Argyrophile and argentaffin reactions in individual granules of enterochromaffin cells of the guinea pig

Summary Successive staining of cells of the enterochromaffin system of the guinea pig by an argentaffin and an argyrophile method shows that, as in man and rabbit, they can be divided into (a) those in which all granules are apparently argentaffin, (b) those in which all granules are argyrophile but none are argentaffin, and (c) those in which some granules are argentaffin (as well as argyrophile) while others are purely argyrophile. The presence of the third type makes it evident that no hard and fast line of distinction can be drawn between the so called argentaffin and argyrophile cells.I am grateful to Mr. Mohan Lal Sharma for help with the photographs and to Mr. Anand Parkash for technical assistance.     

Further observations on the alleged presence of non-argyrophile argentaffin cells in the human gastro-intestinal tract

Summary The relationship between the argentaffin and argyrophile cells of the human gastro-intestinal tract has been studied by staining paraffin sections first by the Masson-Hamperl method and then by the Bodian method. The investigations confirms the author's earlier observation (Singh 1963) that all argentaffin cells of the human gastro-intestinal tract are also argyrophile and that there is no evidence to support the claim of Hellweg (1952) and of Hamperl (1952) regarding the presence of non-argyrophile argentaffin cells.     

Argyrophile and argentaffin reactions in individual granules of enterochromaffin cells of reserpine treated guinea pigs

Summary The individual granules of enterochromaffin cells of normal and reserpine treated guinea pigs have been studied by staining slides of the duodenum first by an argentaffin method and subsequently by an argyrophile method. Some argentaffin cells can be shown to contain not only argentaffin granules, but also granules that are purely argyrophile. The relative number of such argentaffin cells is greatly increased following administration of reserpine, as depletion of their 5-hydroxytryptamine content converts argentaffin granules into purely argyrophile ones. On the basis of this finding it is confirmed that the argyrophile granule is merely an argentaffin granule depleted of its 5-HT content, and that the argyrophile (nonargentaffin) and the argentaffin cells represent different phases of a secretory cycle.This investigation was supported by a grant from the Indian Council of Medical Research. I am grateful to Ms. Ciba of India Ltd. for making available reserpine (Serpasil) and solvent for reserpine. It is a pleasure to thank Mr. Anand Parkash for technical assistance and Mr. M. L. Sharma for the photographs.     

The relative proportion of pre-enterochromaffin (argyrophile) and enterochromaffin (argentaffin) cells in the gastro-intestinal tract of human foetuses

Summary The proportion of argyrophile cells that are argentaffin varies considerably over different parts of the gastro-intestinal tract of the human foetus. In the small intestine plots of the number of argentaffin cells against the number of argyrophile cells, at various cranio-caudal levels, reveal no correlation. The percentage of argyrophile cells that are argentaffin decreases in a caudal direction in the proximal part of the small intestine, but markedly increases in the distal part. In the large intestine there is a positive correlation between the numbers of argyrophile and argentaffin cells, but the percentage of argyrophile cells that are argentaffin also varies with cranio-caudal level. The pattern of this variation is not consistent in the various foetuses studied.     

Studies of the effects of aminopterin on the crypts of Lieberkühn in rats

Summary Studies have been made on the effects of an intramuscular injection of aminopterin on the crypts of Lieberkühn in rats. A decrease in the mitotic counts was accompanied by a rapid increase in the number of abnormal cells present in the epithelium of the crypt. Three hours after administration of the aminopterin, an almost complete absence of true metaphase chromosoms was found. By 24 hours, a partial return towards normal mitotic activity was observed but the number of abnormal cells present was still very high. It is suggested that the mitotic changes are in keeping with the conclusion of Grampa and Dustin (1952) of an arrest at interphase but that a secondary arrest at metaphase cannot be excluded.A morphological feature of some of the abnormal cells was the presence of a Feulgen positive granule in the cytoplasm, which by electron microscopy was also shown to contain many cytoplasmic constituents. It is suggested that material is lost from the nucleus and incorporated into a granule in the cytoplasm. A possible explanation of the purpose and function of the granule, as a means of disposing of unwanted or aberrant material, is put forward. Acknowledgements. I am grateful to Professor R. J. Brocklehurst for his continued interest and support of this work, and to the Stage II, B. Sc. students (1964) who counted the cells in many of the specimens as a laboratory exercise. My thanks are also due to Mr. J. Clements for technical assistance.     

The depletion of the granules of argyrophile cells of dog duodenum by reserpine

Summary The influence of reserpine on the argentaffin and non-argentaffin argyrophile cells of the duodenum has been studied in 8 dogs. In two dogs that were given 5 mg/kg of reserpine there was a complete disappearance of both types of cells. In another dog given 5 mg/kg and in dogs given 2.5 mg/kg and 1 mg/kg reserpine, the number of argentaffin cells was reduced to almost nil while the number of non-argentaffin argyrophile cells showed a statistically significant reduction. The degree of depletion of non-argentaffin argyrophile cells was proportional to the dose of reserpine given. The findings show that the non-argentaffin argyrophile cells are closely related to 5-hydroxytryptamine and their distinction from argentaffin cells is only a quantitative one.     

Electron microscope observations of some peripheral synapses in the sensory pathway of the sucker of Octopus vulgaris

Summary A synaptic axo-dendritic linkage is described between primary receptors lying in the epithelia of the sucker of Octopus and encapsulated nerve cells found near the rim of the sucker in the subepithelial connective tissue. These synapses are postulated to perform a drastic reduction of inputs between the primary receptors of the order of more than ten thousand and the subjacent encapsulated nerve cells of the order of some hundreds. The morphology of these cells as well as that of the synaptic structures are described from electron microscope studies. Aknowledgement. This work was done at University College London, while I was in receipt of a Medical Research Council grant.I am deeply indebted to Prof. J. Z. Young F. R. S. for support and criticism, and to Dr. E. G. Gray for advice and discussion. My thanks are due to Mr. A. Aldrich for the photographs.     

Changes in the fine structure of the human testicular interstitial cells after treatment with human gonadotrophins

Summary Study of the fine structure of the human interstitial cells after prolonged stimulation with human gonadotrophin reveals a striking increase in the quantity of the agranular endoplasmic reticulum. This is accompanied by an increase in the number of mitochondria which exhibit more extensive cristae, collections of intramitochondrial lipid and aggregations of electron-dense granular deposits. A rise is also evident in the number of lipofuscin pigment deposits and granular membrane-bounded bodies, both of which exhibit acid phosphatase activity. These changes after gonadotrophic stimulation are discussed in relation to steroid biosynthesis.In the pretreatment biopsies of these patients aged between 25–35 years, some interstitial cells contain intranuclear crystals which exhibit a hexagonal structure. The relationship of these intranuclear crystals to the cytoplasmic crystals of Reinke is discussed.The author is indebted to Dr. J. W. Johnstone and Dr. A. Long for the human material used in this study. Thanks are also due to Dr. H. P. Taft for helpful suggestions in the management of these patients, to Professor B. Hudson for the estimations of plasma testosterone and to Dr. J. B. Brown for the supply of human pituitary gonadotrophin and the estimations of urinary oestrogens. The technical help of Mr. T. Mezciems and the photographic assistance of Mr. J. S. Simmons F. R. P. S. and Miss S. Flett is gratefully acknowledged.     

Electron microscopic classification of amine-producing endocrine cells by selective staining of ultra-thin sections

Summary The argyrophil, argentaffin and chromaffin reactions were performed directly on ultra-thin sections for examination in the electron microscope. Glutaraldehyde fixation was appropriate for the argentaffin and chromaffin reactions; additional fixation with osmium tetroxide, however, caused impairment of these reactions. Fixation with formaldehyde, but not with glutaraldehyde, was adequate for the argyrophil reaction; post-fixation with osmium tetroxide did not affect this staining. At the light microscopic level the staining reactions were correlated with fluorescence histochemistry according to the method of Falck and Hillarp. The techniques described were used to study certain amine-producing endocrine cell systems: adrenal medullary cells and thyroid parafollicular cells of the mouse, gastric endocrine cells from the oxyntic gland area of the mouse, rat and rabbit. All these cells stained argyrophil. The adrenal medullary cells and one cell type in the oxyntic gland area of the rabbit were strongly argentaffin and chromaffin. The remainder of the cells were non-argentaffin and non-chromaffin but could be induced to give an argentaffin (and chromaffin) reaction after injection of the animals with l-3,4-dihydroxyphenylalanine or l-5-hydroxytryptophan, a treatment which is known to result in the accumulation of the highly reducing dopamine and 5-hydroxytryptamine, respectively, in these endocrine cells. Without exception the precipitates formed in all the staining reactions accumulated selectively over the secretory granules of the cells.The techniques described permit differential staining of consecutive ultra-thin sections for electron microscopic characterization of one and the same cell. They will provide information necessary for correlative studies of the stainable cells at the light and electron microscopic levels.     

Fine structure of argyrophil and argentaffin cells in the gastro-intestinal tract of the fowl

Summary The fine structure of granular cells in the proventriculus gizzard and intestine of the fowl has been described. The proventriculus and gizzard have only one type of granular cell, the pure argyrophil cell. In the intestine two cell types are shown. One corresponds to the argyrophil cell seen in the proventriculus and gizzard. The other, more common, is considered to be the argentaffin cell of the intestine. Structural differences between these two cell types confirm the known differences in histochemical reaction. Acknowledgement. I wish to thank Mr. R. N. C. Aitken, Department of Veterinary Histology and Embryology for supplying the birds used in this study.     

Hypothalamic control of the pars intermedia in Xenopus laevis tadpoles

Summary In Xenopus laevis tadpoles the relation between a paired nucleus of bio-amine producing neurons in the caudal hypothalamus and the pars intermedia of the hypophysis was studied.Treatment of the animals (stage 49 to 50 of Nieuwkoop and Faber's normal table) with reserpine caused aggregation of the skin melanophores within one hour, followed by redispersion five to six hours after the beginning of the experiment. This was at exactly the same time as the bio-amines in the caudal hypothalamus disappeared. However, the drug was ineffective if the nuclei had been removed. This indicates that reserpine acts via these nuclei and does not influence the skin melanophores directly.It was concluded that the initial aggregation of the melanophores may be the result of a reduced extrusion of MSH from the pars intermedia, caused by an increased output of a MIF by the bio-amine producing nuclei. The redispersion was explained by assuming that the bio-amines were depleted and the nuclei stopped with the extrusion of the MIF. This does not mean that the production of a MIF is the only function of the paired bio-amine producing nucleus in the caudal hypothalamus.The author thanks Prof. Dr. P. G. W. J. van Oordt for his helpful comments and criticism. Mr. J. H. I. J. M. ten Berge and Mr. E. W. A. Kamperdijk provided great assistance during the course of the experiments. Mr. H. van Kooten made the diagram and the photograph.     

Fine structure of cilia in rat cerebral cortex

Summary Cilia have been demonstrated on granular neurons and astroglial cells in the fascia dentata, a part of the hippocampal region, in the rat. Every granular cell seems to possess one cilium, which shows an 8+1 pattern in the greater part of its length. This 8+1 pattern is shown to result from the displacement of one peripheral doublet of a 9+0 cilium into the middle of the cilium. The neuronal cilia have a two-centriole basal organization, and fine rootlets radiate from the basal body proper into the cytoplasm. The possible function and significance of these cilia are discussed on the basis of earlier literature.This study was supported in part by Grant NB 02215 of The National Institute of Neurological Diseases and Blindness, U.S. Public Health Service, in part by The Norwegian Research Council for Science and the Humanities. This aid is gratefully acknowledged. I am greatly indebted to Dr. Th. Blackstad for encouragement and advice during this study, to Mrs. J. L. Vaaland, Mr. B. V. Johansen and Mr. E. Risnes for technical assistance, and to Dr. B. Afzelius for valuable discussions.     

The distribution of enterochromaffin cells in the human small intestine

Summary The distribution of argyrophile and argentaffin cells has been studied in the small intestine of five human adults. In proceeding cranio-caudally the characteristic feature of their distribution is the presence of eight to ten waves of rising and falling density. A progressive decrease in density of cells from duodenum to terminal ileum (described by previous workers) is not present.Re-examination of findings reported earlier in the small intestines of human foetuses shows that a predominant U shaped pattern of distribution is present in younger foetuses. This changes to the adult pattern by full term. The appearance of the adult pattern occurs earlier for argyrophile cells than for argentaffin cells.     

Studies by negative staining on the structure of collagen fibrils in normal and lathyritic rats

Summary Electron microscope studies on collagen from rat-tail tendon using the negative staining technique indicate the presence of fine filaments 15–30 Å in diameter within the fibres. The banding of the fibrils was only slightly affected by aminoacetylnitrile bisulphate, but an increased amount of fine fibrous material was present in preparations from experimental animals. It is suggested that this material represents a soluble form of collagen which is known to predominate after administration of the drug. Despite the fact that this would be a chemically abnormal collagen, its structure by electron microscopy corresponds remarkably well with the form suggested from X-ray diffraction and physico-chemical studies on normal soluble collagen.The author would like to thank Mr. J. Clements for technical assistance and Mr. R. Thomas (Dental School, Bristol) for discussion on the effects of aminoacetylnitriles in rats.     

Hypothalamic neurosecretory activity in relation to the reproductive cycle in the common striped skunk (Mephitis mephitis nigra; order carnivora)

Summary The histological appearance of certain cell groups in the anterior hypothalamus was studied during various phases of the reproductive cycle of the common striped skunk (Mephitis mephitis nigra), a carnivore having a very restricted period of sexual activity. Pronounced changes occur in the amount of stainable neurosecretory material in cells of the supraoptic, paraventricular and anterior hypothalamic nuclei. Material staining with aldehyde fuchsin is stored (or inhibited from being released) in these cells during the sexually quiescent period in both the female and male skunk. The time of approaching sexual activity is characterized by the first signs of release of neurosecretory material from these hypothalamic cells and the peak of estrus and rut coincides with a minimal content of the material.Dedicated to Professor Berta V. Scharrer in honor of her 60th birthday.This study was supported by U.S.P.H.S. Training Grant 5 T 1-GM 102 and Research Grant BN-00840, from the National Institutes of Health. This represents a portion of a dissertation, written under the guidance of the late Professor Ernst Scharrer, and submitted in partial fulfillment of the requirements for the degree of Doctor of Philosophy at Yeshiva University, June, 1965. Preliminary reports of this work were presented at the 78th annual session of the American Association of Anatomists, Miami, Florida, and at the VIIIth International Congress of Anatomists, Wiesbaden, Germany, (Hagedoorn, 1965a, b). I thank Dr. H. W. Deane and Dr. J. Osinchak for their critical reading of this manuscipt.     

The effect of metopirone and aldactone on the interrenal tissue of intact and pars distalis-ectomized common frogs Rana temporaria

Summary The effects of metopirone (SU-4885) and aldactone (SC-9420) on the histology and histochemistry of the interrenal tissue of male common frogs, Rana temporaria, have been studied. From the changes in nuclear and cellular size, mitotic rate, and lipid and cholesterol content it has been concluded that in intact animals the administration of metopirone as well as of aldactone resulted in an increase of the interrenal secretory activity. The effects were very similar to that of ACTH. Ablation of the pars distalis prevented the interrenal activation by metopirone. The effect of aldactone was markedly reduced in operated frogs, but some stimulation remained. It is suggested that both substances act by way of enhancing pituitary ACTH ouput. The dependence of the secretion of corticosteroids on pituitary ACTH is discussed.I wish to thank Dr. W. J. van Dongen for his cooperation and criticism and for enabling me to do this work in his laboratory. I am endebted to Prof. Dr. P. G. W. J. van Oordt for his valuable and constructive suggestions. Thanks are also due to Miss Mieke Roelofsen and Miss Ineke Wienen for their conscientious technical assistance.     

The effect of glycerination on the fibrillar flight muscles of belostomatid water-bugs

Summary The progressive effects of glycerol extraction on the myofibrils and membranous structures of insect flight muscle were examined. The myofibrils appear to be intact after extraction for several months. The mitochondria are completely disrupted within 3 days, but later the fragments come together to form mitochondrion-like bodies. The sarcoplasmic reticulum around the Z-lines is preserved for many months, but the T-system and dense bodies of the dyads disappear after about 7 days extraction.I am indebted to Professor J. W. S. Pringle, F.R.S. for his interest and helpful discussions throughout the course of this work. I should like to thank Miss Christine Williams for her technical assistance.     

Electron microscope studies on collagen

Summary The present paper describes the effect of intensive tryptic digestion of native tropocollagen (TC) macromolecules in solution. Contrary to earlier investigations it has been found that the trypsin treatment results in a fragmentation of the TC molecules. The addition of ATP to solutions exposed to the enzyme yields SLS fragments, 2250 Å in length. Comparison between these and normal SLS type aggregates shows that the scission occurs in a well-defined locus adjacent to the -1/2-line seen after positive staining. The significance of this finding is discussed.This study was supported by grant NB-02215-04 from the National Institute of Neurological Diseases and Blindness, Public Health Service, U.S.A. and a Student research fellowship from the Norwegian Research Council for Science and the Humanities. This aid is gratefully acknowledged.I am indebted to Mrs. J. Line Vaaland and Mr. B. V. Johansen for technical assistance.     

The ultrastructure of the test of the tadpole larva ofCiona intestinalis

Summary The test of the tadpole larva ofCiona intestinalis consists of an amorphous background substance in which are long 4–7 nm fibrils. There is a narrow outer dense region of the test where both the background material and the test fibrils are more concentrated and orientated. Fibrils of similar dimensions are found in the epithelial cells below the test, and also in cells associated with the outside surface of the test. No such fibrils are found in the cells located within the test substance. There are some regions where the external cell membrane of the epithelial cells is indistinct and in these regions the fibrils of the test are continuous with the fibrils within the epithelial cells. No large or small vesicles opening from the epithelial cells into the test have been seen.This evidence has been interpreted as support for the tunicization theory of test formation. It is suggested that the cells within the test are the source of the polyphenols that are necessary for quinone tanning that gives rigidity to the test.The fibril-containing cells outside the test probably add fibres to the cuticular region of the test.I wish to thank ProfessorJ. Z. Young, F.R.S. for much advice and encouragment, also Dr.R. Bellairs for the use of electron microscope facilities and Mr.R. Moss and Mrs.J. Hamilton for skillful technical assistance.     

Determination of temperature and heat production in insects

Summary Thermojunctions suitable for accurate measurement of the tp. of muscles in insects during activity and when exposed to the resulting air flow are described.A method of artificial heating of the body of a paralysed insect to reproduce the tp. curve of activity and measure the corresponding heat input is described.I am greatly indebted to Mr. Weis Fogh M. Sc. who ably assisted me in this work and introduced certain improvements in technique, and I am happy to offer it as a tribute to the experimental genius of professor Karl v. Frisch.