G. B. West

THE FORMATION OF HISTAMINE IN THE RAT

Factors affecting the determination of histidine decarboxylase activity in adult rat tissues have been studied. The optimal conditions vary from tissue to tissue, and the most potent sources of the enzyme are the pyloric part of the stomach, the liver and the duodenum, with less in the kidney. There is no relationship between the histidine decarboxylase activity of a rat tissue and the amount of histamine it contains.

Tissue mast cells and tissue amines.

HISTORICAL INTRODUCTION THE first description of tissue mast cells was made in 1877 during an investigation of the staining of fresh tissues by the new aniline dyes then being produced by the German chemical industry. Ehrlichl observed that some cells in the connective tissue of animals contained granules which changed the colour of toluidine blue and certain other dyes as staining proceeded. The numbers of such cells were greater when chronic inflammation or conditions characterised by increased local nutrition were present. For this reason, Ehrlich considered these cells to be overnourished connective tissue elements and called them “Mast cells” (Mastzellen = well-fed cells). In choosing the name of mast cell, he was also influenced by his finding of a high mast cell content in connective tissue subjected to lymph stasis ; as we now know, in the selective lymphatic blockage of elephantiasis, the hypertrophied connective tissue is packed with mast cells and a similar histological picture, though on a smaller scale, is seen in young well-vascularised keloids. Two years after his first discovery, Ehrlich2 found similar cells in the blood, but whereas the blood mast cells, or basophils, take origin in the bone marrow and with the other leucocytes enter the peripheral blood, the more common tissue mast cells are born, live and die in the connective tissue. The similarity in the two kinds of cell lies in their content of watersoluble cytoplasmic granules which have a strong affinity for basic dyes, several of which change colour as staining occurs (metachromasia). Besides discovering and naming the mast cells, Ehrlich also described their morphology, staining properties and distribution, but he then left it to others to elucidate the chemical nature of the granules. But, for the next 60 years, research on this subject remained almost entirely histological and then in 1937 Swedish workers solved one of the riddles of the metachromatic granules of the mast cell. Jorpes and his colleagues in Stockholm had for long been working on the powerful anticoagulant material first isolated from dog liver and hence called heparin. On finding that heparin stained metachromatically with toluidine blue, these workers3 searched the tissues for metachromatism as a possible clue to the site of formation of heparin. And so it was that Jorpes was able to show that there is a good correlation of the mast cell count of a particular tissue and the amount of heparin that can be extracted from it. At that time mast cells were thought to be perivascular in location so as to produce heparin which pours into the blood stream. But a later study of themovement of the perivascular mast cells revealed that they migrate from the blood

The histamine-heparin complex.

Some factors concerned in the formation in vitro of a complex between the base histamine and the acid heparin have been studied. More histamine is contained in the complex when precipitation is carried out in acid medium in the presence of adenosine triphosphate. Although the staining properties and morphological characters of the complex resemble those of natural mast cell granules, it has not been possible to release most of the histamine from the synthetic granules without dissolving them. 5‐Hydroxytryptamine, noradrenaline and adrenaline failed to form complexes with heparin and did not alter the formation of the complex of histamine and heparin.

Studies on 5-hydroxytryptamine and 5-hydroxytryptophan.

The 5‐HT content and the 5‐HTP decarboxylase activity of several tissues of seven mammalian species has been estimated. Whereas 5‐HT is concentrated in the spleen and gastrointestinal tract, the highest enzyme activity is found in the kidney, liver, gut and brain. In rats, treatment with reserpine or chlorpromazine lowers the 5‐HT content of many tissues and reduces the 5‐HTP decarboxylase activity of the kidney, but prolonged treatment with cortisone only lowers the 5‐HT content of the skin.

ABSENCE OF CAPILLARY PERMEABILITY RESPONSE IN RATS TO DEXTRAN AND EGG-WHITE

Intradermal dextran and ovomucoid fail to increase capillary permeability in rats resistant to intraperitoneal dextran although intradermal histamine, 5‐HT and compound 48/80 are as effective as in control rats. When the skin of control rats is depleted of its histamine, intradermal dextran and ovomucoid are first ineffective but later increase capillary permeability although the skin histamine remains low. Chronic treatment of control rats with intraperitoneal dextran (which only slightly reduces both the skin histamine and 5‐HT) prevents the local dextran and ovomucoid responses but does not affect those of compound 48/80, histamine and 5‐HT. It is concluded that intradermal dextran and ovomucoid increase capillary permeability in rats by a mechanism involving substances other than histamine and 5‐HT, and that this mechanism is absent in rats which do not respond to intraperitoneal dextran.

Paper chromatography of some tissue amines.

When solutions of adrenaline, noradrenaline, histamine, 5‐hydroxy‐tryptamine and related amines in trichloroacetic, trifluoroacetic or picric acid are chromatographed in various organic solvent systems, the active material divides into two areas. If a basic amino acid is included in the solution before chromatography, the separation becomes more complete. It is suggested that the second area consists of a loose complex between the amine and the corresponding acid. When solutions of the amines in hydrochloric, acetic or oxalic acid are chromatographed, the active material resides only in one area.

Capillary permeability responses to snake venoms

Russells viper venom is many times more toxic to mice on intravenous administration than is saw‐scaled viper venom, and yet on local intradermal injection the two venoms have equal capillary permeability effects in mice, rats, guinea‐pigs and rabbits. Their effects are completely prevented by drugs possessing both antihistamine and anti‐5‐ht properties. It is concluded that the increased capillary permeability induced by the venoms is largely mediated through the release of histamine and 5‐ht.

THE INFLUENCE OF DIET ON THE TOXICITY OF ACETYLSALICYLIC ACID.

The toxicity of acetylsalicylic acid in rats is greater when animals are fed a high carbohydrate diet than when they receive a high protein diet. Magnesium deficiency increases this difference, particularly in pregnant animals. The maximum B.P. human dose of acetylsalicylic acid is well tolerated by pregnant rats fed on the high protein diet but one quarter of this amount produced foetal resorption and deaths in rats fed on the high carbohydrate diet. Acetylsalicylic acid is not teratogenic under the conditions used and appears to be unlikely to be teratogenic in man since its dose‐toxicity curve is steep. It appears essential to report the composition of the diet in toxicity tests.

INDOLE DERIVATIVES IN TOMATOES

DURING systematic examination of edible fruits, the bananal and the tomat0~9~ have been found to contain indole derivatives. Whereas the banana contained much 5-hydroxytryptamine (5-HT), the predominant indole derivative in the tomato was shown to be tryptamine. This communication describes the initial work being carried out to trace the formation and assess the importance of these indole compounds in the life history of the tomato plant. As soon as the first flowers had formed on young tomato plants the different parts were dissected for extraction. The tomato fruit was collected at different stages of maturation, from green and about 1 in. in diameter till over-ripe and very soft. Each of these fruits was then dissected into the skin, the pulp and the pips. Extracts were made with acetone (1 g./5 ml.) for 24 hours. After reducing the extracts to a small volume, aliquots were either tested for 5-HT activity on the isolated uterus of the rat in oestrus or subjected to two-dimensional paper chromatography. The solvents were sodium chloride solution (8 per cent w/v), isopropanol : ammonia : water (20 : 1 : 2), and n-butanol : acetic acid : water (4: 1 : 5). The indoles were detected on the chromatograms with Ehrlich’s reagent. Duplicate spots were eluted and the eluates tested biologically for 5-HT activity. The concentrations of other indole derivatives were estimated visually by comparison with the colours produced by known amounts of standard substances. These were 5-HT creatinine sulphate, tryptamine hydrochloride (T), tryptophan (TP), 5hydroxytryptophan (5-HTP), 5-hydroxyindoleacetic acid (5-HIAA), and indoleacetic acid (IAA). All values shown are the means of three observations. Young tomato plants. No indole derivatives were detected in extracts of the roots of these plants. In the main stem, TP was found in a concentration of 2.5 pg./g. The concentration of 5-HT in this region was less than 0.01 pg./g. When the leaf stems were examined, the concentration of TP had not increased but that of 5-HT was 0.3 pg./g. In the leaves and leaflets 5-HT was present in an even higher concentration (0-5 pg./g.). Lastly, in the flowering tip, both TP (2-5 pg./g.) and 5-HT (0.3 pg./g.) were detected. These results are shown in Table I. T, 5-HTP, 5 H I M and IAA were not detected in any of the extracts obtained from the parts of the young tomato plants. Tomato fruit. When the green tomato fruit was examined, the predominant indole was T (1 pg./g.), and smaller amounts (0-2pg./g.) of both 5-HT and TP were also present. In the ripe fruit, the concentration of T

Drugs and Rat Pregnancy

SIR,-Whilst investigating the role of histamine in rat pregnancy, it became necessary to study the action of certain drugs in animals which had been mated. The results however may be of interest clinically as some of the drugs which are used in human pregnancy produced abortion in rats. No foetal abnormalities like those seen after thalidomide in humans and rabbits have so far been produced but this may only be because conditions for such malformations have not been achieved. Besides, experimental work on laboratory animals during the past 10 years has revealed that many chemical substances have the power when administered to the pregnant female of producing congenital malformations in the young (see Millen, 1962). We tried to discover why the rat foetus forms much histamine during the last third of gestation and why the maternal urinary excretion of free histamine increases more than 10-fold during this period. The uterine and placental histaminase activity increases to about the same extent at this time yet the maternal uterus becomes relatively insensitive to histamine. Thus, the hypothesis was made that the function of histamine formed by the rat foetus is to help control the blood flow through the placenta (Kameswaran, Pennefather and West, 1962).