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Under specially controlled conditions in the healthy unanesthetized dog we have measured the resistance offered to the flow of bile through the lower common duct. The average resistance 4 to 12 hours after a feeding was found to correspond to the pressure of a column of bile 100 to 120 mm. in height. After a 24 to 72 hour fast the resistance was such as to support a much higher column of bile, one of 300 mm. at times. The exhibition of food to the fasting animal usually precipitated a reflex lessening in the resistance to the flow of bile to the intestine, the actual taking of food always brought it about. This was transient and was soon followed by a period of increased resistance lasting 10 to 30 minutes after food had entered the stomach. There succeeded a drop in resistance which was gradual and fluctuating. We have observed an increase in the resistance to the flow of bile into the intestine after alkali has entered the stomach, and a decrease after acid has been administered. We attribute the sudden changes chiefly to the activity of the sphincter of Oddi.

Local intradermal injections of dye have disclosed the fact that the skin lymphatics in regions of cardiac edema are patent, full of fluid and much widened. Intercommunication between them is ready and dye escapes from them more rapidly than from the vessels of normal skin. A retrograde distribution of dye by way of the lymphatics often occurs and it may pass unseen along the deeper channels to emerge in the skin at unexpected, distant situations. A valvular incompetence of the lymphatics consequent on dilatation would appear to be the cause of these phenomena. In regions of cardiac edema lymph stagnates, despite the fact that the channels are open. In nephritic edema the lymphatic capillaries are wider than normal but not as wide as in cardiac edema. No sign of incompetency of the valves is to be observed. Instead lymph flow is considerably greater than normal, even when edema fluid is accumulating. It was noted to be greater in the periods of fluid equilibrium also and extraordinarily rapid in periods of diuresis.

The gradient of permeability along the capillaries of voluntary muscle and the capillaries and venules of skin exists independently of the hydrostatic conditions, though influenced by them. Its presence cannot be explained by a graded tonic contraction of the capillaries. The evidence,—like that of previous papers,—points to local differences in the barrier offered by the wall of these vessels as responsible for the gradient.

The gradient of permeability which exists along the cutaneous capillaries and venules is accentuated and broadened in scope by increasing the venous pressure moderately. Under such circumstances transudation leading to edema takes place most abundantly from the venules. The permeability of the portion of the capillary web that is near the arterioles increases only when the venous pressure rises so high as to approximate that in the arteries. Under such circumstances the gradient of permeability along the small vessels disappears, the capillaries and venules everywhere leaking fluid. The character of the vital staining developing under such circumstances indicates, like the evidence of previous work, that the cause for the gradient is to be sought in a structural differentiation.

The ears of rabbits were perfused with defibrinated rabbit's blood in such a way that pulsation could be imparted to the perfusate or withheld from it at will. In the absence of pulsation there was almost no lymph flow, whereas when it was present lymph flow was rapid despite the fact that the "systolic" pressure of the perfusate never exceeded the constant pressure in the non-pulsatile instances and the volume flow was far less. Non-pulsatile perfusion led to a slight flow of lymph in ears that were becoming edematous, whereas when it was pulsatile the lymph flow was enormous. The pulse exercises an influence to move fluid into the lymphatics and along them.

We have endeavored to learn whether rabbits manifesting the symptoms characteristic of the advanced stage of liver insufficiency1, 2 can be clinically improved by the circulation of their blood through the livers of healthy animals or by cross transfusion with normal rabbits.“Liver transfusion.” In an initial series of 14 experiments, rabbits, hepatectomized under ether and with cannulae placed in the left carotid artery and left jugular vein, were given sufficient glucose to maintain the blood sugar level well above normal. Fourteen to 24 hours later, when the characteristic signs of advanced hepatic insufficiency in the rabbit1, 2 had appeared, the portal vein and vena cava connecting with the liver of a healthy animal were rapidly cannulated, the hepatic artery was tied and the organ was removed. It was submerged at once in a bath of paraffin oil at 40° C., and connected with the circulation of the liverless animal by means of the cannulae already present in the carotid artery and jugular vein of the l...

The changes of intracutaneous pressure in the limbs of mice and human beings have been followed during and after periods of venous obstruction with almost unhindered arterial flow. During the first 30 minutes of obstruction the interstitial pressure in the tense skin of the lower legs of mice, a pressure which is slightly higher than that in the loose skin of the ears, backs, and thighs (21), rose from 2.6 to 4.6 cm. of water to about 32 cm., thereafter remaining constant. It would appear that the escape of fluid from the capillaries is checked at this pressure. In the skin of the arm and leg of man the interstitial pressure rose from 2.5–3.7 cm. of water to 15.0–23.0, within 15 to 27 minutes after venous obstruction had been produced, mounting no higher during the period of observation. When venous obstruction had existed for about 20 minutes or more the subjects sometimes experienced sensations of relief from congestion as if some tissue adjustment or the opening of some venous by-pass in the marrow had occurred, preventing a further rise of pressure. However this may be, the pressures still appeared to be great enough to prevent further escape of fluid from the capillaries, at least for the time being.

The selective deposition of hemosiderin in the liver parenchyma during pernicious anemia does not constitute evidence that there is a hemolytic cause for the disease located in the portal region. The repeated introduction of small amounts of free hemoglobin into the general circulation, by the subcutaneous route, leads, as we have shown, to an identical siderosis. Larger amounts of hemoglobin cause a renal pigmentation equalling or exceeding the hepatic, a fact that is in keeping with what is known of the physiology of hemoglobin excretion and of the findings in human beings afteroutspoken hemolysis.

A standardized solution of a vital dye which escapes with some difficulty from the lymphatics of the ear of the mouse has been utilized in tests of the permeability of the lymphatic wall under various conditions. It has been found that this permeability is subject to great change. The slight pressure that suffices to prevent lymph flow from the ear,—an organ in which such flow goes on normally,—soon results in increased permeability of the obstructed lymphatics without as yet any perceptible dilatation of these vessels. Mechanical stimulation as for example a stroke with a blunt wire, or scratching so light as not to break the epidermis, results in a practically immediate, great increase in lymphatic permeability, which is sharply localized to the region pressed upon. This increase in permeability, though so great that even hemoglobin is let pass by the lymphatics, endures but a few hours. Warming the ear to 43°C. or exposure to mild sunlight increases permeability considerably. Slight chemical irritation increases it greatly, though not so much that particulate matter is let pass. The edema developing as result of lymphatic obstruction or mechanical, thermal, or chemical stimulation is preceded by and associated with a large increase in lymphatic permeability. The facts are discussed in relation to their bearing upon fluid accumulation within the tissue. It is plain that influences within the realm of the normal suffice to increase lymphatic permeability and that those which lead to edema cause a very great increase in it. In proportion as this increase occurs the lymphatics cease to be channels demarcated by a semipermeable membrane. It seems certain that the changes must be in some part responsible for the local accumulation of fluid. There exist possibilities, on the other hand, of a correlation between the functionings of the blood and lymph vessels under certain pathological conditions, as during the resorption of edema.

In rats fasted after the removal of about seven-tenths of the liver the remaining fragment undergoes ordinarily a marked simple hypertrophy and attains the weight reached through a process of simple atrophy by the entire liver of fasting controls. Under circumstances of exceptionally severe inanition, the hypertrophy may not occur, the fragment remaining unchanged or even undergoing a slight atrophy. But since in comparable controls the hepatic atrophy is extreme, the duplication in the liver weights still manifests itself. It is difficult to avoid the conclusion that the liver changes which take place in fasting animal are essentially conditioned on functional demands made upon the organ. Yet great as is the hepatic atrophy then occurring, this does not necessarily mean that these demands are much lessened during inanition. For a comparison of the liver weight with that of the soft body (net body weight less the weight of the ligamentous skeleton) brings out the fact that the organ is, relatively speaking, three-fourths as large at extreme inanition as in the well nourished individual. Of the missing fourth a part may be foodstuffs on temporary deposit. And the rest is perhaps to be accounted for by the existence of special conditions during inanition which favor the vicarious assumption of a part of the usual liver work by the other tissues. While the findings show that the liver size is essentially dependent on functional demands, they do not enable one to say whether the special demands in question come normally to the organ by way of the portal stream.

Means have been described for the study of pressure conditions in normal and pathological skin of living human beings and mice. The true pressure in normal skin cannot be measured directly by any of the means hitherto described, because there is insufficient free fluid to make manometric determinations. However, for practical purposes, the intracutaneous pressure has been approximately estimated by introducing into skin exceedingly small amounts of a relatively unabsorbable fluid, a mixture of Locke's solution and a vital dye, and then finding the least pressure required to overcome the resistance of the skin to the passage of this fluid through it at the lowest rate measurable with accuracy by the apparatus at hand. In the present paper measurements of this pressure have been termed the interstitial resistance. In normal skin the interstitial pressure, as estimated by measurements of the interstitial resistance, is low, slightly less, on the average, than 1.7 cm. of water in the skin of the mouse, and less than 3.1 cm. of water in human skin. It remains unchanged in states of active hyperemia. In edematous skin the interstitial pressure can be directly measured by determination of the edema fluid pressure. It has been compared with determinations of the interstitial resistance and found to be only 0.5 cm. of water lower in both the mouse and man. Under the conditions of our experiments, in skin rendered slowly edematous by the introduction of irritant chemicals or their topical application, little rise in pressure took place. On the other hand, in rapidly forming edema of the skin the edema fluid pressure and the intradermal interstitial resistance rose and became great enough to hinder materially the further escape of fluid from the blood vessels. The edema fluid pressure rose in proportion to the rapidity with which the edema formed. When a rapidly formed edema subsided, the edema fluid pressure and interstitial resistance fell, but if inflammation and induration followed later, the interstitial resistance became high again. As these conditions subsided the interstitial resistance fell, at times to normal levels, even in the presence of edema. In mouse skin injured by squeezing according to a standard procedure, with result in pronounced edema, the intradermal interstitial resistance rose within a few hours to levels of 10 to 15 cm. of water. In those instances in which the injury progressed to induration, the interstitial resistance rose to such high levels that it seemed impossible that fluid could continue to escape from the capillaries. Such a state of affairs may be of great importance in determining whether necrosis follows trauma.

The sheath of omentum that forms about a long rubber tube left within the peritoneal cavity provides so efficient a barrier to ascending infection that such a tube can be employed for the permanent drainage of the common duct, whereas a short tube will regularly come away after a few days. A method is described whereby the total bile of a considerable number of dogs has been obtained in a sterile state during periods up to 3 months. Save for some anemia, the animals remained in good health throughout. The method has obvious uses in connection with the collection of other secretions.

Methods have been devised to bring microscopic amounts of fluid into contact with cutaneous connective tissue, under pressure or without pressure, in such a manner that it enters neither blood capillaries nor lymphatics directly. The take-up of fluid brought into contact with the tissues in this way has been measured and its characteristics studied. Elaborate control tests, here described and discussed, have indicated the possible errors in the employment of these methods. Locke's or Tyrode's solutions brought into contact with the cutaneous tissues, by the method described and at atmospheric pressure, pass into the tissue intermittently. Forced into the skin by pressures of 1.0 to 2.0 cm. of water the take-up is still intermittent in character. From this it follows that either the absorption of interstitial fluid from localized regions is periodic or the movements of interstitial fluid are influenced by intermittent physiological changes.

The tubules of the mammalian kidney, vitally stained with erythrolitmin, show a significant color pattern, the cells of certain regions appearing bright blue and others brilliant red. The dye is segregated within the cytoplasm, staining fine granules diffusely. Under normal circumstances of renal function erythrolitmin is stored in the lining cells of the glomerulus and the epithelial cells of the proximal convoluted tubules in the blue, alkaline form. In the cells of the final portion of the distal convoluted tubules the dye is red. Between these regions a narrow transitional zone can be found, at times above, at times below the loop of Henle, in which the erythrolitmin-stained granules in the tubule cells are violet, or red and blue within the same cell. Alterations in the relative reaction of certain regions of the tubules, as disclosed by the color of the dye within the cells, can be induced by means that alter the reaction of the urine. When acid urine is being secreted there is a change from relative alkalinity to acidity high up in the tubular canal, in the proximal convoluted tubules. During the secretion of markedly alkaline urine the intracellular granules appear blue most of the way down the tubular canal, even to the first portion of the distal convoluted tubules. When the urine is neutral the cells above the descending limb of the loop of Henle are alkaline to erythrolitmin, and those below this point appear acid. The granules within the endothelial cells of the glomerulus and the epithelial cells of the first portion of the proximal convoluted tubules are always alkaline to erythrolitmin; while those within the cells of the final portion of the distal convoluted tubules are always acid regardless of the reaction of the urine. Only after complete suppression of urine as result of massive doses of acid do granules of the sort first mentioned manifest the color indicative of acidity. The interpretation of the findings waits upon further work.

We have devised methods for the separation and isolation of the important indicator constituents of litmus, azolitmin, and erythrolitmin, with a view to employing them as vital stains. Analysis of the color intensities of these dyes shows slight differences in them, azolitmin being the weaker pigment, weight for weight. Study of a third coloring matter, erythrolein, which exists in litmus has shown it to be an unsatisfactory indicator, and toxic for animals. Analyses with the spectrophotometer of the absorption of light by erythrolitmin and azolitmin, prepared by our methods, and tested over a wide acid-alkali range, show them to be pure substances, comparable in this respect with synthetic indicators. The errors in the interpretation of the indicator phenomena on vital staining, which are incident to changes in the concentration of the dyes, are so slight as to be negligible. The salt and protein errors on the other hand are large. The factors responsible for the Donnan equilibrium fail to influence the distribution of the indicators between fluid and gelatin. Erythrolein was found useless when employed for vital staining, and azolitmin proved unsatisfactory since it colors poorly and is toxic. But erythrolitmin can be used to great advantage. It is readily absorbed, and in non-toxic doses stains intensely. The range of pH at which it changes from red to blue fits it for the demonstration of changes in the reaction of living tissues. By reason, however, of the salt and protein errors to which it is liable, the pH cannot be accurately ascertained. Intravital staining with erythrolitmin yields results similar to those following injection of purified "whole litmus."

The pulsation of blood vessels in the ear of the rabbit greatly increases the rate of the spread of dye introduced into the subcutaneous tissue. The appearance of edema in tissues perfused at a constant pressure leads to very little increase in the rate of dye spread. By contrast, a rapid interstitial spread of dye occurs in tissues becoming edematous while perfused with a pulsatile flow of blood. The significance of these facts is discussed.

The bile ducts from three-quarters of the liver substance in dogs and monkeys can be obstructed without any clinical evidence developing of pigment or cholate accumulation in the organism. And in the dog nineteen-twentieths of the liver substance can be placed in stasis without the occurrence of tissue icterus such as regularly follows total obstruction in this animal. There is no reason to suppose that this will not be found true in the monkey as well. Always a local obstruction results sooner or later in atrophy of the affected tissue with compensatory hypertrophy elsewhere. Thus as time passes the derangement of function produced by the sudden stasis is progressively lessened. The plasma of the dog and monkey, unlike that of man, is normally free from bilirubin, and this pigment so readily escapes from the blood into the urine that bilirubinuria is often to be found in the dog in the absence of bilirubinemia, while the latter is never met with alone in either animal. It follows that in both species the renal threshold for bilirubin is much lower than in man,—if indeed one can be said to exist at all. The amount of biliary obstruction required to produce jaundice in human beings is probably as great as in the experimental animals with which we have dealt. The clinical jaundice encountered in association with local liver lesions should be viewed not as the result of local bile resorption, but as due to a general injury to the hepatic parenchyma or ducts, or to blood destruction.

It has been known definitely since the time of Koch that certain essential oils have considerable disinfectant value. It has been assumed since his time also that substances which have disinfectant value when in watery solution are largely rendered inactive by solution in oil. This assumption arose as a wide generalization from what was actually observed by Koch in only one case, namely, that of phenol in vegetable oils, such as olive oil or cotton seed oil. In this case the validity of the observation is undoubted, but there have been a very few practitioners who have believed that in spite of the test tube indications the best way to apply phenol as a wound disinfectant is in solution in vegetable oil. The recent application by Dakin7 and his associates, of solutions of a water-insoluble oil-soluble substance (dichloramin-T) to the treatment of wounds indicates that from the practical point of view the question of the usefulness of disinfectants in oil is by no means closed. As a consequence of these general considerations, I undertook to study these questions anew. The recognized methods of determining quantitatively the antiseptic value of substances presuppose water solubility. The immiscibility of water and oil prevents the use of broth culture (or the suspension of bacteria in salt solution) in the testing of an oily substance, or of material which has been in contact with an oil. It is doubtless for this reason the physical difficulty of testing by standard methods that the many new antiseptic oils and oil-soluble antiseptics are accepted solely on the basis of clinical experience and not on that of known germicidal power. The value of such oils has not up to the present been expressed in terms of a known standard, comparable to the "phenol coefficient" of water-soluble antiseptics. Face to face with the necessity of deciding accurately the antiseptic power of a large number of oils the need for a preliminary investiga

Fat combustion is carried on adequately in rabbits deprived of the liver or brought into a condition of extreme liver insufficiency. Even 24 hours after hepatectomy fat combustion goes on as well as in the normal animal. Evidently the liver plays no essential part in the breaking down of fat.

With the aid of solutions of vital dyes the lymphatic capillaries in the ear of the mouse have been studied during the period of immediate reaction to injuries of various sorts and during the period of repair. The behavior of lymphatics severed by incision differs greatly from that of the small blood vessels. Instead of closing they sometimes remain open for as long as 48 hours. Materials introduced into the wound pass directly into the lymphatics through their gaping ends, a fact which will explain why infection from incisions is predominantly along the lymphatics. All around an injury the lymphatics are rendered abnormally permeable. So, too, are the blood vessels, a fact well recognized in the past. Twenty-four to 48 hours later, at a time when the blood vessels in the edematous tissue surrounding the injured area are still much more permeable than normal, the draining lymphatics allow far less to escape than usual. The possible reasons for this have been discussed. The lymphatics participate in the removal of fluid from the edematous tissue. As repair after injury takes place severed lymphatics may reunite when as yet there are no functioning blood vessels. Later an active hyperplasia of the lymphatic channels occurs, an extraordinarily abundant plexus of minute lymph capillaries budding into the area under repair.

In a previous paper the point has been brought out that the influence of the gall bladder upon the bile differs entirely from that of the ducts, the one organ acting to concentrate the secretion markedly and the other to dilute it slightly. The question arises, in species lacking a gall bladder, whether the concentrating function of this organ will be found lodged in the ducts. To test the point, observations have been made upon the mouse and rat, since these animals though so nearly related have, the mouse, a gall bladder and the rat, none. The normal bile was first studied. Both animals were found to secrete larger quantities than do cats and dogs, but less than the guinea pig and rabbit. Methods were worked out for the quantitation of the pigment which was used as the index to changes in concentration. Bladder bile of the mouse was regularly found to be more concentrated than that collected from the common duct of the same animal. The bile collecting during stasis regularly showed a great increase in pigment content, such as in other species is brought about by the action of the gall bladder. In the rat, on the other hand, stasis bile never became more concentrated in pigment than the normal. The gall bladder, then, is not only absent from the rat in form, but in one at least of its important functions. That its other obvious function—that of a reservoir—cannot be assumed in the rat by the ducts would seem to be indicated, not only by the small size of these channels, but by the recent observation of Mann that the tonus of the sphincter of Oddi is almost negligible in the rat, in contradistinction to animals which possess a gall bladder. It is an interesting fact that the bile of the rat, which as has been said, undergoes no condensation of bulk after leaving the liver, contains on the average eight times as much pigment as does the liver bile of the mouse which is submitted to concentration. Whether it is correspondingly strong in substances useful for digestion, and therefore ab initio requires no concentration, is a matter upon which little can be said at present. However, in this connection the fact that the bulk of bile secreted per gram of liver weight is identical in both animals may be significant. Although this output is the same, the mouse liver when compared with the body weight (1 to 14.6) is relatively larger than that of the rat (1 to 21.7), so that the mouse secretes somewhat more bile per 100 gm. of body weight. This bile as it comes from the liver is but one-eighth as strong at least in pigment as rat bile, but the concentrating activity of the gall bladder is so great that the products yielded to the intestine may become not dissimilar.

Following single intravenous injections of a foreign protein antigen, bovine γ-globulin, into mice and rabbits, antigenic material persisting in the liver could be detected for several weeks. Ground liver tissue—taken from the mice and rabbits, just mentioned, either 4 or 6 weeks after injecting the antigen—when transferred repeatedly, at 2 or 3 day intervals, to the peritoneal cavities of normal, or unilaterally adrenalectomized, recipient mice, rendered the recipients sensitive to active anaphylaxis when they were challenged after a suitable interval by intravenous injections of the original antigen. The work throws some light on the state of the antigenic material that persists for 4 to 6 weeks in the livers of the donor animals. Obviously it is sufficiently unchanged, at least in its reactive groups, to engender in the recipient mice antibodies capable of reacting with the original antigen.

Methods were developed for a study of the relations existing between viruses and living cells. It was found that vaccinia and the virus causing the infectious fibroma of rabbits (Shope) rapidly become fixed upon tissue cells freed as individuals and submitted to virus in suspension. This happens whether the cells are alive or have been killed with heat or ultraviolet light. The virus does not come away during agitation of the cells with Tyrode solution and repeated washings with large amounts of it. The exposure to neutralizing antisera of cells carrying virus fails to affect this latter significantly if the cells are alive, whereas if they are dead the activity of the virus is nullified. Cells freed as individuals from tissue cultures of vaccinia and the Shope tumor carry these viruses in abundance through repeated washings, and, if living, protect them from the influence of a neutralizing serum, whereas killed cells exert no such protection. The findings would appear to throw light on the way in which viruses gain a foothold in the host; and they suggest reasons for the persistence of some viruses in recovered animals and for the unsatisfactory results of serum treatment instituted during the course of virus diseases. The virus causing the Shope fibroma has been successfully maintained in cultures of the growth. It is closely associated with the cells, almost none being present in the culture fluid. Certain of its other attributes have been determined. Vaccinia greatly damages the cells of cultures of rabbit embryo in which it is under propagation.

A technic is described for the demonstration of lymphatic capillaries in living skin and for their study. By means of vital dyes injected intradermally these vessels can be rendered plainly visible. They form an extraordinarily abundant anastomotic web. The least scratch, one which does not penetrate through the epidermis, gives rise to such conditions that lymphatic absorption readily takes place from the abraded surface; and so close-meshed is the lymphatic web that an intradermal injection with even the finest hypodermic needle tears some of the constituent vessels open with result that they undergo direct injection. In many individuals much of the fluid introduced at an ordinary intradermal injection, like that made in the clinic, spreads through the superficial lymphatic network, whereas in others it tends to enter the deeper lymphatics at once, the difference being due to merely physical factors determined by skin texture. Normal flow along the skin lymphatics is rapid even when the body is at rest, dye introduced into the skin of the resting forearm reaching the axilla within a few minutes. The observations make plain the fact that every intradermal injection is an intralymphatic one, often preponderantly such, while furthermore every local injection into the skin becomes within a few minutes a general one, so rapidly is the introduced material transported to the blood. The normal permeability of the skin lymphatics of man is approximately the same as that of the mouse. Tests indicate that in both instances the lymphatic wall behaves like a semipermeable membrane. The permeability of the human lymphatic wall like that of the mouse is subject to rapid and great changes. A stroke on the skin with a blunt instrument to produce a wheal, causes the lymphatic capillaries to become so permeable temporarily that dyes pass through their walls as if practically no barrier existed, instead of being held back for a greater or less period. Slight inflammation due to heat, ultraviolet light or bacterial products has a similar effect. So, too, has histamine. When fluid pours rapidly into the tissue from the blood, as when a wheal is formed, the lymphatics are compressed and their efficiency as drainage channels is interfered with. These facts are briefly discussed in their bearing upon skin phenomena in general. The lymphatics cannot be disregarded in considering such phenomena, in which it is plain that they have a large share.

The gall bladder and ducts exert opposite influences upon the bile. The ducts fail to concentrate and thicken it with mucus as the bladder does, but dilute it slightly with a thin secretion of their own that is colorless and devoid of cholates even when the organism is heavily jaundiced. The fluid may readily be collected into a rubber bag connected with an isolated duct segment. It continues to be formed against a considerable pressure, and, in the dog, is slightly alkaline to litmus, clear, almost watery, practically devoid of cholesterol, and of low specific gravity to judge from the one specimen tested. In obstructed ducts separated from the gall bladder, or connecting with one so changed pathologically that the concentrating faculty has been lost, such fluid gradually replaces the small amount of bile originally pent up. It is the so called "white bile" of surgeons. When obstructed ducts connect with an approximately normal gall bladder the stasis fluid is entirely different, owing to the bladder activity. At first there accumulates in quantity a true bile much inspissated by loss of fluid through the bladder wall, darkened by a change in the pigment, and progressively thickened with bladder mucus. As time passes duct secretion mingles with the tarry accumulation and very gradually replaces it. The inspissation of the bile, as indicated by the pigment content, is at its greatest after only a day or two of stasis. The differing influences of the ducts and bladder upon the bile must obviously have much to do with the site of origin of calculi and their clinical consequences. The concentrating activity of the bladder cannot but be a potent element in the formation of stones. We have discussed these matters at some length. Intermittent biliary stasis is admittedly the principal predisposing cause of cholelithiasis; and the stasis is to be thought of as effective, in many instances at least, through the excessive biliary inspissation for which it gives opportunity. In this way a normal gall bladder can become, merely through functional activity, a menace to the organism. In patients with the tendency to stones frequent feedings may lessen the danger of their formation.

The effects of a single injection of a bacterial endotoxin on the cellular changes of a primary immune response to a standard dose of sheep red blood cells were studied in the spleens and mesenteric lymph nodes of mice. Daily histological comparisons of these organs in mice, injected with endotoxin, or with antigen, or both, showed that endotoxin given simultaneously with sheep red blood cells, as antigen, significantly enhanced all of the cellular changes that appear in the mesenteric lymph nodes and spleens of mice that form antibody when that antigen is given alone. First, in the white pulp of the spleens and cortical regions of the nodes, there appeared an early and excessive proliferation of the large pyroninophilic cells which seems to be responsible for the earliest formation of antibody, as judged by this work and that of others cited in the body of the paper. Polymorphonuclear cells invaded the spleens of these animals early after simultaneous challenge with antigen and endotoxin, and in far greater numbers than have ever been seen in mice given the same antigen without endotoxin. "Activated" germinal centers formed in the lymphoid tissue either 1 day before the appearance of antibody in the blood stream or on the same day, and they became larger than in the mice given antigen only. On the other hand, these specific and characteristic cellular changes failed to appear in mice prevented from forming any antibody at all by injections of endotoxin given 2 days before the antigenic challenge. These findings are discussed in the light provided by data from recent reports of others as well as in the light of the accompanying paper (1) which demonstrated not only the enhancement of antibody formation following simultaneous injections of antigen and endotoxin, as already known, but a totally unexpected, complete suppression of its formation when endotoxin was given 2 days before antigen.

By a method of permanent intubation the entire output of bile from dogs has been obtained in a sterile state over long periods of time, and studied quantitatively. The secretion of the first few days after the initial operation is scanty, contains more pigment than that secreted later, and is sometimes so viscid as to cause temporary obstruction within the collecting tube. The small amount and thick consistency of the fluid are referable to direct injury of the liver and ducts, while the abundant bilirubin is derived, in part at least, from the hemoglobin of extravasated blood. In some animals in which there was accidental mercuric chloride poisoning at the time of operation a suppression of the bile followed which, in one instance, was complete during the 48 hours before death. In a dog developing mixed infection of the biliary tract, an almost colorless fluid, glairy as white of egg, gradually took the place of the bile. Not until a week or 10 days after operation does the bile acquire the character which it maintains later. The quantity of this later bile, as measured in successive 24 hour specimens, is greatly less than that recorded by previous investigators, a difference attributable to disturbing influences inevitable to the method of collection they employed. By our method, it averaged from 3.5 to 9.5 cc. per kilo of dog in 24 hours, though transient variations from 1 cc. up to 14 cc. were encountered. Some dogs give consistently far more bile than others. The frequently recorded effects of fasting to lessen the rate of secretion, and of a meat diet to increase it more than a carbohydrate one, were noted often. Contrary to expectation, vigorous exercise does not act to increase the quantity of the bile. During hot weather this may sink greatly, although the animal remains in good condition; and during intercurrent diseases unassociated with jaundice, the flow may almost cease. One of the best of cholagogues, bile by mouth, fails of effect under such circumstances. It acts best when the animal is healthy, the weather not oppressive, and the food intake abundant. The bilirubin output, after the immediate effects of the operation have worn off, remains nearly constant from day to day, though often exhibiting slow, wave-like variations, each extending over a week or more. These slow changes are synchronous with similar alterations in the hemoglobin percentage of the circulating blood. A recognition of them is important to studies of the bilirubin yield. A mild anemia, absent in controls maintained under like conditions, regularly develops in unexercised dogs completely deprived of bile, despite their apparent health, and as it does so the bilirubin output falls off. Prior to the development of the anemia the output averages about 7.5 mg. per kilo in 24 hours, a yield which closely corresponds with that recorded by previous workers. Vigorous exercise of animals previously sedentary causes an increased pigment output presumably as result of increased blood destruction. Since the daily pigment output is approximately constant, whereas the fluid quantity undergoes frequent and great changes, it follows that the pigment concentration must vary inversely as the fluid quantity, and vary greatly. It does both. No matter how large or small the 24 hour specimen of bile may be within "normal" limits, therein will be found the customary quota of pigment. When, under pathological conditions, the bile flow almost ceases, the pigment concentration becomes extremely great. A similar reciprocal relationship between concentration and fluid quantity would seem to hold for the mucinous element of the bile. Scanty biles are ropy as a rule, and copious ones are watery. Temporary obstruction was produced by clamping the outlet tube from the common duct, and the bile yield following upon its relief was studied. Secretion was for a time far more copious than usual, with a low pigment content per cubic centimeter, and tended to remain so until the accumulated pigment had been voided. As much extra bilirubin is put forth after 24 hours of obstruction as if the liver continued to manufacture the pigment without interruption during it. Obstruction for this length of time appears to cause no subjective disturbance in most instances, though bilirubinuria develops. Even this sign of bile retention may be lacking when obstruction endures but 12 hours. The physiological and clinical significance of these facts is briefly discussed.

Pronounced vascular changes occurring in the ears and claws of mice during anaphylactic shock are described. Practically at once after a foreign serum (pig, horse, or rabbit) enters the blood stream of sensitized animals both the arterial and venous vessels undergo marked, local or generalized constriction in the organs mentioned. Usually spasm of the vessel walls occurs simultaneously in the arteries and veins, but it may appear first in the arteries, or occasionally in the veins. When venous spasm precedes arterial spasm, the true capillaries become distended with cells; if the reverse order holds, the ears appear bloodless. There is no active constriction or dilatation of capillaries; the capillary behavior follows passively the changes in the large vessels. Peripheral vascular spasm occurs while the carotid blood pressure is high, but a few minutes later, while this still holds true, the ear vessels begin to relax and the circulation is resumed. Shortly afterwards the blood pressure falls to levels far below normal, but the vessels remain open. If the circulation of one ear is obstructed while anaphylactic shock is produced, no vascular spasm occurs in it. Release of the obstruction during the animal's recovery results in belated constriction of the blood vessels of this ear although by now the vessels in the other ear are dilated and the general systolic blood pressure is very low. The vascular reactions in the ears appear to be uninfluenced by the blood pressure in the large vessels, and they are not a response to nervous stimuli. They are local in origin. The vascular changes are often not clearly perceptible in the gross but are plainly to be seen under a low power of the microscope. They occur in some sensitized mice exhibiting no manifest signs of shock, differing only in degree from the changes taking place when shock is severe or fatal.

In dogs intubated for the collection of all of the bile, a marked falling off in the yield of bilirubin is regularly to be noted after operation, followed soon by an anemia of secondary character. Though, in the absence of complications, the anemia is mild, it persists despite the excellent general condition of the animal. Intercurrent changes in the hemoglobin percentage take place from time to time, and these are accompanied by very similar fluctuations in the bilirubin quantity. At first inspection the data strongly suggest that blood destruction finds accurate quantitative expression in the yield of bile pigment. But this is not the case. True, the destruction finds expression in terms of bile pigment and practically at once; and the data support the conception that bilirubin has no other sources besides the hemoglobin of destroyed blood. But our experiments show that the amount of it put forth during the development of an anemia from gradual blood destruction, either intercurrent or induced, is far below that derivable from the net quantity of hemoglobin disappearing from the circulation. The discrepancy is referable to a process of pigment conservation which varies in proportion to the body need. We have shown in a previous paper that bile pigment may on occasion beabsorbed from the intestinal tract. This fact and those reported in the present communication lead one to question the accepted view that bilirubin is mere waste material eliminated by way of the liver. The anemia which develops upon total loss of the bile is not improbably consequent on the pigment loss therewith. Current methods of computing the rate of normal and pathological blood destruction from the bilirubin (or urobilin) yield are unsound in principle and open to large error in practise. Yet there is no doubt that day-to-day variations in the output of bile pigment result from changes in the blood; and on occasion they provide enlightening evidence of the nature of hemic events.

The mounting gradient of permeability along the small vessels of the corium is essentially unaltered by active hyperemia produced by heat, cold, or light. Only when the vascular walls are so damaged that rapid leakage ensues, as shown by the development of edema, does the permeability of the capillary web as a whole approximate that of the venules. It is plain that the normal gradient of vascular permeability depends upon the integrity of the vessel wall. The method of experiment described can be utilized for a study of the functional changes which result in the lesions due to burning and freezing.

The development of a method for the collection of total bile from dogs in a sterile state and uninfluenced by the gall bladder, day after day for weeks, has rendered possible an accurate study of the influence of diet upon the cholesterol output of the secretion. When a diet rich in cholesterol is given the amount of the substance in the bile greatly increases. Not only this but the concentration per cc. is, in almost every instance, greater. An increase in the total food intake, by the addition to the ordinary ration of a bone mash diet containing only a slight additional amount of cholesterol (200 mg.) produces a similar, though lesser, increase. In the fasting dog, the cholesterol yield is greatly cut down. The increase in the cholesterol after food rich in the substance does not depend on the cholagogue action of this latter, though it is true that the concentration of cholesterol in the bile usually increases with the bile volume. Though fasting cuts down the cholesterol of bile, the concentration of the substance per cc. is greatly increased. On an ordinary diet the yield of it fluctuates abruptly and considerably from day to day. In general the rule holds that an animal eating largely puts out not only much more bile but much more cholesterol. The relation between bile quantity and cholesterol yield is anything but a fixed one, however. The cholesterol yield of the bile does not parallel that of bilirubin. The pigment output from day to day remains relatively constant as compared with that of cholesterol.

The bile coming at one time from different portions of the liver of the dog has nearly the same amount of pigment per cubic centimeter. With this determined we have studied the power of the gall bladder to concentrate bile directed to it, using as criterion the pigment strength of a sample collected throughout the period of experiment from a duct branch. The extent and rapidity of the concentration are alike remarkable. A gall bladder emptied at the beginning of one experiment and left to fill from the liver, concentrated the 49.8 cc. of bile reaching it in 22½ hours to 4.6 cc., that is to say reduced its bulk 10.8 times; while another bladder left distended with a bile of known constitution and receiving in addition fresh increments from the liver concentrated the secretion 8.9 times in 22 hours. A series of five emptied bladders concentrated the bile coming to them in about 24 hours on the average 7.1 times, or a little more than the 6.4 times of seven organs left full. The conditions in both cases were relatively unfavorable to the withdrawal of fluid from the bile because this takes place by osmosis and diffusion, with the ultimate Δ always that of the blood, and the secretion in our animals was notably rich in solids as an indirect result of the operation. The rapidity with which fluid is withdrawn through the wall of the bladder may be judged from some experiments in which a bag was connected with the tip of the organ by a large cannula. Merely in its passage through the bladder the bile was concentrated 2.3 to 4.8 times. The finding indicates a potential source of error in observations upon samples of bile obtained from fistulous channels of which the bladder forms a part. The bile ducts do not withdraw fluid from the secretion they convey but tend to dilute it, as we shall show in a companion paper. The restriction of the concentrating activity to the receptaculum chyli is good evidence that the latter has special significance for the organism. The nature of this significance is briefly discussed.

A technique has been developed for the demonstration of lymphatic capillaries in the ear of the mouse by means of vital dyes and for tests of their permeability under normal and pathological conditions. The lymphatics become visible as closed channels from which the dyes escape secondarily into the tissue. Some of them, cross-connections, with extremely narrow lumen, would seem ordinarily not to be utilized. There is active flow along the lymphatics of the mouse ear under ordinary circumstances. The movement of dye was always toward the main collecting system. The valves of the lymphatics as well as fluid flow prevented distal spread. There was in addition slow migration, apparently interstitial in character, but in the same general direction, of dots of color produced by the local injection of dye. The normal permeability of the lymphatics was studied with dyes of graded diffusibility. Their walls proved readily permeable for those highly diffusible pigments that the blood capillaries let through easily, but retained those that the latter retained. Finely particulate matter (India ink, "Hydrokollag"), they did not let pass. No gradient of permeability was observed to exist along them such as exists along the blood capillaries of certain organs. The observed phenomena of lymphatic permeability, like those of the permeability of the blood capillaries, can be explained on the assumption that the lymphatic wall behaves like a semipermeable membrane.

Minute amounts of Locke's or Tyrode's solution have been brought into contact with the interstitial connective tissue of the skin of the living mouse, at atmospheric pressure, in such a manner that the blood or lymphatic vessels are not entered directly. Under such circumstances these absorbable fluids enter the tissue spontaneously. Entrance is strikingly intermittent, not continuous, and so too when very slight pressures are brought to bear on the fluids (1). Hyperemia of the tissues, with accompanying dilatation of the blood vessels, increases the entrance of fluids at atmospheric pressure but it is still intermittent. By contrast, venous obstruction leads to intermittent backflow into the apparatus, but reflex hyperemia, following release of the obstruction, is attended by an increase of flow into the tissues in spite of the great reactive dilatation of vessels. The inflow is also intermittent. If the skin is deprived of circulation, fluid does not enter it at all at atmospheric pressure, though it moves in regularly and continuously if slight pressure is put upon it. Edema-forming fluids, described in the text, also enter in a continuous manner when forced into the skin of either living or dead animals. So too do serum and sperm oil. The findings indicate that the passage of interstitial fluid into the blood vessels may be intermittent under normal circumstances and its escape from them as well. The observed occurrence of intermittent flow in the blood vessels of several tissues (9, 15–25) will go far to account for the intermittent entrance of fluid into the skin.

The findings presented in this paper indicate that 60 gm. of liver tissue, taken from adult donor rabbits 2 weeks after injecting them with large amounts of bovine γ-globulin, and transferred to the peritoneal cavities of recipient rabbits prepared for a secondary reaction, contain enough of the antigen to induce the formation of detectable amounts of precipitin in the recipient rabbits. On the other hand, liver taken from donors 21 and 28 days after injecting them with the same antigen fails to bring about a similar effect. The findings outlined in the paper suggest that the antigen is degraded after the 2nd week to such an extent that it fails to engender the formation of complete antibody in the recipient animals, although previous work has shown (2–4) that the transfer of liver tissue from similar donor rabbits to mice renders the recipient mice anaphylactically sensitive to the antigen, bovine γ-globulin. The implications of the findings are discussed.

A rapid and simple method for the production of marked liver insufficiency in rabbits has been described. The necessary operation can be carried out by an unassisted operator in a few minutes. The method should further the study of liver physiology. The changes as concerns blood sugar, urea formation, and uric acid metabolism would appear to be the same in the rabbit suffering from hepatic insufficiency as in the dog.

Rabbits in which a chronic anemia of moderate grade is induced by repeated bleedings repair the hemoglobin loss much more rapidly when given subcutaneous injections of hemoglobin than when this is not the case. But the effect of the injections is not manifest for several weeks, during which many pale corpuscles are put out by the marrow; whence it follows that the introduced pigment does not find its way in quantity direct to the new-formed cells but must follow a more or less roundabout metabolic route, perhaps the same one as that of ordinary iron compounds destined for the blood. The rapid replacement of the circulating hemoglobin in the injected animals occurs chiefly through an increased production of corpuscular substance having the same color index as that found in uninjected, anemic controls. By color index in this connection is meant the relation of hemoglobin to the volume of the massed corpuscles. Late in the period of bleedings and hemoglobin injections the demand for stroma for the new-formed blood is far greater than in control animals that have been merely bled, yet the circulating corpuscles show no lessening in resistance to salt solution, such as might perhaps be expected were there a stroma lack. The hematopoietic tissue of the injected animals undergoes an extensive increase—a fact which speaks strongly for the view that the elements out of which stroma is formed are still abundant. The factor which determines the spread of red marrow during anemia is shown by our experiments to be the presence in the body of hemoglobin, or perhaps of its precursors, in excess of the amount which can be utilized by the marrow already existing. Numerous illustrations in support of the point can be adduced from human pathology. Two will suffice. The widespread "currant jelly" marrow of pernicious anemia is found in an organism rendered anemic but supplied with hemoglobin in excess; while the pale, restricted marrow of cases suffering from chronic anemia due to repeated hemorrhages is associated with depletion of the constituents necessary for pigment production.

A method has been devised to measure the spread of vital dyes in the skin of mice. Spread is greatly influenced by physiological and pathological changes which add fluid to the tissue or abstract it. Spread is greater in the quiet, living ear than in the ear of an animal just killed. It is equally considerable in the frankly edematous ears of living and dead animals, and not greater in either case than in normal, quiet tissues. During the early stages of edema formation on the other hand, dye spread is notably rapid. It is still greater in the ears of normal animals actively moving about, and is greatest in tissues subjected to very gently intermittent changes in external pressure. The significance of these findings is discussed.

The pressure in the cutaneous lymphatic capillaries of normal mice anesthetized with nembutal ranged between 0.0 and 2.7 cm. of water. Measurements of the interstitial pressure in the tissue immediately next the lymphatics showed that, in more than half the instances studied, there was a slight gradient of pressure from the tissues to the lymph. In nearly all the other instances the pressures inside and outside the lymphatic capillaries were approximately equal. In two cases in which lymph flow in the capillaries was rapid, the lymph pressure may have been negative. Under these circumstances there must have been a considerable gradient of pressure from the tissues to the lymph. In skin which was rapidly becoming, or had recently become, edematous as result of the application of xylol or of heat, the intralymphatic capillary pressure generally was increased, yet when compared with the pressure prevailing in the edema fluid outside of the capillaries it was usually found to be relatively much lower, at times by as much as 5.9 cm. of water. The findings indicate that a pressure gradient is an important factor in lymph formation under normal and pathological circumstances.

With the aim of determining the structural conditions which affect fluid movement in the cutaneous connective tissue of mice, various test fluids were brought into contact with it under conditions such that neither blood vessels nor lymphatics were directly entered. Locke's solution, mouse serum, and a mixture of Locke's solution with a dye which causes edema were all employed. At atmospheric pressure, Locke's solution entered the tissues intermittently. When subjected to very low pressures it continued to enter the skin intermittently and at approximately the same rate. At pressures above 4.5 cm. of water, however, the flow became continuous but it did not increase in rate significantly until pressures of about 8.5 cm. were employed. There was no relationship between the rate of flow and the pressure employed. At a pressure of about 8.5 cm. the resistance of the tissues seemed to give way abruptly as if the formed elements had been separated. This has been termed the "breaking point." After it had been reached each further increase of pressure produced a proportionately greater inflow. Under the conditions of our experiments, the dye-Locke"s solution and also the homologous serum failed to enter the tissues at atmospheric pressure. It was necessary to subject these fluids to pressure to force them into the skin at the same rate at which the Locke's solution entered it spontaneously. Under these circumstances the dye-Locke's solution and the serum entered the skin continuously, not intermittently like the plain Locke's solution. As the pressure was gradually raised, no significant increase of flow into the tissues occurred until a point was reached, on the average 8.5 cm. of water, at which fluid suddenly began to enter very rapidly. This point, the "breaking point" already mentioned, was reached at the same pressure irrespective of the character of the fluid employed, showing that the phenomenon was produced by the fluid bulk. Once it had been attained, further increases in pressure caused proportionately greater inflow of fluid. The circulation had nothing to do with the phenomenon, for it occurred in the skin of dead mice." The findings indicate that under normal circumstances the movement of fluid in the interstitial tissue does not take place as though in preexisting channels. The experiments confirm previous observations from this laboratory (13, 14) that in normal skin tissue the state of affairs is such that fluid cannot flow freely. However, when fluid is introduced into the skin under pressure spaces are forcibly opened up. Inflammatory edema in the skin changed the phenomena of fluid entrance into it under pressure. The reason is that there then occurred a separation of the formed elements and the interstitial fluid moved as in preformed channels. Even when very low pressures were employed (3.0 to 7.0 cm. of water), there appeared usually a linear relationship between the pressure and the rate of flow.

The escape of a vital dye from the lymphatics of the ears of living mice and its subsequent movement through normal and pathological connective tissue have been observed at high magnification. The dye first appears outside such channels as bristly, wavy lines of color, which can be bent and twisted by pressure with a micro probe and spring back to their original positions when the pressure is removed, as if the dye were fixed upon or between some tissue elements. Our findings indicate that this is the case, that the bristly lines of color are formed by dye moving between connective tissue fibers or along them. With the onset of mild edema, such as the dye induces secondarily, the bristles disappear, the coloration becoming diffuse and freely movable with the micro probe. When edema is induced before dye is introduced into the lymphatics, the character of its escape is wholly different. It first appears as a colored cloud, freely movable in the edema fluid, the manner of its passage into the tissues being completely changed. In the ears of mice partly dehydrated by bleeding, or in those of dead animals, the bristly or wavy lines were more evident than in normal individuals. It was plain that dehydration did not change the mode of transportation of the dye through the tissue but merely emphasized some of the characteristics of its passage. In animals injected intravenously with large amounts of physiological saline, with result in the presence of more tissue fluid than usual, the colored bristles were seldom seen. It is plain that connective tissue fibers serve indirectly as pathways for the transport of substances of large molecule. We have not been able by the dye method to demonstrate the presence of any free fluid in the normal tissues of the mouse ear.

An antiviral principle is elaborated within the regional lymph nodes draining skin into which vaccinia is injected. The immunity conferred by clinical Jennerian vaccination may be largely of lymph node origin.

Vital dyes injected intradermally enter lymphatic capillaries directly, rendering them visible, and appear later in the draining lymphatic trunks as colored streamers. The method enables one to perceive the state of the lymphatic channels and the rate of lymph flow within them. It yields consistent results when tested under physiological conditions known to increase or decrease lymph flow. In the horizontally placed normal limb at rest there is slight lymph flow. In a normal leg or arm hanging downward lymph flow ceases although fluid in the limb increases. When a previously dependent arm is raised above the head, or when the foot of a seated subject is propped on a table, lymph flow in the raised limb becomes active. It ceases in the skin of an arm subjected to partial obstruction of the veins by pressure from without, but very active lymph flow appears during the reactive hyperemia which follows upon the release of venous obstruction. It is still greater following release of total circulatory obstruction, and seems to be the same whether or not the limb has previously been engorged with blood. In the ischemic patches which appear in the skin of a limb during total circulatory obstruction (Bier's spots) the lymphatic capillaries are definitely and considerably constricted, whereas they are slightly dilated in the purple, congested regions of the skin round about. On release of obstruction there occurs a strikingly rapid, equal lymphatic drainage from both regions. The significance of all the findings is discussed. When dye is injected intradermally and the skin sucked, much of the foreign material is driven into the lymphatics draining the injected area.

After feeding a dog, forceful contractions of the gall bladder occur that are sufficient in strength to expel part of the contents of the viscus against a considerable pressure resistance. The pressure within the gall bladder of a healthy, unanesthetized dog fasted 24 to 48 hours is usually about equal to a column of bile 100 mm. high. After a few swallows of food there is a rapid increase in the pressure to more than 200 mm. with a gradual fall in it again, and repeated similar rises and falls occur thereafter. The gall bladder contractions responsible for these alterations are accompanied by a lessening in the resistance to the passage of bile to the intestine, a resistance which is maintained by the muscles at the lower end of the common duct. There would appear to be a reciprocal response on the part of the two structures to the one stimulus. The maximum pressure developing within the temporarily obstructed biliary tract in an animal with the gall bladder excluded about equals that of a column of bile slightly more than 300 mm. in height. The taking of food acts as a stimulus on the rate of bile secretion, but does not alter the maximum secretion pressure. When the gall bladder is connected with the duct system, obstruction does not lead until after some hours to the development of a pressure of more than 100 to 150 mm. within the biliary tract,—that is to say the pressure does not rise above the normal. Its failure to rise further is referable to the activity of the gall bladder to store and concentrate the bile as secreted. The physiological and clinical significance of these findings is discussed.

Methods have been developed whereby animals can be totally, partially, and intermittently deprived of their bile, without infection of this secretion or of the duct system. In all the instances considered the bile remained sterile throughout the period of the experiment. With this aid we have studied urobilin physiology. We have been able to show that the normal presence of urobilin in the bile and feces of dogs depends on the passage of bile pigment to the intestine, either through the normal channels, or by abnormal ones, as when it is fed by mouth. Complete loss of the bile from the body resulted in the total disappearance of urobilin and urobilinogen from the bile, feces, and urine. Rarely very faint traces remained in the feces, the origin of which has been discussed. Partial loss of the bile resulted in a corresponding reduction in the urobilin of the dejecta. Feedings of sterile urobilin-free dog bile to intubated dogs losing all of their bile and having no urobilin in it, or in feces or urine, were followed by the appearance of the pigment in the hepatic bile secreted shortly thereafter. When the feedings were stopped the urobilin soon disappeared. Total obstruction of the bile flow caused disappearance of the urobilin of the bile and stool. Later as the animals became heavily jaundiced the pigment appeared again in very small quantity in the feces. Autopsy at this time showed that the intestinal mucosa was deeply tinted with bilirubin some of which undoubtedly had passed into the lumen of the bowel and had there been changed to urobilin. Employment of the "altercursive intubation," by which an intermittent diversion of the bile stream of animals from the intestine to a collecting apparatus could be effected, showed that while bile pigment still reached the intestine urobilin was present in the bile secreted by the liver but that almost at once after the bile had been diverted from the gut urobilin disappeared from it. In relation to this finding it was noticed that in animals from which merely a small fraction of the bile was collected, that from a single liver lobe, while the greater portion reached the intestine there was most urobilin in the bile at times when most bilirubin was entering the intestine.

In hepatectomized rabbits a progressive decrease in blood fibrinogen occurs. Partial defibrination in the liverless rabbit invariably results in a progressive decrease in blood fibrinogen preceded by a temporary and slight rise. No evidence has been secured of fibrinogen regeneration in the absence of the liver. From this it follows that the liver is the essential source of fibrinogen and in all probability the sole one.

As previous papers from our laboratory have shown, there exists a well defined tendency for calcium carbonate to come out of solution in the normal liver bile of the dog, and for it to be deposited on certain nuclei not infrequent in the secretion under pathological circumstances. Gall stones that had arisen in this fashion were a frequent occurrence in the intubated animals we studied. The present paper is concerned with the reasons for the absence of such stones from dogs with an intact biliary tract. The solubility of calcium carbonate is known to be markedly affected by the reaction of the fluid in which it is contained. The normal liver bile, out of which it tends to precipitate, is alkaline, with an average pH of 8.20 but in the gall bladder where conditions might otherwise seem especially favorable to precipitation, the secretion undergoes a change toward the acid side, becoming on long sojourn there, strongly acid to litmus (pH 5.18 to 6.00). From bile as thus altered, no carbonate precipitation takes place, even when it becomes greatly concentrated as in fasting animals or after obstruction of the common duct. Furthermore, carbonate which has precipitated out of liver bile on standing dissolves again in it when the fluid is rendered slightly acid in vitro, or, in some cases merely neutral to litmus. There are several obvious reasons for the absence of carbonate stones from the normal ducts under ordinary conditions,—notably the motility of these latter, the flushing that they undergo from an intermittently quickened bile stream, and the cleansing and possibly antagonistic action of the secretion elaborated by the duct mucosa. In the fasting animal, one at least of these influences is almost done away with, the rate of bile flow is so greatly cut down; while furthermore the calcium concentration of the secretion undergoes a considerable increase. But pari passu with these changes there occurs one in the bile reaction, a diminution in alkalinity so great that the pH often approximates that of the neutral point for litmus. That this change is not a direct consequence of the increase in calcium, may be inferred from the findings with stasis bile, the calcium content and reaction of which were observed to vary independently, if in general in the same direction. These adjustments within the organism, some of which may be thought to exhibit an element of the purposeful, when considered with the test-tube experiments, strongly suggest that the reaction of the bile plays a critical part in determining the occurrence of carbonate stones, as furthermore that their absence from the normal gall bladder is a consequence of the changes in the bile reaction there occurring. The changes come about through a functional activity of the bladder. This being the case, one might suppose that the failure to act would be followed by a formation of carbonate stones. There is sufficient evidence available in the literature to indicate that this happens, in rabbits at least. It is important to know whether changes in the bile reaction play any part in determining the cholelithiasis of man. To determine the matter will require a large material. But this much we have shown, that carbonate spheroliths not infrequently serve in human beings as centers in a formation of secondary stones of carbonate and cholesterol, as further that cholesterol precipitation out of human bladder bile can be induced or prevented by slightly altering the reaction of the fluid toward the alkaline and acid sides, respectively. The possibility that cholelithiasis may be a consequence of sins of omission on the part of the biliary channels and reservoir deserves to be considered.