weigh carefully all evidence of the splenic phagocytosis of whole red cells. Phagocytes have not yet been seen ingesting and cytolizing erythrocytes in living unstimulated spleens (Knisely, ’36). They are phagocytized in dying spleens, and red cells and their decomposition products are not infrequently present in the phagocytes of fixed spleens. Red cells can be either stationary or moving in the pulp partitions for considerable time without being phagocytized. Apparently splenic phagocytosia of erythrocytes does not necessarily progress at a uniform rate as is sometimes assumed. DISCUSSION OF AMPULLA OF THOMA One of the problems in this work has been the identification of the structures Mall named ‘ampulla of Thoma’ (Thoma TRAUMATIZED SPLEENS AND DYING SPLEENS 143 1895 and 31al1, ’00 and ’02). Mall saw the connections of ampullae with veins for in his 1900 paper (p. 33) he says: “There are then marked channels or ampullae of Thoma, connecting the arteries with the veins, which can be demonstrated by certain methods.” Thoma showed that the distal end of the ampulla had a mechanism which prevented injected material from entering the veins, for he says (p. 48) : Letzterer zeigt, wenn die Injectionsmasse nicht in die Venen vordrang, haufig eine ampulltire Erweiterung, welche beweist, dass der Druek der Injectionsmasse in dem Arterienendzweig ein sehr hoher war, somit irgend welche Vorrichtungen bestehen miissen, welche das Weiterschreiten der Injectionsmasse verhindern. Since Mall’s papcrs were written, different authors have used the term ‘ampulla of Thoma’ to designate various structures. For example, Robinson ( ’26) used the term to indicate an exaggerated space in the splenic pulp tissue. He says (p. 352), “ F o r these reasons we believe that the ‘Ampulla of Thoma’ is merely an exaggerated pulp space produced, no doubt, by the concentration of blood flow at this point.” MacNeal ( ’29) describes small ampullary dilatations at the ends of several types of capillaries. Such pulp space dilatations as Robinson and XacNeal described have not been seen in living spleens. Tait and Cashin ( ’25, p. 424) equated the term ‘ampullae of Thoma’ to the term ‘sheaths of Schweigger-Seidel.’ McNee (’31) followed their usage. But Mall (’02) clearly distinguished between the ellipsoids and the ampulla of Thoma, for he says, “The ellipsoid of Schweigger-Seidel continued to the end of the artery as a small group of round cells and marks the beginning of the Ampulla.” The structure of his ‘ampulla of Thoma’ cannot be confused with that of the sheaths of Schweigger-Seidel. H e says, The first third of the ampulla is lined with spindle-shaped cells which are directly continuous with the endothelial cells 144 MELVIN El. KBISELY of the artery. The second third branches and often communicates with neighboring ampullae. The last third of the ampulla is difficult to demonstrate but under certain conditions it can be injected as has been shown by Thoma and by me. Mall also says, ‘‘Free communications between the ampullae and veins have been seen from time to time by numerous investigators from W. Xiillcr to Helly, but no one has ever been able to find them in great number nor free from extravasations.” And describing the connections of the structures Mall calls ‘ampullae,’ he says, If a spleen, made oedematous by injecting gelatin into either the vein o r the artery or by filling the puly! with blood by ligating the vein for half an hour, is injected through the artery with some fine granular mass, like Prussian blue, it is found that at the end of each arterial-capillary there are a number of ampullae which communicate with the pulp spaces, with one another, and with the veins. Frequently, looking into a living unstimulated spleen, one sees a somewhat elongated, cucumber-shaped structure, filled with blood cells and appearing to have no efferent connection with the venous system. Brief, uncritical observation of such a structure could convince one that this is Thoma’s o r Mall’s ‘ampulla of Thoma.’ Each time that such a structure has been carefully watched for a long time, a previously unohserved efferent sphincter has suddenly opened wide and the structure has discharged its stored blood cells into an adjacent venule. This structure, the static condition of which roughly fits Mall’s description, proves by discharging that it is a venous sinus in the storage phase of its activity. Such structures always proceed to repeat the regular venous sinus cycles. Mall’s statement that efferent connections with venules have been found from time to time but not in large numbers, supports the view that his ‘ampullae of Thoma’ were venous sinuses because, in a fixed preparation, the only efferent connections that would be discovered are the patent ones. Under most conditions more would be closed than open. Mall’s statement that in a spleen distended with gelatin (or with TRAUMATIZED SPLEENS A B D DYIKG SPLEENS 145 blood by ligating the veins) the ‘ampullae’ connect with the gulp spaces, with one another, and with the veins, clearly describes the postmortem anatomical relationships of the individual sinuses constituting a multiple sinus system (compare I, 11,I11 and I11a in fig. 2 of the first paper of this series). Further, 3Call’s statement that the first third of the ampulla is lined with “ spindle-shaped cells which are directly continuous with the endothelial cells of the artery” equates nicely with Mollier ’s ( ’11) description of the spindle-shaped cells lining the venous sinuses in his preparations. Mall says, “The second third branches and often communicates with neighboring ampullae.’’ This sentence is an apt description of the interconncctions of a multiple sinus system. He says that the last third is difficult to demonstrate, but that Thoma and he have been able to inject it. He states also that the last third connects with a venule by a channel or channels which have bridges of tissue across them. This latter statement rather accurately describes the ragged condition of some sinuses’ efferent ends after the agonal changes have gone to completion. The fact that Mall’s fluid injection masses ‘extravasated’ through the walls of his ampullae also checks well with our present knowledge of the permeability of the sinus wall io blood colloids (Knisely, ’36). From these considerations I am led to conclude that the structures which Mall ( ’00) described and called ’ampullae of Thoma’ were venous sinuses fixed in different phases of physiological activity and in different stages of agonal disintegration. After another series of researches (’02)’ during which Mall injected spleens with, as he says, “ a great variety of mixtures of asphalt, turpentine and granules” and gelatin, he concluded that “It seems as if the ampullae are only large holes within the spongy pulp.” This is the same conclusion that Robinson reached after injecting spleens from abattoirs and autopsies with asphaltum in turpentine, hot lard, etc. THE ANATOMICAL RI~CORD,VOL. 65, NO. 2 AND SWPPLJCMEXT 146 MELVIN H. KNISELY SUMMARY A variety of manipulative stimuli initiate two reactions in living spleens : 1) Simultaneous complete contraction of a great many physiological sphincters, particularly the ellipsoids of Schweigger-Seidel, and 2) the passage of large numbers of red cells from sinuses in storage phase into the pulp partitions. This explains 1) the difficulty of injecting the splenic pulp by may of the arteries, 2) the reason why material injected into the veins does not come out the arteries, 3) the presence of some of the holes in the walls of fine vessels of injected specimens, and 4) the presence of part of the red cells found in the pulp ‘cords’ in sections of spleens. The agonal changes consist of 1) migration of red cells into pulp tissue, 2 ) disappearance of portions of arterial-capillary and capillary walls, 3 ) the appearance of irregularly shaped, moving, constantly changing cells which rapidly ingest and cytolyee erythrocytes, 4) the development of spaces in the venous sinus walls large enough for red cells to pass through without being distorted, and 5) the relaxation of distended sinuses and stretched pulp partitions, which alters the tissue so that the relative magnitudes of its parts are not the same as in the functioning organ. The most striking features of the agond changes are the ,instantaneousness of their onset and the rapidity with which they go to completion. The observations on the reactions of the spleen to trauma, and the rapid agonal changes which take place in the organ show why it is so difficult to trace the connections of the arterial capillaries with venous sinuses in fixed sections. The term ‘ampulla of Thoma,, coined by Mall, was used by Robinson to indicate large pulp spaces present in the tissue after injecting foreign materials. It mas used by Tait and Cashin, and later by &Nee, as a synonym for the terms ellipsoid, Capillarhiilsen, and sheath of Schweigger-Seidel. Mall used it to designate the structures commonly known as ‘venous sinuses’ in the papers describing his early experiments. But in his 1902 paper, after injecting asphalt in TRAUMATIZED SPLEENS A N D DYIFG SPLEENS 147 turpentine, etc., he applied the term ‘ampulla of Thoma’ to the large pulp spaces he found, just as Robinson did 24 years later, after similar treatment of the organ. The term ‘ampulla of Thoma’ does not indicate a specific antttomical structure, but has been applied to two real structures and to artificial dilatations produced by injection. The observations reported in these two papers show the activities and reactions of the splenic vascular system under a limited set of conditions. 1 do not think that we can yet safely conclude that the activities and reactions of the system are necessarily similar under all other conditions. LITERATURE CITED BARCROFT, J. 1923 Recent knowledge of the spleen. Lancet, vol. 208, pp. 3 19-322. BARCROIT,J. AND J. B. STEPHENS 1927 Observations on the size of the spleen. J. Physiol., v01. 64, pp, 1-23. FOOT,N. C. 1927 On the endothelium of the venous sinuses of the human spleen. Anat. Rec., vol. 36, pp. 91-102. GERSH, I. 1932 The Altmann technique for Bxation by drying while freezing. Anat. Ree., TOI. 53, pp. 309-337. ;JANOSIK, J. 1903 Uber die Blutzirkulation in der Nilz. Archiv. f. mikr. Anat., Bd. 62, 8. 580491. K~LLIKER, A. 1847-1849 The article on the spltien in Todd’s Cyclopaedia of Anatomy and Physiology, vol. 4, pp. 771-800. Longman, Brown, Green, Longmans and Roberts, London. KXISELY, M. H. 1936 Spleen studies. I. Microscopic observations of the circulatory system of living unstimulated mammalian spleens. Anat. Rec., VOI. 65, pp. 23-50. MACXEAI,,W. J. 1929 The circulation of blood through the spleen pulp. Archiv. of Path., vol. 7, pp. 215-227. MALL, F. P. 1900 The architecture and blood vessels of the dog’s spleen. Zeitsehr. f. Morphol., Bd. 2, S 1-42. 1902 On the circulation through the pulp of the dog’s spleen. Am. J. Anat., 701. 2, pp. 315-332. MCNEE,J. W. 1931 The Lettsonian Lectures on “The SpIeen: its structure, functions, and diseases." Trans. Med. SW. of London, vol. 54, pp. 185-236. XOLLIER, 5. 1911 Uebor den Bau der Capillaren Milzvene (Xilzsinus). Archiv. f . mikr. Anat., Bd. 76, S. 608-657. 1932 Observations on the structure and funcNISIMAEU, P. AND F. R. STEGGERDA tion of certain blood vessels in the spleen. J. Physiol., vol. 74, pp. 327-337. 148 MELVIN H. KNISELY ROBINSON, W. L. 1926 The vaseuhr mechanism of the spleen. Am. J. Path., VOI. 2, pp. 341-356. TAIT, J. AND M. F. CASHIN 1925 Some points concerning the structure and function of the spleen. Quart. J. Exp. Physiol., vol. 15, pp. 421-445. TELLYESNICZKI, I(. 1926 The article on ' 'Fixation, " in Krause 's Enzykklopiidie der mikroskopisehen Technik, pp. 750-785. Urban and Schmarzenberg, Berlin and Wien. THOMA,R. 1895 Ueber die Blutgefiisse der Milz. Verhandl. der Anat. Gesellwhaft, Bd. 9, S. 45-52. WEIDENREICR, F. 1901 Daa (3efZsssytem der Milz des Menschen. Arehiv. f. mikr. Anat., Bd. 58, 8. 247-373.