Franklin B. Mead

Inactivation of Histamine in Vivo

One of us with Gebauer-Fuelnegg 1 has previously reported on the appearance of a physiologically active substance in the blood and lymph of dogs during anaphylactic shock and on the basis of a pharmacological and chemical study 2 concluded that the active substance was apparently histamine. It was noted in this study that this active substance may be only temporarily detectable and it was concluded that it disappeared rapidly from the blood as it circulates. Various workers have noted the inactivation or detoxication of histamine in intact animals, organ perfusions, etc. (see Best and McHenry 3 , 4 for discussion and literature). None of these studies, however, provide the necessary data to enable us to compare the fate of intravenously injected histamine with that liberated during shock. We were interested, therefore, in inducing shock of varying degrees of severity by intravenous injection of histamine at different dosage levels into dogs and of studying the blood and thoracic duct lymph for its presence and disappearance and of correlating these findings with our observations in anaphylactic shock. Dogs were anesthetized with ether and sodium barbital and freshly prepared solutions of histamine acid phosphate (1-500) injected into the femoral vein. Samples of blood and thoracic duct lymph were collected at various time intervals thereafter. These samples were kept from clotting by means of heparin, the bloods in addition were centrifuged to obtain the plasma, and then assayed for their histamine activity by intravenous injection into cats (under ether or ether and barbital anesthesia and after the preliminary administration of atropine). The essential data of these experiments are outlined in Table I. The degree of sensitivity of the cats used was always such that samples of blood or lymph recorded as negative contained a concentration of histamine less than 1-200,000. It is therefore apparent that injected histamine disappears very rapidly from the blood stream.

Role of Histamine in Circulatory Effects of Rattlesnake Venom (Crotalin)

Essex and Markowitz 1 have made an extensive study of the physiologic action of crotalin and have pointed out a number of similarities to the action of histamine. Feldberg and Kellaway 2 have recently shown that crotalin has the property of liberating histamine into the perfusate when it is added to the per fusion fluid with which the lungs of cats or guinea pigs are being perfused. Their studies demonstrate a mechanism whereby the characteristic action of histamine can be superimposed upon the direct effects of the venom. The extent to which the added action of histamine influences the action of the venom in intact animals is not indicated by these experiments. In a series of studies on anaphylactic shock 3 and peptone shock 4 in the dog, we have been able to show that the degree of the vascular reaction is correlated with and dependent upon the amount of histamine liberated from the dogs tissues in these reactions. We were, therefore interested in determining whether histamine is liberated when venom is administered to the intact dog and whether the amount of histamine liberated is adequate to account for the degree of the vascular effect produced. Crotalin∗ (from Crotalus atrox) was injected intravenously into 5 anesthetized dogs, in doses of 0.5 to 1.3 mg. per kilo, and samples of blood and thoracic duct lymph were collected during the ensuing reaction and tested for histamine. All of the reactions were severe, 2 being fatal within 20 minutes. Histamine was found in the blood in 2 experiments and also in the lymph in one of these. The identification of histamine was based upon the presence in extracts made by Codes 5 method for the determination of histamine, of a physiologically active substance which lowered the blood pressure of the atropinized cat, contracted the guinea pig intestine, and was inactivated by diazotized sulfanilic acid.

Effect of Iron on Hemoglobin Regeneration in Gastrectomized Dogs

We have previously shown 1 that the gastrectomized dog has a markedly reduced capacity to regenerate hemoglobin when maintained at anemic levels on various diets. The average production capacity is approximately 10% of that of normal dogs and is around 0.2 gm. of hemoglobin per day on a meat diet. It was also noted that liver feeding or the administration of liver extract 343 (Lilly) is comparatively ineffectual in increasing the level of hemoglobin production. Liver extract 55 (Lilly), however, caused a definite increase in hemoglobin production. As this preparation contains added iron, the present studies were directed to determining the effect of the oral administration of iron. Incidentally, the character of the blood findings during the period of anemia and of iron therapy were followed. The same 4 dogs used in the previous study were used, the gastrectomy operation thus preceding the present experiments by from 2 to 3 years. The technique of study was similar to that used previously. The dogs were maintained at an anemic level (e. g., hemoglobin levels of 6 to 9 gm. per 100 cc.) by intermittent bleeding, the hemoglobin production for a given period being determined as the total hemoglobin removed by the bleedings plus or minus the gain or deficit in total circulating hemoglobin during the same time. This is the method developed by Whipple and his coworkers. The hemoglobin determinations were made with a Sahli-Hellige Haemometer and the blood volume determinations by the method of Keith and Rowntree as modified by Whipple. One dog (No. 12) was spontaneously anemic and was not subjected to bleedings. After several consecutive periods of standardization on a meat diet alone, iron in the form of Blauds pills (Pills of Ferrous Carbonate), ferric ammonium citrate, or saccharated iron oxide was administered orally and the hemoglobin production during the iron regime determined.

Relationship Between Peptone Shock and Anaphylactic Shock

We have recently reported that the intravenous injection into dogs of a solution of peptone produces shock indirectly by leading to the liberation of a vasodepressor substance, identified as histamine, from the tissues. 1 Thus the mechanism of peptone shock is apparently similar to that of anaphylactic shock. 2 Biedl and Kraus 3 pointed out the remarkable similarities between these conditions. They also reported that peptone shock desensitizes an animal to anaphylactic shock and vice versa. They concluded that the fundamental mechanism concerned in these reactions was identical. They interpreted the reactions as indicating that peptone contains some active substance, such as Popielskis vasodilatin, which can produce shock directly, and that in the anaphylactic experiment a mother substance is formed as the result of the sensitization which then discharges this active peptone constituent when the shocking injection of antigen is made. Subsequent investigators have had variable results in attempting to influence an anaphylactic reaction by the prior administration of peptone. Hill and Martin 4 in their review summarize the reports as follows: “In short, there is some, but not striking evidence that peptone may inhibit shock within certain defined limits.” In view of our demonstration that both peptone shock and anaphylactic shock are reactions that are brought about indirectly by the liberation of histamine from the tissues of the shocked animal, the relationship between these reactions has considerable significance. For example, the question whether desensitization is related to a depletion or exhaustion of the mobilizable store of histamine in the tissues is subject to direct approach by such a study. The present investigation, therefore, was undertaken in the effort to determine whether or not one reaction would desensitize to the alternative reaction.

Common Bile Duct Obstruction and Anaphylaxis

We have elsewhere reported that blockade of the reticulo-endothelial system with India ink or saccharated iron oxide does not materially modify anaphylactic shock in the dog. In the preceding paper we have shown that the hyperbilirubinemia resulting from injections of bilirubin leads to a retention of bromsulphalein. Klein and Levinson 1 and more recently Mills and Dragstedt 2 have reported studies indicating that reticulo-endothelial blockade causes a retention of bromsulphalein. Interpreting the dye retention from a hyperbilirubinemia as a form of physiological blockade of the reticulo-endothelial system it seemed worth while to repeat our anaphylactic studies on jaundiced animals. Theoretically at least, the blockade occurring in dogs with obstructive jaundice should be more complete than that produced with foreign materials as it is constant and progressive. Anaphylactic shock was induced in 8 horse serum sensitized dogs in which the common bile duct had been obstructed. In one animal the obstruction preceded the sensitization, in the remainder the operation was done after the sensitization and at varying intervals before the test or assaulting injection of serum. Dye retention tests and determinations of the plasma levels of bilirubin were made just prior to the assaulting dose. For this the animals were anesthetized with ether and sodium barbital and the degree of shock evaluated by the blood pressure records as previously described. 3 The essential data are shown in the accompanying table. Severe to fatal reactions occurred in all but one animal. This animal was in poor condition and the blood pressure was extremely low before the horse serum was injected. No additional effect was produced. In 2 animals the shock was somewhat slower in developing than is usually the case. There is therefore no evidence of any mitigating effect on anaphylactic shock. Actually the percentage of fatal reactions obtained is higher than that occurring in a large series of normal animals.