ne Str ee ey ee Pe : . rs ae a a ee > [Se %e ee ale ee > & ., a PF Pe 7, ee MOEA Me , rv ¥. STAY oF Pe tA Res 4 nd A aes 99) PP? 5 > Nes AA »- > a2 y , IF td yy nN ake Pa ad Loryy awe a SP ee Porpeaes , , ? y > vy ; ? wp AAD N AAD AD ARADO SP rary PNP yy? ary’ r> Ce % 7 . Se / we - oe st o.458 , - r ey ~~ . L4G SE ee a z e ‘ f Ane A . “ Ln, ttm ase fon, 4 RAAARAABZAAAA . % - Oe ew: — Ceara eae RATA SORES, . oe et ce SP ONAN Sth 2 eet > 4? a ~ » a ee Oe Z " 2a, 2u th fe IN Pe Te AN ae ON : 2 ‘! KL oath 1S IRAN AN NS OS . a LNA . “ RBRAA . » 7 a . ae ms tana. a7 ear yore > ? > A > \? Z a A Ces > A Me 44 Ser Rens 2. ae w ‘as Po? oe POLY > as POPE? xo » a apres ‘as a ee eS nm RAB ADO I. > PnP?! Sasi om? >> EEE Se Nasa sacd > >? 0 te te Le ANP PAR NM Ne ANAM Hy OS de eA NT oe 28 ns On am ay eB ANE PS eI Om A INA IS A EON AL ARLEN eA, DAN DN Be A Ae A OR RN NON tAARA pe Bees ae ie Soca 5 O€PLTOO TOEO g HONEY 1IOHM/TEIN i. i aie ut) beg ' INTERNATIONAL SERIES OF MONOGRAPHS ON PURE AND APPLIED BIOLOGY Division: ZOOLOGY GENERAL EpITor: G. A. KERKUT ' VOLUME 6 ANIMAL HORMONES— m COMPARATIVE. SURVEY Part I. Kinetic and Metabolic Hormones Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. Vol. _— WN khwWNeR SOOONI DRO No — FSO oN RN OTHER TITLES IN THE SERIES ON PURE AND APPLIED BIOLOGY ZOOLOGY DIVISION RaveN—An Outline of Developmental Physiology RaveN—Morphogenesis: The Analysis of Molluscan Development Savory—Instinctive Living Kerxut—ZIJmplications of Evolution TartTar—The Biology of Stentor Cor.tiss—The Ciliated Protozoa Grorce—The Brain as a Computer ARTHUR—Ticks and Disease RaveN—Oogenesis Mann—Leeches BIOCHEMISTRY DIVISION Pirt-Rivers and Tata—The Thyroid Hormones BusH—The Chromatography of Steroids ENcEL—Physical Properties of Steroid Hormones BOTANY DIVISION Bor—Grasses of Burma, Ceylon, India and Pakistan TURRILL (Ed.)—Vistas in Botany ScuuLtes—Native Orchids of Trinidad and Tobago CooKE—Cork and the Cork Tree MODERN TRENDS IN PHYSIOLOGICAL SCIENCES DIVISION FLORKIN— Unity and Diversity in Biochemistry BracHET—The Biochemistry of Development GEREBTZOFF—Cholinesterases BrouHa—Physiology in Industry Bacg and ALEXANDER—Fundamentals of Radiobiology FLORKIN (Ed.)—Aspects of the Origin of Life HOLLAENDER (Ed.)—Radiation Protection and Recovery Kayser—The Physiology of Natural Hibernation FRANGON—Progress in Microscopy CHARLIER—Coronary Vasodilators Gross—Oncogenic Viruses MeErRcER—Keratin and Keratinization HEATH—Organophosphorus Poisons CHANTRENNE— The Biosynthesis of Proteins RivERA—Cilia, Ciliated Epithelium and Ciliary Activity ENSELME—Unsaturated Fatty Acids in Atherosclerosis PLANT PHYSIOLOGY SUTCLIFFE—Mineral Salts Absorption in Plants SIEGEL— The Plant Cell Wall ANIMAL HORMONES A comparative survey Part I—Kiunetic and Metabolic Hormones PENELOPE M. JENKIN M.A., D.Sc. Senior Lecturer in Zoology, Bristol University Associate of Newnham College, Cambridge with a foreword by JouN E. Harris, C.B.E., M.A., Ph.D., F.R.S. Professor of Zoology in the University of Bristol PERGAMON PRESS NEW YORK - OXFORD - LONDON 1962 PARIS PERGAMON PRESS INC. 122 East 55th Street, New York 22, N.Y. 1404 New York Avenue N.W., Washington 5, D.C. PERGAMON PRESS LTD. Headington Hill Hall, Oxford 4 & 5 Fitzroy Square, London, W.1 PERGAMON PRESS S.A.R.L. 24 Rue des Ecoles, Paris V° PERGAMON PRESS G.m.b.H. Kaiserstrasse 75, Frankfurt-am-Main Copyright © 1962 Miss P. M. JENKIN Library of Congress Card Number 60-8977 Set in Imprint 10/11pt and Printed in Great Britain by Cox and Wyman Ltd., London, Reading and Fakenham CONTENTS PAGE List OF ‘TABLES eit ACKNOWLEDGEMENTS ix PREFACE Xill CHAPTER 1 INTRODUCTION 1 1.1 Discovery of Hormones 1 1.2 Chemical Activators 2 1.3. Mechanical Activation + 1.4 Definitions of Hormones 5 1.5 ‘Types of Hormones 6 1251 Kinetic hormones 9 1252 Metabolic hormones 10 1253 Morphogenetic hormones 11 1.6 Identification 12 1.7 References 16 Zz Sources OF KINETIC AND METABOLIC HORMONES 18 2.1 Ectodermal Sources 18 2.11 Secretory cells derived from the nervous system 19 ZA2 Endocrine glands derived from ectodermal epithelium 38 2.2 Endodermal Sources in Vertebrata 44 Z21 Isolated cells in the gut 44 222 Endodermal endocrine glands 46 2.3. Mesodermal Sources in Vertebrata 50 Z.31 Endocrine gland cells derived from coelomicepithelium 50 2.4 References 53 3 Kinetic HormMones—I. CONTROL OF MUSCLES AND PIGMENTARY EFFECTORS 56 3.1. Control of Muscles 37 3.11 Visceral muscle 37 g2 Somatic muscles 69 3.2 Control of Pigmentary Effectors f 0 5 BA Chromatophores with muscles 72 3.22 Pigmentary effectors with movable pigment granules = 3.3. References Vv 79829 vi CONTENTS CHAPTER 4 KInetT1c HorMoNES—II. CONTROL OF EXOCRINE AND ENDOCRINE GLANDS 4.1 Exocrine Glands 4.11 Digestive glands 4.12 Oviducal glands 4.13 Milk-secreting glands 4.14 Skin glands 4.2. Endocrine Glands 4.21 Ectodermal endocrine glands of Arthropoda 4.22 Endodermal endocrine glands of Vertebrata 4.23 Mesodermal endocrine glands of Vertebrata 4.3, General Considerations 4.31 Characteristics of kinetic hormones 4.32 Stimulation of the secretion of kinetic hormones 44 References 5 METABOLIC HORMONES 5.1 General Metabolic Rate Sti Respiration S12 Fat metabolism 5.2 Intermediary Metabolism of Carbohydrates and Proteins sw Carbohydrate metabolism 522 Protein metabolism 5.3. Balance of Monovalent Electrolytes and Water Bio! Balance of sodium ions (Nat) and of associated monovalent electrolytes (K+ and Cl-) 5.32 Water balance 5.4 Balance of Calcium and Phosphates 5.41 Balance of calcium 5.42 Balance of phosphates 5.5 General Consideraticns 551 Characteristics of the metabolic hormones 552 Control of the secretion of metabolic hormones §.53 Hormones and the environment 5.6 References GLOSSARY INDEX OF AUTHORS INDEX OF ANIMAL NAMES INDEX OF SUBJECTS fis Por rxaBpLlesS TABLE PAGE 1 Summary of the Main Types of Action of Vascular Hormones 8 2 Steps in Establishing Direct and Indirect Actions of Two Interacting Hormones 14 3. Ectodermal Sources of Kinetic and Metabolic Hormones 22 4 Cells in the Pars Distalis of the Adenohypophysis 44 Endodermal Sources of Kinetic and Metabolic Hormones in Vertebrata 46 6 Mesodermal Sources of Metabolic Hormones in Vertebrata 51 7 Kinetic Hormones Controlling Muscles 58 i Kinetic Hormones Controlling Pigmentary Effectors 71 9 Kinetic Hormones Controlling Chromatophores with Movable Pigment 87 10 Crustacean Hormones Controlling White Pigment in Relation to Light 96 11. Crustacean Hormones Controlling Red and Black e Pigments in Chromatophores in Relation to Light 98 12 Changes in Melanophore Index in Ligza 100 13. Illumination of Different Areas of the Eyes of Ligia 102 14 Kinetic Hormones Controlling Exocrine Glands in the Gut 118 15 Kinetic Hormones Controlling other Exocrine Glands 129 16 Endocrinokinetic Hormones Stimulating Endocrine a Glands of vil Vill TABLE 31 LIST OF TABLES PAGE Means of Controlling the Secretion of Kinetic Hormones 156-157 Metabolic Hormones Controlling Respiration and Fats 169 Changes in Oxygen Consumption in Astacus, follow- ing Sinus Gland or Eyestalk Removal 179 Changes in Fat Content of the Body of Crabs (Hemi- grapsus nudus) following Starvation and Sinus Gland Removal 188 Metabolic Hormones Controlling Carbohydrates and Proteins 190 The Effect of Asphyxia on the Concentration of Blood-Sugar 2 Changes in Body Composition of Crabs (Hemigrapsus nudus) following Starvation and Sinus Gland Removal 200 Hormones Associated with Nitrogen Excretion in Crabs 204 Metabolic Hormones Controlling Electrolytes and Water 208 Changes in Sodium and Potassium Concentration of Plasma and Muscle, following Adrenalectomy yA Changes in Excretion of Water, Sodium and Potas- sium, following Adrenalectomy 229 Metabolic Hormones Controlling the Balance of Calcium and Phosphates 241 Effect on Blood Calcium of Injection of Hypophysial Hormones into Xenopus 245 Changes in the Calcium Content of the Blood of the Crayfish (Astacus), following Removal of either the Sinus Glands or the Whole Eyestalks 248 Means of Controlling the Secretion of Metabolic | Hormones 254-255 ACKNOWLEDGEMENTS I ACKNOWLEDGE with gratitude the help and stimulus that I have received from many friends and experts, with whom I have discussed different sections of this book. Among these I would mention particularly Professor B. Hanstrém, who read the whole of chapter 3; Professor M. Thomsen and Dr. Ellen Thomsen who discussed the general plan with me; Professor H. E. Heller who read § 5.3 on electrolyte and water balance; Dr. J. A. Kitching, who read §§ 4.1 and 5 and Sir Francis Knowles and Dr. D. B. Carlisle who gave me much information on crustacean hormones and discussed the nomenclature which has been adopted here and, in part, in their own recent book. Professor J. E. Harris went far beyond the duties of an Editor in encouraging me in every way and not least in his penetrating discussion of fundamental problems of hormone action. Though all these have saved the book from many errors, I must accept sole responsibility for any that remain, as I do for the selection, interpretation and presenta- tion of the material. I am particularly indebted to Sir Francis Knowles for allowing me to reproduce some of his beautiful coloured photographs of chromatophores in Leander and to the publishers of Endeavour for supplying copies of the blocks for these. My warm thanks for permission to reproduce or adapt their figures are also due to Dr. E. Thomsen and Dr. S. P. Carlson, who provided photographs for Figures 2-3 and 3-19; to Dr. W. Junk of the Hague, for the block of Figure 5-6, and to all the other authors and publishers indicated in the legends and references, as well as to the following publishers and sponsors of journals: Academic Press Inc. (Figs. 2-10, 4-10, 4-11, 5-10). American Institute of Biological Sciences, Washington (Fig. 3-23). American Physiological Society (Figs. 3-7, 4-2, 4-3, 4-4, 4-5, 5-9, 5-22). ix x ACKNOWLEDGEMENTS Chas. J. Branford, Co. (Fig. 3-3). Butterworths Scientific Publications (Figs. 5-14, 5-23). Cambridge University Press (Figs. 3-1, 3-5, 3-6, 3-14, 3-16, 3-20, 3-21, 3-22, 3-24, 5-1, 5-2, 5-3, 5-12, 5-13, 5-16, 5-20, 5-24, 5-25, 5-26). Professor I. Chester Jones (Figs. 5-12, 5-13, 5-20, 5-24). Colston Research Society (Figs. 5-14, 5-23). Company of Biologists (Figs. 2-1d, 2-4, 5-1, 5-2). Council of the Marine Biological Association of the United Kingdom (Figs. 3-1, 5-3, 5-26). DD: C. Heath & ‘Co.(Fig. 5-8): Koninklijke Vlaamsche Academie voor wetenschappen, Brussels (Fig. 5-11). Marine Biological Laboratory, Wood’s Hole, Massachusetts (Figs. 2-1c, 2-5, 2-7, 2-9b, 3-2, 3-12, 3-15, 5-4, 5-7). Masson et Cie. (Figs. 2-le, 2-9a). Oxford University Press (Figs. 2-1d, 2-4). Springer Verlag (Figs. 2-1b, c, g, 2-2, 3-9, 3-10, 3-11, 5-18, 5-19). Stazione Zoologica, Naples (Fig. 2-6). Charles C. Thomas (Figs. 2-14c, 2-15). University of Chicago Press (Figs. 5-5, 5-17). Verlag Birkhauser (Fig. 2-3). Wistar Institute of Anatomy and Biology, Philadelphia (Figs. 3213; 3-17): I am most grateful for the skill and understanding co-operation of Mr. W. R. B. Buchanan and his staff in redrawing many of these figures and interpreting my sketches; and for the clear photographs of my drawings, taken by Mr. Ken Wood, for Figs. 2-1 (a—f) and 2-14. I also wish to thank Miss J. McKinney for her help in completing the indices and not least Mrs. P. M. Richards for her unstinted help and encouragement in the prepara- tion of the typescript and the main part of the indices, and in correction of the proofs. Finally it is a pleasure to acknowledge my indebtedness for the many facilities afforded me by the Director and Library staff of the Marine Biological Laboratory, Plymouth, as well as by the staff of the Zoological Department at Bristol. This book 1s dedicated to my Students, who, by their questions, have stimulated me to write it FOREWORD A foreword, like an aperitif, should whet the appetite without dulling the critical appreciation of what is to follow. Many wise people therefore avoid them. Nevertheless, it is a personal pleasure for me to be asked to provide one to this volume, for the initiation of which I was, at least in part, responsible. Specialized scientific publications in these days may be broadly, and thus inaccurately, divided into scientific papers summarizing experiments, reviews summarizing scientific papers, and books summarizing reviews. Among the multitude of these last, the really interesting book is all too rare—one with a broad but scholarly treatment, which stimulates the reader to think about the subject, to produce his own ideas and to design his own experiments. Such a book must provide a sufficiently clear account of the experimental techniques for the student to appreciate the methods of study and their limitations; it must establish a theoretical background which gives coherence to the subject as a whole; finally it must tread sufficiently near to the frontiers of knowledge to provide a glimpse of what may lie beyond. Such a stimulus has already reached several generations of Bristol students through Dr. Jenkin’s lectures on hormones; I hope that in its present form her book will successfully challenge a wider audience. JoHn E. Harris PRE ACE THE IDEA of writing this book arose from lecturing on hormones to second and third year students of zoology, for whom the subject formed part of a course in comparative physiology. It was found that no introductory book covered the whole subject equally; even Hanstrém’s admirable Hormones in Invertebrates (1939) dealt with only a part of the field and was already out of date in 1956, when he assured me that he would not be rewriting it and encouraged me to attempt this general survey. To do so necessitated evolving a scheme within which to consider and select suitable examples from the mass of available material. This resulted in a comparative arrangement, which should be of general application, since it is based on the actions of hormones, rather than on their sources or on their phyletic distribution. The actions of hormones were then seen to fall into three well-defined groups, the kinetic, the metabolic and the morpho- genetic, although these had not all been named nor clearly defined at that time. Subdividing these groups brought together examples acting upon similar effectors, such as muscles, chromatophores or glands, or having similar metabolic actions, such as increasing water excretion, blood-sugar or respiration. Still further sub- division brought together the hormones that stimulate a given action or facilitate a given process and separated them from those having the opposite effects. When consistently adhered to, this approach helped to give a clear picture of hormone actions, to emphasize cases where antagonistic hormones were known and to draw attention to apparent gaps in recorded knowledge. In writing the book, invertebrates and vertebrates were placed side by side to show the extent to which both are now known to have hormones with similar actions. Describing the invertebrate examples before those from vertebrates was a deliberate attempt xill XiV PREFACE to emphasize this fact. To have given pride of place to the verte- brates might have given a more clear-cut picture, and could certainly have provided more abundant and detailed examples; but it would have thrown the intended comparison out of perspec- tive. The search for good examples among invertebrates proved unexpectedly successful. It has been decided, therefore, to publish the book in two parts instead of one; but the unified plan of relatively simple presentation is being maintained. The present part of the book covers only the kinetic and metabolic hormones, their sources, actions and the ways in which their secretion is controlled. The second part* will contain a similar treatment of the morphogenetic hormones, namely those concerned with growth, differentiation and reproduction; it will also discuss such topics as the relation of the chemical constitution of hormones to the sources from which they are derived and their type of action. A consideration of the distribution of hormones in the animal kingdom may also throw some light on the possible evolution of these chemical activators, as well as suggesting prob- lems for investigation. Many of these problems must be apparent to anyone who surveys the field of hormone research; yet it is permissible to assume that few research workers have time to undertake such a survey, as their own work becomes more and more specialized and results in the publication of books that are confined to single classes of animals or single endocrine organs. It is therefore hoped that the present work may be useful to some specialists as well as to the teachers and students for whom it is primarily intended. It has clearly not been possible for the writer to review the whole literature of so rapidly expanding a subject; but the main original papers on the kinetic and metabolic hormones of inverte- brates have been covered up to the summer of 1958, while the vertebrate examples have been checked by recent reviews and reports of symposia. The references at the end of each chapter show the sources used, but make no pretence to being complete, though they should provide a useful starting point for anyone wishing to go further. * Animal Hormones, a comparative survey. Part II. Morphogenetic Hormones, in preparation. : PREFACE —“ It is much to be regretted that these references are not more nearly up to date; but publication has been delayed by various unforeseeable causes, including the printing industry’s national dispute during 1959 and the writer’s serious illness. Finally a word of explanation about some of the things which have not been included in the book. Examples in which extracts of one kind of animal have been tested upon another kind have been avoided, on the grounds that they are apt to lead to unsound physiological deductions. Details of standard techniques are omitted, since the reader can refer to any physiological text- book for an account of such methods as recording muscle con- tractions by means of levers that mark a revolving smoked drum (e.g. Figs. 3-1 and 3-3). ‘The use of commercial hormone prepara- tions and methods of quantitative estimation of hormones by biological assay are also omitted, as being primarily of clinical interest. Since the book is intended for zoologists and comparative physiologists, the mammalian examples have been chosen from species other than man, while reference to pathological and clinical material has been omitted, as being outside their chosen field. Such material is easily accessible elsewhere, and should not be difficult to fit into the present framework, if the reader so desires. Bristol. Ps Mak CHAPTER. 1 INTRODUCTION 1.1 DiIscovERY OF HORMONES THE DISCOVERY of hormones was a late-comer in the study of physiology; the circulation of the blood was demonstrated in the seventeenth century by Harvey (1628), but it was more than two centuries before it was realized that chemical messengers could be carried in that circulation. The first hint of this was when Berthold (1849)* showed conclusively that the morphogenetic effects of transplanting the testes of cockerels must be transmitted by some factor in the blood. It was even longer before Oliver and Schafer (1895) found that a chemical extract of the adrenal medulla, if injected into the circulation, could induce a pronounced rise in blood pressure. In 1901 the active substance in this extract was isolated, identified and called ADRENALINE. The general term “Hormone” is derived from the Greek épuaw, meaning “I arouse’, and indicates the stimulating action of such chemicals; it was first used by Starling (1905) for SECRETIN, that had been discovered in 1902 and shown to induce the flow of alkali from the pancreas. Two hormones concerned with the cure of human disease, INSULIN for the control of diabetes mellitus, and THYROXINE for cretinism, were armong the more spectacular discoveries of the early twentieth century, and led to an intensive search for more hormones in man and other mammals. This resulted in the gradual discovery of some thirty kinds of endocrine cells and glands that can produce minute quantities of chemical substances which are carried in the blood, to stimulate or inhibit various specific effectors, or to control different aspects of metabolism and morphogenesis. * See Harris (1955) for a translated account of his experiments. B 1 Z INTRODUCTION The first indication of any hormone in an invertebrate was that postulated by Kopeé (1922) as carrying the brain stimulus for moulting in Lymantria. 'Then Koller (1927) found a blood-borne factor controlling the colour changes of certain shrimps, and Perkins (1928) located its source in the eyestalk. The discovery of other hormones has followed, mainly in crustaceans and insects, where they have almost as many actions as those carried out by the better-known hormones of vertebrates. 1.2 CHEMICAL ACTIVATORS During this period, when hormones were being discovered in ever-increasing numbers, different kinds of chemical activators were being found in other fields of biology. Substances akin to hormones were found in plants; nerve transmission in vertebrates and some invertebrates was found in many unrelated species to be due to release of either acetylcholine or adrenaline, at the point of contact between one neuron and the next, or between the motor axon and its effector. The control of the pattern of development in early embryos of Amphibia was found to be due to the diffusion from cell to cell of particular chemical substances or organizers; these substances were not specific in that they were capable of producing similar effects in a wide range of genera (Spemann and Mangold, 1924). Some order was brought into the variety and diversity of these and other chemical activators by Huxley (1935) in an important scheme of classification. Its main weakness was that it did not include neurosecretory cells derived from nerve cells and capable of yielding hormones. These cells had been recognized histologi- cally in vertebrates by Dahlgren (1914), and in some invertebrates by Hanstrém (1931); but their action in releasing hormone-like substances into the blood was first established by the Scharrers (1937). They are now well known in Annelida, Arthropoda and some other invertebrates as well as in vertebrates. Huxley’s (1935) classification of chemical activators may therefore be modified as follows, to include neurosecretion: A. Para-ActivaTors. By-products of normal and pathological metabolism with effects on correlation or differentiation, e.g. carbon dioxide in its effect on the respiratory centre. § 1.2 CHEMICAL ACTIVATORS 3 B. ‘TRUE ACTIVATORS. Chemical substances produced by the organism and exerting specific functions in regard to corre- lation or differentiation: 1. Local activators, with effects on the same cell, or cells, within which they are produced. (a) Intracellular activators (‘intracellular hormones” of Goldschmidt), acting in each cell singly and being the direct expression of gene activity, in relation to regional differentiation. (b) Regional activators, responsible for the chemodifferentia- tion of specific regions in embryos and for growth gradients. 2. Distance activators, with effects on cells other than those in which they are produced. (a) Diffusion activators, distributed by diffusion through the tissues. (1) Direction of transport restricted by structural organization. Growth hormones in plants. (11) Diffusion restricted to tissues in direct contact. Organizers in embryos and “‘organisines”’ in animals without a circulatory system. It is possible that the cortical releasing factor, CRF, from the brain of vertebrates should also be included here (§ 4.323). (iii) Diffusion restricted mainly by chemical means. Neurohumoral secretions at nerve- and _ neuro- secretory cell-endings (“neurohormones”’ of Welsh, 1955). (iv) Diffusion unrestricted, the substances passing out of the tissues and into the surrounding medium to act on other individuals, usually of opposite sex. These include ‘‘gamones”’ and ‘‘ectohormones’’. (b) Circulatory activators or vascular hormones, distributed to all parts of the body in the blood circulation, so that their actions must be limited by the sensitivity or com- petence of the tissues which they reach. They may be secreted by: : peg! (i) Isolated cells such as those of unknown origin in the gut mucosa of vertebrates (§ 2.21). 4 INTRODUCTION (ii) Neurosecretory cells with the swollen ends of their original axons forming storage-and-release organs (“neurohaemal organs” of Knowles and Carlisle, 1956), that make contact with blood vessels (§ 2.11). (iit) Endocrine gland cells, which secrete internally into the blood and are formed from almost any tissue of the body, including the nervous system (in which case the distinction from neurosecretory cells is only one of the degree of their histological modi- fication). The chemical activators to be considered as “animal hormones” in the present book are the circulatory activators, or vascular hormones (2b). Yet since there is really no logical point at which some of them can be separated from other neurosecretions, or even from the organisines which have actions so much like those of morphogenetic hormones, reference to these will have to be made in the relevant sections, as will chemicals involved in nervous stimulation, where their functions overlap those of hormones. The complex interaction of many of these chemical activators is well illustrated by considering the way in which genes initiate, and hormones complete, the differentiation of the gonad rudiment into a testis or an ovary within an embryo, the development of which starts under the general control of an organizer, continues by progressive chemcdifferentiation, and cannot be completed without the combined action of the nervous system and yet other hormones! 1.3 MECHANICAL ACTIVATION The only other means made use of by animals for co-ordinating their activities is purely mechanical. This can act in the absence of nerves or of any chemical activators, and may well be a primitive way of transmitting control. The action of a current of water stimulating the sponge osculum to remain open appears to be purely mechanical, but it is not certain that some chemical may not be diffusing from cell to cell. The best example is in the locomotion of the earthworm, where the muscles contracting in one segment stretch those in the adjacent segment behind, and § 1.4 DEFINITIONS OF HORMONES 5 stimulate them to contract in their turn to give a wave of con- traction passing back from segment to segment. This is a primitive method of control which may be supposed to have preceded that by the nervous system. A distinct contrast to this is afforded by the use of mechanical distention of the stomach as a means of initiating the secretion of GASTRIN, one of the hormones from the mammalian gut. For in this case mechanical stimulation, like the direct chemical stimulation which acts upon other endocrine cells in the same region, seems to be part of a highly specialized system for harmonizing the succes- sive stages of digestion, and to have succeeded the nervous control that is used for a similar purpose by cold-blooded vertebrates 4:11), 1.4 DEFINITIONS OF HORMONES The simplest and earliest definition of hormones as ‘“‘chemical messengers’? must be amplified, if it is to limit the use of the term “animal hormones” to the circulatory activators. A well-established definition of a hormone is ‘‘a physiologic organic compound produced by certain cells and carried by the blood to distant cells, the activities of which it influences” (Selye, 1947). This is still rather too loose a definition; it accords more nearly with what has been termed a “humoral mechanism”’, or ‘“‘a process which has been demonstrated to be independent of nervous connections between the site of stimulation and the effector site, and is, therefore, considered to be transmitted by a blood-borne substance, but in which the hormonal or non- hormonal nature of the blood-borne substance may be uncertain”’ (Grossman, 1950). This refers particularly to substances like histamine, which may occur in the blood in the abnormal condi- tions of an experiment and yet play no part in normal physiology, and to the so-called ‘‘secretagogues’’. These last are substances that may be found in extracts of gut cells; they have the capacity to stimulate enzyme secretion by the gut glands but are not natural secretions (§ 4.11). It has already been indicated that a hormone is not necessarily secreted by a gland, nor is its secretion by any means always stimulated by nerves, as some elementary definitions have 6 INTRODUCTION suggested (e.g. Yapp, 1942). A sound definition must be such as to include GASTRIN, which comes from isolated cells and for which the stimulus to secretion is the direct action of mechanical pressure in the gut lumen, and also INSULIN, which comes from small groups of gland cells and for which the stimulus to secrete is the level of glucose concentration in the blood. It must also include the hormones which stimulate the secretion of other hormones, like the INTERSTITIAL-CELL-STIMULATING HORMONE from the adenohypophysis, which stimulates the secretion of TESTOSTERONE from the testis, as well as those hormones the action of which is inhibitory rather than activating. Huxley (1935) suggests that a hormone is “‘a chemical substance produced by one tissue, with the primary function of exerting a specific effect of functional value on another tissue’’; but, as he admits, this has the teleological implications of any functional account. Moreover, it loses sight of the fact that some chemical substances, such as adrenaline, are present as by-products with no apparent function in many primitive animals, and seem only to have been salvaged for use as hormones in the more highly evolved phyla. It is also well to remember that hormones, or their active constituents, are usually rather stable compounds, able to persist for some time in the blood stream and yet composed of molecules sufficiently small to pass through the walls of blood capillaries and cell membranes to reach their targets. The vascular hormones, with which this book is mainly con- cerned, may best be defined as specific organic substances produced by isolated cells, or by a tissue which may form a gland; they activate or inhibit effects of functional value occurring in other cells or tissues, to which they are carried in the blood. 1.5 TYPES OF HORMONES Although many actions in a wide range of animals can now be attributed to hormones, it cannot be expected that any functions will belong to hormones alone; rather must they be recognized as playing but a small part within the complex co-ordination of metabolic and other processes that supply and direct the food and energy as between the multifarious daily activities of the animal §1.5 TYPES OF HORMONES 7 and its growth and reproduction. These processes must be con- trolled to fit the frequent changes both within the animal and in its environment. At different times of the day, or the tide, or the year, the energy must be directed to different purposes, to serve the needs of both individual and race survival. The greater part of this control is achieved by the nervous system; but hormones may come in at almost any point, sometimes independently, but more often in direct or indirect response to nerve stimulation. It is not merely convenient to review the hormones in relation to their actions; it also allows of some interesting comparisons being made between those having similar actions in invertebrates and vertebrates, and reveals a notable degree of correlation between some of their actions and the sources from which the hormones come. It also shows certain striking gaps: some animals lack hormones with particular actions; many invertebrate phyla, or classes, lack hormones altogether. It may be supposed that in many cases the main detectable actions of a hormone represent its physiological functions within the normal animal; but other actions, which are apparent experimentally, may be accidental and without true functional significance. Their actions will be considered under three main headings: (1) Kinetic, or the control of effectors (§ 1.51 and §§ 3 and 4); (2) Metabolic, or the control of cell biochemistry (§§ 1.52 and 5); (3) Morphogenetic, or the control of growth and differentiation (§ 1.53, and Part II, to be published separately). Each of these groups can be further subdivided in relation to the particular organs or processes controlled (Table 1). This grouping of hormones is not yet widely used; but it has been found in practice to afford a very good working framework within which to consider the available information, with the minimum of ambiguity or overlap. It has been accepted in a recent presenta- tion of crustacean hormones (Carlisle and Knowles, 1959), together with the terms kinetic and endocrinokinetic suggested to them by the writer (Carlisle and Jenkin, 1959). The term kinetic hormone corresponds to their previous term of ‘‘energetic hormone (Knowles and Carlisle, 1956). INTRODUCTION TABLE. 1. SUMMARY OF THE MAIN TYPES OF ACTION OF VASCULAR PART I 5.3 5.4 PART II § 3 § 4 HORMONES EXAMPLES * ACTION . VERTEBRATE INVERTEBRATE KINETIC Contraction of muscle | Adrenaline | Corpus car- diacum Concentration and dis- | W and By PLH and PDH persion of pigment é Secretion of exocrine | Secretin Gonad glands Secretion of endocrine | ACTH Intercerebral glands neurosecre- tory cells METABOLIC Control of respiration rate Carbohydrate and pro- tein balance Eloctrolyte and water balance Ca and P balance MORPHOGENETIC General growth Moulting Metamorphosis Regeneration Growth of glands Gonad maturation Gamete release Differentiation of geni- tal ducts Development of second- ary sexual organs Thyroxine Corpus allatum Insulin Sinus gland Brain Parathormone | Y-organ ADH ““Growth”’, Sinus gland ? ool al Thyroxine Y-organ Thyroxine Prothoracic gland STH ““Organisine”’ Thyroxine ? (source unknown) FSH Corpus allatum LH Corpus allatum Oestrone or Testis Testosterone Testosterone | Vas deferens gland | * See tables in each section for complete lists of the hormones and their sources referred to in this book. + See glossary. § 1.51 KINETIC HORMONES 9 1.51 KINETIC HORMONES The kinetic hormones act upon effector cells or organs, to produce repeatable reactions mainly concerned with feeding, digestion and protective colour change, and so are often related to the short-term interaction of the individual with its environ- ment. The results that they produce are relatively rapid compared with other types of hormone action, but considerably slower and more long-lasting than the nerve action which often controls similar effectors. Their action is also more widespread than that of nerves, since they are distributed by the circulation throughout the body, and cannot, as a rule, be used to cause the contraction of one muscle and not another, or to produce a pattern by con- centrating some chromatophores and dispersing others, unless the effectors themselves are differentiated. Many, but by no means all, of the hormones in this group are neurosecretory substances (2b. ii, p. 4). Most of the others come from ectodermal glands (2b. iii), except the notable group produced from the isolated cells (2b. i) in the mammalian gut. The last-named are stimulated directly (§ 4.11); but the rest are all controlled by the nervous system, apart from a few anomalous hormones which appear to have kinetic actions but to be derived from mesodermal glands (§ 4.324). There is one group of hormones within the kinetic type which calls for special mention at this stage, and for an elucidation of the rather confused nomenclature associated with it. These are the hormones for which the name endocrinokinetic is adopted here. They all stimulate the secretion of other hormones from endocrine glands (2b. iii), and it seems logical to class this action, with that of stimulating exocrine glands, as kinetic. But the situation in which a series of two hormones is involved is more complex than those with but one hormone, and it seems to warrant somewhat different treatment. These endocrinokinetic hormones are frequently desig- nated by the suffix “trophic” or “tropic’’, as in thyrotrophic or gonadotropic ; but the suffix is not confined to this type of hormone, since it also occurs in chromatophorotrophic, where the effector is a chromatophore, and not an endocrine gland at all. Even the international decision taken in 1939 that the form trophic should be used in all cases has led to no uniformity of spelling! 10 INTRODUCTION Among vertebrates all the endocrinokinetic hormones are secreted by the adenohypophysis (anterior lobe of the pituitary body); but so also are hormones with such morphogenetic actions as stimulating the growth and maturation of the gonads. Yet the term trophic or tropic has been applied indiscriminately to both, as in the case of the gonadotrophins, of which the interstitial- cell-stimulating hormone, ICSH, is endocrinokinetic and causes hormone secretion from the gonads, but the follicle-stimulating hormone, FSH, is morphogenetic and causes their growth. There are as yet no separate names for the similarly separable actions of the thyrotrophic hormone, 'TSH, or of the adrenocorticotrophic hormone, ACTH; but there is a mounting body of evidence to show that the two types of action are often, and perhaps always, due to distinct, albeit closely similar hormones (§ 4.2). If in some cases the two actions are really inseparable, they may perhaps be likened to the motor and trophic actions of one and the same nerve. There are still a few effectors for which no example of kinetic hormone control is known: namely, luminous and electric organs among those usually controlled by nerves, and flagella and nema- tocysts among those for which no internal control is known. 1.52 METABOLIC HORMONES The metabolic hormones are concerned particularly with the control of metabolic activities, at the physico-chemical or bio- chemical level, within the cells of the animal, e.g. with adjustment of respiratory rate (§ 5.1), supply of sugars and proteins to the tissues (§ 5.2), and their electrolyte and water balance (§§ 5.3 and 5.4). Such processes often have a basic rate that seems to be an intrinsic or genetic property of the cells which carry them out. The rates are rarely under nerve control, but hormones may induce changes in them; in many instances, a pair of hormones act together, one increasing the rate and the other decreasing or inhibiting it. ‘This is particularly clear in the control of electrolytes and water in the vertebrate kidney (§ 5.3). Many of the metabolic hormones in Arthropoda are products of neurosecretion, and stimulate the rate of the process in question but are themselves subject to nervous inhibition. Other metabolic hormones, especi- a a) MORPHOGENETIC HORMONES 11 ally in vertebrates, are secreted by endocrine glands and are sub- ject to control by endocrinokinetic hormones. The nervous system takes but a small share in their control, except in emergencies such as haemorrhage or other forms of shock. More often the equili- brium is maintained by a “feed-back” system, whereby the accumulation of some product of the hormone’s action inhibits further secretion of the hormone until the accumulation is again reduced. This applies directly to some metabolic hormones, and indirectly to others, through its control of the endocrinokinetic hormones that stimulate them (§ 5.5). 1.53 MORPHOGENETIC HORMONES The morphogenetic hormones produce long-term changes that involve cell division, growth and differentiation; in contrast to the quick-acting kinetic hormones, their effects can neither be reversed nor repeated, at least for a considerable time. Formerly, these hormones were grouped under a more widely defined ‘‘metabolic’”’ heading. ‘There was some justification for this, in that growth is not possible without an adjustment of the metabolic processes to provide the growing cells with the necessary building materials for protein synthesis, and energy supplies in the form of glucose (cf. §§ 5.2 and 5.5). Yet these and other metabolic processes con- tinue throughout the life of the animal, and are not necessarily linked to morphogenesis, which is often intermittent, as is easily seen in moulting and metamorphosis and in the changes associated with seasonal reproduction. Morphogenetic hormones affecting growth and regeneration are present in several phyla, including Annelida and Mollusca, from which no metabolic hormones have so far been reported, and the means of controlling them is, in many cases, still unknown. It is only in Crustacea, Insecta and Vertebrata that nearly all the morphogenetic factors are secreted as vascular hormones and are controlled by endocrinokinetic hormones, which link the processes of growth, differentiation and reproduction indirectly to the nervous system, and thence to seasonal changes in the environ- ment (§ 4.232). Hormonal control of moulting and metamorphosis is now well known in Crustacea and Insecta, in both of which ectodermal 12 INTRODUCTION glands from the antennary or maxillary segments secrete moult- promoting hormones. Their secretion is stimulated by an endocrinokinetic hormone, prothoracotrophin, in Insecta (§ 4.211), and probably by a similar hormone in Crustacea. Otherwise the two classes differ in that, in the latter, moulting is restrained by a moult-inhibiting hormone; this does not occur in Insecta, in which a so-called juvenile hormone from the corpora allata inhibits metamorphosis only. The initiation of metamorphosis in Amphibia by thyroxine, with its dependence on the availability of iodine, was an early discovery; its control by the endocrino- kinetic thyrotrophin, TSH, was established later. The differentiation of the genital ducts and other sexual characters has also been found to depend on hormones in a number of invertebrates, as well as in vertebrates, where the pattern of control differs in detail in different classes (§ 4.234 and Part II, § 4). Only a few of the hormones producing these effects are shown in Table 1; their detailed treatment is reserved for the second Part of this work. The so-called “organisines”, which stimulate regeneration in Platyhelminthes, are not vascular hormones, since in the absence of any circulation in these animals it must be assumed that the substances diffuse through the tissues in a way that is reminiscent of embryonic organizers, as their name suggests (Dubois and Lender, 1956). 1.6 IDENTIFICATION The technique of finding, testing and confirming the presence and action of a hormone is exacting, and needs many controls if the results are to be conclusive. It is difficult if only a single hormone with a relatively clear-cut effect is under investigation; for quite a long series of experiments is needed to elucidate the situation. All too often some steps in the proof are missing, either for technical reasons or because their importance was not fully realized during the early stages of hormone investigation. Histological examination of tissues that are suspected of sec- reting hormones is one side of the investigation, since cells capable of this type of chemical activity often have a recognizable cytologi- cal appearance (Figs. 2-2 and 4-7), with granular precursors of the § 1.6 IDENTIFICATION 13 secretion and enlarged nuclei. Once the secreting cells have been located, it may be possible to make extracts of tissue containing them, and to compare this with extracts of adjoining tissue containing no such cells, in order to arrive at more definite results than can be obtained from extracts of whole structures like the crustacean eyestalk (§§ 3.12 and 3.223). A carefully planned experimental investigation is also essential if the action of the hormone is to be fully established. This usually falls into one of two categories, the pharmacological or the physiological: in the first, it is shown that certain extracts, or chemicals, have effects upon the animal, such as stimulating muscle contraction, or raising the salt output in the urine; in the second, and usually more difficult category, an attempt is made to prove that the chemical in question is used in the normal physi- ology of the animal to control the same process. In general, it can be said that if only one hormone is concerned, its control of a certain reaction can be sufficiently proved if adequately controlled experiments show that: (1) removal of the source of the hormone is followed by loss of the reaction, (ii) injection of an extract from the source can restore the reaction, (iii) removal of any other structure does not cause loss of the reaction, (iv) injection of any other extract does not restore the reaction. It is better if the reaction is shown by an organ with no nerve connections, and if it can be interrupted by ligation of its blood supply. If the reaction can be restored by injection or transfusion of blood from another individual in which hormone secretion has been stimulated by natural means (cf. Fig. 3-3), the proof that this hormone plays a part in the natural physiology of the animal is more convincing. To complete the identification of any parti- cular hormone, it may be necessary to separate it from others which can be extracted from the same source, and the problem is never finally elucidated until the chemical constitution of the pure hormone is known. . If more than one hormone is involved in the control of a reaction, 14 INTRODUCTION TABLE 2. STEPS IN ESTABLISHING DIRECT AND INDIRECT ACTIONS OF TWO INTERACTING HORMONES CONTROLS ON LITTER EXPERIMENTS ON RATS RESULT MATES PREFERABLY OF | RESULT SAME SEX 1. Operative removal | Death 1a. Mock operation | Survival of the adrenal cor- tex 2-Opsas 1 =- injec- ‘| Survival® "| 2a. Opsias t+ other | Death tions of pure cor- injections} : tex extract 3. Op: removal of | Death 3a. Mock operation Survival Adhp: (including source of ACTH) leaving cortex in- tact but unstimu- lated 4. Op: as 3 + injec- | Survival* | 4a.Op: 3 + injec- | Death tion of one frac- tions of any tion of Adhp: ex- other Adhp: ex- tract (ACTH) tracts 5. Op: removal ‘of | Death 5a. Mock operations | Survival Adhp: and cortex of equal severity 6: Op: as: 5 +‘ anjec- | Death 6a. Op: as 5 + injec- | Death tions of ACTH (as tion of other ex- 4) tracts 7. Op: as 5 + cortex | Survival* | 7a. Op:as 5 + myjec=") Meath extract (as 2) tion of other ex- tracts * For as long as injections are maintained. + NaCl injection can mitigate the effect of cortex removal for some time (§ 5.311). Adhp = adenohypophysis. ACTH = adrenocorticotrophin. Argument: Experiments 1 to 4. The result of removing either the cortex or the adenohypophysis is the same (1 and 3). So is the effect of injecting extracts of the gland removed, i.e. replacement of either missing hormone can maintain life (2 and 4). The operation itself and post-operative shock as causes of death are ruled out by survival of controls (1a and 3a), on which operations of comparable severity are performed, but without touching the endocrine organs. Injections of other materials are shown by controls to be ineffective (2a and 4a). By fractionation of the extracts of the adenohypophysis, and by testing § 1.6 IDENTIFICATION 15 each separately (4), it is shown that ACTH is the only effective substance. At this point it still remains an open question as to whether the two hormones that have been identified have independent actions, or whether one is direct and the other indirect, as it would be if one were an endocrinokinetic hormone. Experiments 5 to 7. The results of these further experiments and their controls give the minimum of information upon which these questions can be decided. CoRTEX EXTRACT alone is effective in the absence of both sources (7), and is therefore the direct metabolic hormone, whereas ACTH is ineffective (6), unless the CORTEX TISSUE is present (4). Since the unstimulated cortex is ineffective (3), the action of ACTH must be to stimulate the cortex to secrete (as in the normal animal, and in experiments la and 4). Conclusion: ACTH is, therefore, the endocrinokinetic hormone stimulating the secretion of a metabolic hormone from the adrenal cortex. the experimental investigation becomes more complex. The tabulated theoretical scheme for a set of experiments, to show the relation of a metabolic hormone from the adrenal cortex and the adrenocorticotrophic hormone, ACTH, from the adeno- hypophysis (Adhp. ‘Table 2), is given as an indication of the mini- mum number of experiments involved. Taking ‘‘death” or “survival” of the animal as the criterion of the hormone’s action is obviously much too crude, and should be replaced by some surer physiological test of the metabolic activi- ties affected, such as the measurement of blood-sugar; but in that case, the opposing action of insulin has also to be taken into account (§§ 4.2 and 5.2). For obvious reasons of space it will not be possible in the cases cited below to give full details and results of all the experiments upon which the conclusions are based; but an attempt will be made to give some of the clearer examples in enough detail to indicate how far the technique of the original work was satis- factorily controlled. 16 INTRODUCTION 1.7 REFERENCES BERTHOLD, A. A. (1849). Transplantation der Hoden. Arch. anat. Physiol., Lpz. 42-46. : CaRLISLE, D. B. and JENKIN, P. M. (1959). Terminology of hormones. Nature, Lond. 183: 336-337. CaRLISLE, D. B. and KNow _es, F. G. W. (1959). Endocrine Control in Crustaceans. Cambridge: University Press. DAHLGREN, U. (1914). The electric motor nerve centers in the skates (Rajidae). Science, 40: 862-863. Dusois, F. S. and LeNnper, T. (1956). Corrélations humorales dans la régénération des planaires paludicoles. Ann. Sci. nat. (b) Zool. 18: 223-230. Grossman, M. I. (1950). Gastrointestinal hormones. Physiol. Rev. 30: 33-90. Hanstrom, B. (1931). Neue Untersuchungen iiber Sinnesorgane und Nervensystem der Crustaceen. I. Z. Morph. Okol. Tiere, 23: 80-236. Harris, G. W. (1955). Neural Control of the Pituitary Gland. London: Edward Arnold Ltd. Harvey, W. (1628). Exercitatio anatomica de motu cordis et sanguinis in animalibus. Frankfort: Fitzer. Hux ey, J. S. (1935). Chemical regulation and the hormone concept. Biol. Rev. 10: 427-441. KNOWLES, F. G. W. and Car .isLe, D. B. (1956). Endocrine control in the Crustacea. Biol. Rev. 31: 396-473. Kotter, G. (1927). Uber Chromatophorensystem, Farbensinn und Farbwechsel bei Crangon vulgaris. Z. vergl. Physiol. 5: 191-246. Koperé, S. (1922). Studies on the necessity of the brain for the inception of insect metamorphosis. Biol. Bull. Wood’s Hole, 42: 323-342. Ouiver, G. and ScuArer, E. A.(1895). The physiological effects of extracts of the suprarenal capsules. 7. Physiol. 18: 230-276. Perkins, E. B. (1928). Color changes in Crustaceans, especially in Palae- monetes. F. exp. Zool. 50: 71-106. Scuarrer, E. and Scuarrer, B. (1937). Uber Driisen-Nervenzellen und neurosekretorische Organe bei Wirbellosen und Wirbeltieren. Biol. Rev. 12: 185-216. SELYE, H. (1947). Textbook of Endocrinology. Montreal, Canada: Acta Endocrinologica, Université de Montréal. SPEMANN, H. and Manco p, H. (1924). Uber Induktion von Embryonal- anlagen durch Implantation artfremder Organisatoren. Arch. mikr. Anat. 100: 599-638. STARLING, E. H. (1905). The Croonian lectures on the chemical correla- tion of the functions of the body. Lancet, 2: 339-341. § 1.7 REFERENCES il We LsH, J. H. (1955). Neurohormones. In The Hormones, edited by G. Pincus and K. V. THimann. New York: Academic Press Inc. 3: 97-151. Yapp, W. B. (1942). An introduction to Animal Physiology. Oxford: Clarendon Press. Youne, J. Z. (1957). The Life of Mammals. Oxford: Clarendon Press. CHAPTER: 2 SOURCES OF KINETIC AND METABOLIC HORMONES BEFORE DESCRIBING the actions of the various kinetic and metabolic hormones, an account will be given for reference of the sources from which they are derived and of some of the ways in which they reach the blood stream. The cells in the animal body which are able to secrete hormones into the blood can be conveniently grouped by their embryological origins. Invertebrate examples will be given first. It will then be noted that the sources of those kinetic and metabolic hormones that are so far known from invertebrates all come from the ectoderm (§ 2.1) and that it is only in the vertebrates that the endoderm (§ 2.2) and the mesoderm (§ 2.3) also provide sources for these kinds of hormones. The sources of morphogenetic hormones, which affect growth, differentiation and reproduction, include the gonads of both invertebrates and vertebrates, as well as the ectodermal glands which secrete moulting hormones in the Arthropoda, namely, the Y-organ in Crustacea and the prothoracic glands and their homologues in Insecta. Passing references will be made to some of these morphogenetic hormones in the chapters that follow, but their main actions and details of their sources will be described in Part II. 2.1 ECTODERMAL SOURCES The hormone-secreting, or endocrine, cells which are formed from the embryonic ectoderm can be divided into those which arise from the nervous system (§ 2.11) and those which arise from non-nervous epithelium (§ 2.12); but the distinction may be rather arbitrary, since the stomodaeal epithelium of the cephalo- 18 § 2.11 SECRETORY CELLS FROM THE NERVOUS sysTEM 19 pods gives rise to non-nervous cells, whereas that of insects gives nervous cells. 2.11 SECRETORY CELLS DERIVED FROM THE NERVOUS SYSTEM A large number of hormones in many phyla are now known to be secreted by nerve cells or their derivatives (Fig. 2-1). Some of these have become so specialized for secretion that they have lost almost all histological semblance of neurons, and their connection with them is only apparent in their development or in the quality of their secretion. Of such are the cells of the corpus cardiacum of insects and the adrenal medulla of vertebrates (Fig. 2-1e and f). On the other hand, many secretory cells derived from neurons come within the histological definition of NEUROSECRETORY CELLS; these have only recently been recognized as sources of hormones because they are less easy to discern than the compactly aggregated endo- crine glands, formed by most other cells of internal secretion. They differ from neurons in secreting microscopically visible quantities of granules or droplets, while retaining such characters as Nissl bodies in the cytoplasm and axons with neurofibrillae; they may or may not have dendrites (Fig. 2-15, c and d). There seems to be no real need to separate neurosecretory cells, which secrete hormones into the blood, from any other endocrine cells derived from either the nervous system or any other part of the ectoderm, for their secretory activity is similar, although their histological form is different. Since, however, much attention has been focused recently upon neurosecretion, it will be well to summarize some of the main points about it. Neurosecretory cells have been identified in animals represent- ing most of the phyla with centralized nervous systems; but as yet their production of vascular hormones has only been demon- strated in a relatively small proportion of these phyla. In some of these phyla, and notably in Annelida, they are only known to yield morphogenetic hormones (Part II); but in the Mollusca, Arthropoda and Vertebrata neurosecretory cells are known to secrete vascular hormones with kinetic and metabolic actions. Neurosecretory cells can usually be recognized within the ner- vous system by their large size (often 30 » or more in diameter), 20 SOURCES OF KINETIC AND METABOLIC HORMONES with large nuclei and secretory granules in the cytoplasm; but the latter may vary with the phase of the secretory cycle. This will affect both the microscopic appearance of the cells (Fig. 2-2) and their reaction to histochemical tests which can be applied to the secretion. Some of these cells appear to release their secretion, or neurohormone, where it can only diffuse through the closely adjacent tissue without reaching the circulation. It is then difficult to distinguish the action experimentally from that of a normal motor nerve, especially as the release of secretion is probably accompanied by electrical changes in the axon similar to those accompanying the nerve impulse. The distinction between such cells and ordinary neurons seems only to be one of degree, for it depends upon the presence or absence of “granules”. This in turn depends rather arbitrarily upon the limits of resolution of the ordinary light microscope. Since the abundant fine granules of adrenaline, which stain brown with chromates, can be readily seen with the light microscope in cells of the adrenal medulla, there seems little reason to doubt that the minute quantities of the same substance, secreted at sympathetic nerve endings, could also be seen by using the greater magnification that can now be achieved by the electron microscope. Yet these nerves are not usually con- sidered to be neurosecretory. Other neurosecretory cells have simple axon endings that discharge their secretion into blood vessels, thereby clearly acting as a source of a vascular hormone. The secretion is formed as granules or droplets either in the cytoplasm immediately surrounding the nucleus, or within the nucleus itself. Thence the granules have been seen to move slowly along the axon and are probably carried in the axoplasm current, which flows at a rate of about 3 mm per day (the movement of endoneural fluid is about 20 times as fast). They accumulate at the unbranched ends of the axons, which become swollen and are often aggregated together to form a storage-and-release organ at the point where the hormone is passed into the blood. Such structures have been called neurohaemal organs (Carlisle and Knowles, 1953). The secretion can sometimes be detected for a short distance even after its discharge into the blood vessel. It is probable, however, that the visible secretion often acts as a “‘carrier’’, to which is attached the chemical substance that acts § 2.111 SECRETORY CELLS FROM THE NERVOUS SYSTEM 2] as the hormone. The carrier may be some large molecule, like a protein, which helps to anchor the smaller hormone molecule in the cell until the time for its release. The hormone is then separ- ated from the carrier, and apparently becomes free to enter the blood and be passed on to the tissues. The carrier is usually visible in the living cells by dark-ground illumination because of its highly refractile granules which show up as bright spots (Fig. 2-3); in fixed preparations the carrier often stains, in a character- istic but not specific way, with Gomori’s chrome haematoxylin phloxin and other stains, such as Mallory’s triple stain for connec- tive tissue (Scharrer and Scharrer, 1954a). There is increasing evidence that neurosecretory cells not only secrete a greater quantity of some active chemical substance than do typical nerve cells, but that they may also be specialized to produce a greater variety of substances than just the acetylcholine or adrenaline and noradrenaline of nerve endings. Recently, five distinct staining reactions have been found among the neuro- secretory cells terminating in the sinus gland of a crab (§ 2.112; Potter, 1954), and it seems likely that eventually these will be found to be related to separate hormones. The occurrence of neurosecretory cells, which release hormones that have either kinetic or metabolic actions, is given with the other sources in Table 3. The last column shows the later sections of the book in which examples of these actions are des- cribed. A more detailed summary of the occurrence of neuro- secretion in invertebrates can be found elsewhere (Gabe, 1954). 2.111 Epistellar body of Cephalopoda In most octopods there is a small compact body on the outer surface of the stellate ganglion in the mantle cavity. In Eledone moschata it is yellow and about the size of a pin’s head. Micro- scopic examination shows this epistellar body to contain a group of neurosecretory cells (Fig. 2-1d) with their axons converging on a central cavity, which contains secreted granules in a homo- geneous ground substance. The granules presumably release a hormone into the adjacent artery; but its ability to stimulate muscle tone in the mantle (§ 3.12) has only been postulated from extirpation experiments. 22 SOURCES OF KINETIC AND METABOLIC HORMONES TABLE 3. ECTODERMAL SOURCES OF KINETIC AND METABOLIC HORMONES SOURCE OF HORMONE | STORE OF HORMONE TYPE OF SECTION (OR NAME) ACTION* NO. 2.11 CELLS FROM THE NERVOUS SYSTEM 2.111 Epistellar body of Cephalopoda Stellate ganglion |Epistellar body K 342 2.112 Neurosecretory systems of Crustacea Ganglionic-X-organ Sinus gland K S22 and brain ” ” ” ” K 3.223 » ” » » M 5.112 ” ” ” ” M Del oe ” ” ” ” M a a 1 ” ” ” »” M 5.321 ” ” ” ” M 5.422 % se Hanstr6ém’s sensory EK 4.211 pore organ Commissures ? K 3.223 Pericardial organs ? K 3.141 Eyestalk tip ? M 5.224 2.113 Neurosecretory systems and glands of Insecta Protocerebrum Corpora cardiaca EK 4.211 * » ? M 5328 Suboesophageal — K 3.221 ganglion i — M oye ip Corpora cardiaca — K ey |e | 2.114 Neurosecretory systems and glands of Vertebrata Paraventricular and Neurohypophysis K 3.114 supra-optic nuclei - os M 5.342 of hypothalamus ” ” ” ” M Spey Supra-renal tissue (Adrenaline) K 3112 Adrenal medulla (Adrenaline) K 3.492 - % K 3.116 2AD GLANDS FROM ECTODERMAL EPITHELIUM 2.121 Salivary glands of Cephalopoda Salivary glands \((T'yramine) K ee 2.122 Corpora allata of rb Corpora allata — M 5-11 2.123 Adenohypophysis = Ee, Pars distalis (TSH)t EK 4.221 7. ~ ray EK 4.223 a i (ACTH) EK 4.231 “ x (ICSH) EK 4.232 i a (LSH) EK 4.232 Pars intermedia (B or MSH) K 3.223 _ Pars tuberalis (W) K 3.223 * K = kinetic. EK = endocrinokinetic. M = metabolic. + See glossary. Fic. 2-1 (a) ( For legend see over) : Mit (g), Fic. 2-1 (g) Fic. 2-1. Cells derived from the nervous system. (a) ‘Typical motor nerve cell with branched dendrites (de), cell body with Nissl bodies (n.b.) in cytoplasm, nucleus (nu) with nucleolus, and long axon (ax) branching to motor end-plates (mot.e.p.) on muscle fibres (m). (b—d) Neurosecretory cells with stainable granules (gr): (0) with dendrites, from supraoptic nucleus of dog (after Scharrer and Scharrer, 19545); (c) without dendrites, the blunt axon is swollen with secretory granules that pass to a blood vessel (b.v.) from gang- lionic-X-organ of crab, Sesarma (after Enami, 1951). (d) cell with shorter axon, from epistellar body of Eledone (after Young, 1936). All drawn roughly to upper scale. (e) and (f) Gland cells with secre- tory granules (s.g.) but no histological characters of neurons (drawn to lower scale): (e) cells from corpus cardiacum of beetle, Hydrous (cf. Fig. 2-9 after de Lerma, 1956); (f) cells round blood space (b.v.) from adrenal medulla of a tetrapod, with “chromaffin” gran- ules (c.g.) (after Maximow and Bloom, 1942). The differences in quantity of secretion are not characteristic of these cells, but indicate different phases of secretion (cf. Fig. 2-2). (g) Electron micrograph of a highly enlarged section across the axon of a neurosecretory cell from the neurohypophysis of a cat, showing fine granules (Gr) 0.1 to 0.3 1 in diameter, and mitochondria (Mit) that are larger (from Bargmann, 1958). é “if 4 4 Rs , > ¥ er *, aoe se 4 we « J ve ae ‘ e°, a. or st B D om Fic. 2-2. Neurosecretory cells, with axons cut short, from the suboesophageal ganglion of the cockroach, Leuwcophaea maderae. The differences probably correspond to phases in a secretory cycle: (A) laying down fine granules in the cell-periphery in the position of Nissl bodies at the onset of secretion; (B) abundant larger granules passing into the swollen axon above; (C) empty vacuoles replacing secretory droplets; (D) an almost empty cell body, with only a few granules left at the base of the axon. ‘The cycle then probably starts again at (A); but the sequence has not yet been proved. Cells, < 320, fixed in Zenker-formol and stained in Masson (from Scharrer and Scharrer, 1954). Fic. 2-3. Living median neurosecretory cells in the protocere- brum of the blowfly, Calliphora, photographed by dark-ground illumination. The secretory granules in the cells are highly refrac- tile and therefore show as bright areas filling the cell bodies; only the nuclei remain dark. Other brain cells without granules can be seen faintly outlined in the background (from E. Thomsen, 1954). Fic. 2-14. Endocrine cells from the endoderm and the mesoderm of mammals. (a) Follicle of the THyRorID gland, enclosing colloidal store (col.) of secreted diiodotyrosine; this precursor substance is later reabsorbed by the cells, converted to thyroxine and passed to the blood vessel (b.v.) through the outer cell surface (cf. Figs. 4-7 and 4-8). (b) Three kinds of cells in an islet of Langerhans stained with Mallory-azan: a cells that secrete GLUCAGON and have coarse granules that stain red; pale f cells that secrete INSULIN and have fine granules that stain red; D cells that have no known function and stain blue; b.v., blood vessel, c., capsule of con- nective tissue. (c) Mesodermal cells, forming part of ADRENAL COR- TEX; they are shown in close contact with each other, and with capillaries of the blood supply (cf. Fig. 2-15). The vacuolated cytoplasm (v.c.) is shown after removal of all fat, which is abundant in living cortical cells (after Maximow and Bloom, 1942 and Pauly, O57): As fone ; | i kia ni § 2.112 SECRETORY CELLS FROM THE NERVOUS SYSTEM 23 The epistellar body is of interest, not only because the term neurosecretory was first used in this country to describe its cells but also because it provided some of the earliest evidence for the conversion of neurons into secreting cells in an invertebrate (Young, 1936). The corresponding nerve cells in the stellate ganglia of decapod cephalopods still retain the form of neurons, but have their axons fused to form giant fibres to the mantle muscles. They are scattered in Sepia, but collected together in the same position as the epistellar body in Loligo (Fig. 2-4). These neurons, as well as the neurosecretory cells of the epistellar body, are innervated by axons coming from the pedal ganglion of the brain. Fic. 2-4. The stellate ganglia of Sepia (A) and Loligo (B), and the epistellar body of Eledone (C). In (A) the giant fibres (g.f.) and the stellate nerve (st.n.) arise from nerve cells that are scattered throughout the ganglion; in (B) they are collected into a lobe (g.f.l.). In the octopus (C) there are no giant fibres, but instead there are neurosecretory cells (n.s.) whose axons end blindly in the central space of the epistellar body (ep.); their secretion passes in the blood to the mantle muscles. A nerve (n.) to the epistellar body replaces the preganglionic fibres (p.g.f.) to the nerves in (A) and (B), and presumably controls the release of secretion in (C). (From Young, 1936). 2.112 Neurosecretory systems of Crustacea There are four neurosecretory systems in crustaceans. ‘Two of these have their nucleated cell bodies in the brain and in the optic 24 SOURCES OF KINETIC AND METABOLIC HORMONES lobes; the ends of their secreting axons are aggregated into distinct storage-and-release organs, known as the sinus gland and Hanstrém’s sensory pore organ respectively. A third group of neurosecretory cell bodies has been found in Granules Post. com— (a) Fic. 2-5. Neurosecretory cells (n.c.) in decapod Crustacea. (a) Brain, connectives and fused ventral ganglia of a crab, Gecarcinus, and (6) eyestalk of Gecarcinus, cut open dorsally to expose the § 2.112 SECRETORY CELLS FROM THE NERVOUS SYSTEM 25 the commissures and connectives arising from the brain, and extracts showing hormonal activity have been obtained from them (§ 3.223); but the natural point of release for their secretions into the blood stream is uncertain. It may be the post-commissure organs (Knowles, 1953). A fourth system has been found in the pericardial organs of various decapod crabs and of Stomatopoda; these also yield an active extract (§ 3.11), but a natural secretion has not been fully established. Brain, ganglionic-X-organ and sinus gland Some of the neurosecretory cells, that release their secretion in the SINUS GLAND, have their cell bodies in the PROTOCEREBRUM of the brain and others in its extension into the TERMINAL MEDULLA of the optic lobe. The details vary for different orders and species. For instance, in the crab, Gecarcinus, the cells extend from the protocerebrum into the deutero- and tritocerebral lobes of the brain (Fig. 2-5a-b; Bliss and Welsh, 1952). _ The relatively large group of neurosecretory cells (Fig. 2-7) that lies in the terminal medulla is best known as the GANGLIONIC- X-ORGAN (pars ganglionaris X organi of Carlisle, 1953). It is important to remember the positions of these neuro- secretory cells in experimental work. Removal of the whole eyestalk removes the ganglionic-X-organ but leaves the cell bodies outer part of the optic stalk and nerves to the retina (R) (after Bliss and Welsh, 1952); (c) post-commissure organ (Post. com.) of a prawn, Leander, attached to tritocerebral commissure and having neurosecretory granules, (after Knowles, 1955). Protocere- brum (PRO), carrying optic lobes (OP), and deuterocerebrum (DEU), both joined by internal commissures, not shown; tritocere- brum (TRI) with its commissure (Tr. com.) behind oesophagus (Oes); circumoesophageal connectives (Conn.) join brain longi- tudinally to fused suboesophageal (SUBOES) and thoracic ganglia (THOR). Eyestalk contains continuation of optic lobe ending in terminal medulla (MT) with ganglionic-X-organ (GXO), and neurosecretory axons that end in sinus gland (SG) on internal medulla (MI); the external medulla (ME) also has some neuro- secretory axons. Nerve roots to antennae (Ai and Ai), viscera (Vis), mandible (Mdb), maxillae (Mx i and Mx ii) and thoracic appendages (Th ij, etc.). 26 SOURCES OF KINETIC AND METABOLIC HORMONES in the brain still in place; removal of the sinus gland only (by means of a minute punch, like an apple corer; Kleinholz, 1947) leaves both sources undamaged and able to continue their secretion. Dorsal Ventral R SP HSPO ON SN X-SP Fic. 2-6. Eyestalk of prawn, Lysmata, cut open in the vertical plane (cf. Fig. 2-5). Some neurosecretory cells in the ganglionic- X-organ (GXO) in the terminal medulla (MT) have axons which pass in a bundle (X—SG) to the stNus GLAND (SG) on the dorsal surface of the external medulla (ME); others have axons (X—SP) that pass ventrally to HANSTROM’sS SENSORY PORE ORGAN (HSPO) and end in “‘onion bodies’? (ON). Both sinus gland and sensory pore organ also have axons (B—SG and B-SP respectively) from neurosecretory cells in the brain. ‘The sensory pore retains some sense cells (SP) connected by a sensory nerve (SN) to the brain (from Carlisle, 1953). In the Malacostraca with long eyestalks, the SINUS GLAND is usually on the dorsal surface of the optic ganglion, either on the internal or external medulla (Figs. 2-5) and 2-6). It is largely com- posed of an aggregate of swollen ends of the long neurosecretory cell axons from the brain and the ganglionic-X-organ; they con- § 2.112 SECRETORY CELLS FROM THE NERVOUS SYSTEM 27 verge on a half open cavity in communication with the adjacent blood sinus. In the blue crab, Callinectes, there is evidence that the sinus gland is composed of as many as five groups of axon endings, each with a distinctive staining reaction, which can be traced back along the axon (Potter, 1954). It seems highly prob- able that these groups are responsible for secreting and releasing most of the specific chemicals with different hormonal actions which can be found by experiment in the sinus gland. 4e® SHLWHOO 0999 YOY S5Oer8 ASO ( OP SOS orNK \\e NPMT’ Peal SAN “| O ) © \\ 0 O=\\'\ \o0 A\ 0-0 B Op} © Oge:= \ BS 6.2 ¥o/5\ oO © RC Fic. 2-7. Neurosecretory cells (NSC) in the ganglionic-X-organ of a crab, Sesarma. Stained granules of secretion (G) pass down the axons (SGN) to the sinus gland. Some cells (CB) have few granules, others (RC) are almost depleted of granules. Small ganglion cells and nerve fibres (NPMT) of the surrounding ter- minal medulla are also shown (from Enami, 1951). 28 SOURCES OF KINETIC AND METABOLIC HORMONES It has been suggested that, in addition to acting as a storage- and-release organ for the neurosecretion, the walls of the sinus gland may include some secretory cells, but of this there is no clear evidence (Knowles and Carlisle, 1956). It seems more likely that active hormones are being set free here from the inactive carrier substance that travels down the axons. In the sessile-eyed Malacostraca, such as the Isopoda, both the ganglionic-X-organ and the sinus gland itself lie within the head capsule (Amar, 1948). Hanstrém’s sensory pore organ The sensory pore organ, like the sinus gland, is the storage-and- release organ for neurosecretory cells some of which are situated within the BRAIN and others in the GANGLIONIC-X-ORGAN. All the axons have their endings swollen into characteristic onion- shaped bodies in HANSTROM’S SENSORY PORE ORGAN, situated on the ventral surface of the eyestalk in Malacostraca (Fig. 2-6). In addition to these axon endings and to the sensory cells, which give their name to the organ, there are some small secretory cells confined within the organ, at least in Lysmata (Carlisle and Passano, 1953). This structure was originally called the ‘““X-organ” and was identified in several species by Hanstrém (1939) and his pupils; but since then, many workers have used the name X-organ for the neurosecretory cell bodies located in the terminal medulla. The modified name of ganglionic-X-organ is here used for this latter group of cells, from which in fact axons run to both the sinus gland and to Hanstrém’s organ (Knowles and Carlisle, 1956). Commissures and connectives Active extracts have been obtained from many other parts of the crustacean nervous system besides the supraoesophageal ““brain’’; but as yet there is little indication of where any natural hormones, corresponding in their actions to these extracts, may be released into the blood stream. The TRITOCEREBRAL (or antennal) COMMISSURE, which is a cross connection passing below the oesophagus between the tritocerebral parts of the brain (Fig. 2-5a), is particularly rich in chromactivat- § 2.113. SECRETORY CELLS FROM THE NERVOUS SYSTEM 29 ing materials (§ 3.2) in many Decapoda. In the prawns, Leander and Penaeus, it has been shown (Knowles, 1953 and 1954) that the attached POST-COMMISSURE ORGANS are the swollen bases of nerves, some motor fibres of which pass to dorsoventral muscles; the two nerves have a cross connection and contain many neurosecretory fibres and secretory droplets and yield an active extract. Some of the cell bodies are in the commissure (Fig. 2-5c). The CIRCUMOESOPHAGEAL CONNECTIVES and the thoracic and abdominal ganglia also yield extracts of varying activity, particu- larly in forms such as Palaemonetes, in which the ganglia are not fused in one mass. Pericardial organs In Decapoda and Stomatopoda there are some rather unusual neurosecretory axons in the pericardium. They are supported by larger nerve trunks and end blindly in networks of very fine branches spread over the venous openings from the gills; they are therefore exposed to the blood stream and appear to secrete into it one or more chemicals that increase the rate of the heart beat (§3.111). The position of the cell bodies of these PERICARDIAL ORGANS has not yet been found; but recent evidence indicates the presence of very fine granules in the fibrils, and experimental evidence for their secretory activity is good. Similar structures may also be present in Isopoda (Alexandrowicz, 1953). 2.113 Neurosecretory systems and glands of Insecta The nervous system of Insecta has given rise to both neuro- secretory cells and simple secretory cells which have lost any morphological signs of their nervous origin. The former occur within the central nervous system, mainly in the suboesophageal ganglion and the brain, and the latter in the corpora cardiaca. The tritocerebral commissures and the circumoesophageal connectives, though similar in form to those of crustaceans, have not been shown to yield active extracts in insects. Suboesophageal ganglion Neurosecretory cells have been identified microscopically in the SUBOESOPHAGEAL GANGLIA of several insects, including some 30 SOURCES OF KINETIC AND METABOLIC HORMONES Ephemeroptera (Arvy and Gabe, 1953) and Plecoptera, where their axons are connected anatomically to the ventral gland (Fig. 2-8). Experiments have shown that in higher insects the suboesophageal ganglion is the source of a kinetic chromactivating HEAD PROT Ocelli Ge xs OTHOR Lbr Mdb Mxi Mx.ii — Vent. gl Leg I Fic. 2-8. Central nervous system and sources of hormones in the head and prothorax of a hemimetabolous insect, in lateral view. ‘The nervous system, with optic lobes (OP) cut off, resembles that of the crab (Fig. 2-5) and is similarly lettered and named, except that Mx ii is here the labium, and ganglia of the ventral chain are § 2.113 SECRETORY CELLS FROM THE NERVOUS SYSTEM 31 hormone (§ 3.221) and of the metabolic diapause hormone (§ 5.1 12). In the cockroach, Leucophaea, these cells present a variety of appearances in fixed preparations (Fig. 2-2). These probably represent phases in secretion; but this has not been confirmed in the living animal. Neurosecretory cells of the brain Paired groups of neurosecretory cells are to be found in the brains of insects, as in crustaceans; but in most insects they appear to be confined to the PROTOCEREBRUM and not to extend into other parts of the brain or optic lobes. The neurosecretory cells usually lie in groups: the median neurosecretory cells (m.n.c., Fig. 2-3) lie anteriorly near the mid-line and other cells lie ventrally or laterally (I.n.c.). ‘The former are connected to the corpus cardi- acum of the opposite side by an internal nerve, and the latter by an external nerve (Fig. 2-8). Their axons lead into the CORPORA CARDIACA, where their secretion is stored (Fig. 2-9). The lateral neurosecretory cells may represent the frontal organs of Aptery- gota and may even be homologous with the cells of Hanstrém’s sensory pore organ (§ 2.112). It is interesting to note that these groups of neurosecretory cells secrete endocrinokinetic hormones that stimulate endocrine glands in both Crustacea and Insecta (§ 4.21; Scharrer and Scharrer, 1954 5). not fused. The stomatogastric, or visceral, nervous system arises from the stomodaeal ectoderm, and has paired nerves from the tritocerebrum, a median frontal ganglion (Fr. gang), a branch to the labrum (Lbr) and a recurrent nerve to the hypocerebral ganglion (Hy) and the paired ventricular ganglia on the gut (near Oes). HorMONES are secreted by median (m.n.c.), lateral (I.n.c.) and suboesophageal neurosecretory cells (s.n.c.); perhaps also by cells in paired corpora pedunculata (C.ped.). Secretions from these pass in two paired nerves to be stored in the CORPORA CARDIACA (CC), which arise from stomodaeal ectoderm (dashed arrow). Hormones are also secreted by two paired glands that arise as ectodermal invaginations: the CORPORA ALLATA (CA from Mx i), which migrate (dashed arrow) above the oesophagus, where they receive axons from the corpus cardiacum (cf. Fig. 2-9); and the VENTRAL GLANDS (Vent. gl. from Mx ii), which persist in primi- tive insects, but form prothoracic glands (Proth. gl.) in most other orders. (Based on two diagrams by Weber, 1949). 52 SOURCES OF KINETIC AND METABOLIC HORMONES Storage of the neurosecretion from the brain in the corpora cardiaca has been conclusively shown in the cockroach, Leuco- phaea, where the system 1s paired (B. Scharrer, 1952). Unilateral section of the axons from the median neurosecretory cells results <@1 gsc (a) Corpus cardiacum (b) Corpus allatum Fic. 2-9. Sections of parts of (a) the CORPUS CARDIACUM of a beetle, Hydrous piceus (after de Lerma, 1956), and (6) the coRPUS ALLATUM of a grasshopper, Melanoplus differentialis (after Mendes, 1948). Axons (ax.b) from neurosecretory cells in the brain carry granules; some end in swellings (s.ax), like Herring bodies, and release masses of neurosecretion (ns) in the corpus cardiacum. Cells in different phases of secretion (sc and sc’) release masses of granules (mg) that stain with phloxin, and may be the intrinsic secretion of the organ. Tracheoles (t) and non-secreting nerve cells (cn) also occur. Other axons (ax.c) pass to the cells of the corpus allatum, where their granules disappear. It has undifferentiated cells (uc), secreting (sc”) and giant secreting cells with polyploid nuclei (gsc) ; acidophil granules pass with some fluid into intracellular vacuoles and are then extruded (v). in the depletion of all stainable secretion from the corpus cardia- cum of that side. At the same time, increased quantities of the secretion appear in the axons proximal to the cut (Fig. 3-2). The corpora cardiaca are nearly always fused with the dorsal blood vessel or aorta, into which they presumably pass the stored secre- tion as required (Hanstrém, 1940). § 2.114 SECRETORY CELLS FROM THE NERVOUS SYSTEM 33 Gland cells of the corpora cardiaca The CORPORA CARDIACA are not only storage organs for neuro- secretion from the cerebrum but they are also endocrine organs in their own right. Together with the corpora allata (§ 2.122) they lie dorsal to the oesophagus and form the retrocerebral system (Fig. 2-8), the form of which varies in different insects; both organs may be paired, or one or both may be fused in the mid-line. Together with some nerve cells, connective tissue and tracheae, loosely packed secretory cells form the bulk of the corpora car- diaca and produce the intrinsic secretion of this organ (§ 3.111). Like the sympathetic cells of the hypocerebral ganglion, these cells arise from the stomodaeal ectoderm and are undoubtedly nervous in origin, although they have become so much specialized for secretion that they have lost most of the characters of neurons, notably the axons of typical neurosecretory cells (Figs. 2-le and 2-9a). ‘These cells are rich in ribonucleic acid and reveal a Golgi apparatus after suitable treatment. With haematoxylin chrome phloxin, their secretion stains quite distinctively from the neuro- secretion, the former having a greater afhnity for phloxin and the latter for haematoxylin (de Lerma, 1956). 2.114 Neurosecretory systems and glands of Vertebrata There are two main sources of hormones that are derived from the vertebrate nervous system. One is a set of neurosecretory cells in the hypothalamus of the brain with their storage-and-release organ (akin to the sinus gland of the Crustacea) in the neuro- hypophysis. The other is an aggregation of gland cells that are derived from sympathetic ganglia and form the suprarenal body of fish and the adrenal medulla of tetrapods. Neurosecretory cells of the hypothalamus and the neurohypophysis It is now well established that the hormones of the neuro- hypophysis (or posterior lobe of the pituitary body) are not secreted within that body but by neurosecretory cells in the hypothalamus of the brain. The cell bodies are grouped together in the preoptic nucleus in fish and amphibians and separated into two groups, the supraoptic and paraventricular nuclei, in reptiles, birds and D 34 SOURCES OF KINETIC AND METABOLIC HORMONES @ Nucleus paraventricularis Tractus supraopticohypophyseus Nucleus - supraopticus Pars tuberdlis Pars nervosa Pars distalis Pars intermedia Accumulation of neurosecretory material Plane of stalk section (b) Fic. 2-10. Diagram of the pituitary body of a dog, Canis; (a) before and (b) after the pituitary stalk has been cut. Neuro- secretory cells in the paraventricular and supraoptic nuclei of the hypothalamus of the brain pass secretory granules down their axons to the neural lobe of the neurohypophysis (pars nervosa), where they are stored in Herring bodies, or swollen axon endings, and thence pass into the surrounding blood vessels (Fig. 2-12). After cutting across the stalk, the neurosecretion accumulates in the proximal part of the axons and the supply previously accumulated in the neurohypophysis becomes depleted after a time and is not replenished. The pars distalis of the adenohypophysis (and pars tuberalis) lie in front of the pars nervosa, separated from it by the pars intermedia (from Scharrer and Scharrer, 1954a). § 2.114 SECRETORY CELLS FROM THE NERVOUS SYSTEM 35 mammals (Fig. 2-10; Scharrer and Scharrer, 1954a). Unlike most neurosecretory cells of the arthropods, those of the hypothalamus of vertebrates possess dendrites (Fig. 2-10). In fish and aquatic Urodela their axons are not well developed and their function is uncertain; but in all terrestrial vertebrates from the terrestrial Urodela and Anura upwards the axons pass down the infundibular stalk to end in an enlarged neural lobe (pars nervosa). ‘This becomes distinct from the median eminence at the base of the infundibulum and is not present in the lower forms (Fig. 2-11). In the course of evolution, the neural lobe has acquired an independent blood supply from the internal carotid arteries, forming a relatively rich vascular network (Fig. 2-12) with which the axons of the neurosecretory cells make contact by their swollen endings, called Herring bodies, similar in appearance to those of the comparable cells in the crustacean sinus gland. They can be shown by appropriate staining to be filled with neuro- secretory granules. In mammals the secretory granules first become visible in the embryo, where they appear to be carriers for the actual hormones released from the neurohypophysis. The relation between secretion and hormone formation has now been as clearly shown here as anywhere, although the proofs were obtained later than in the invertebrates. In any vertebrate from fish to mammals, section of the axons in the infundibular stalk results in accumulation of the secretion in the parts proximal to the cut and its depletion beyond; and the possibility of obtaining active extracts from the different parts of the system follows the same pattern. It has also been possible to grow cells from the supraoptic nucleus of a dog in tissue culture and to observe (and even to make a ciné film of) the secretory granules passing from the cell body down the axon. The neurohypophysis therefore acts as a storage-and-release organ for the hypothalamic secretion or secretions. ‘Two distinct hormones have been recognized, the antidiuretic hormone and oxytocin, but there does not seem to be any constant arrangement of the neurosecretory cells which produce them. In the dog, for instance, both substances can be obtained from both supraoptic and paraventricular nuclei, although the proportion of oxytocin to the antidiuretic fraction is always small; but in the camel, oxytocin 36 SOURCES OF KINETIC AND METABOLIC HORMONES Primary capillary net sm ~Vascular plexus Secondary capillary net =»> Portal vessels Cyclostome Typical Holostei Urodela Teleostei Elasmobranchii Lacertilia Fic. 2-11. Pituitary body in diagrammatic sagittal section with its circulation shown by thin arrows: a to d in fish, e to # in amphi- bians and reptiles. A.H., adenohypophysis, divided into pars distalis (P.D.), pars tuberalis (P.T.), and pars intermedia (P.I.) in tetrapods; N.H., neurohypophysis, divided into median eminence (M.E.) and neural lobe (N.L.) with separate circulation § 2.114 SECRETORY CELLS FROM THE NERVOUS SYSTEM 37 is the more abundant in the paraventricular nucleus (Van Dyke Adamsons and Engel, 1957). These hormones are concerned with counteracting thirst and the desiccation that is the main risk accompanying the migration from water to land. The antidiuretic hormone facilitates reabsorp- tion of water from the urine (§ 5.322) and oxytocin increases the excretion of Na* and Cl7 (§ 5.312). It is therefore understandable that the neural lobe, where these hormones can be quickly released into the blood, should be best developed in terrestrial animals (Harris, 1955). The correlation has been confirmed by the observa- tion that natural thirst, or an equivalent state caused by injecting rats with saline, is followed by depletion of the secretory granules within a few minutes, to be slowly replaced in a day or so after giving the animals water to drink. Depletion of secretory granules in fish has been observed in response to immersion in hypertonic sea water, which would have the same effect as desiccation (Arvy, 1957), Gland cells of the suprarenal tissue and adrenal medulla The peripheral neurons of the sympathetic nervous system all secrete adrenaline, or noradrenaline, at their motor nerve endings. In most vertebrates some of these neurons become modified to secrete relatively enormous amounts of either or both of these substances; at the same time the cells lose all histological resemblance to ganglion cells (Fig. 2-1f). Their origin and func- tion is nevertheless the same as that of neurosecretory cells; but opinion is divided as to whether they should be regarded as such (Welsh, 1955). They are often referred to as “‘chromafhn”’ cells, because the contained adrenaline gives a characteristic olive-brown colour with any chromic salts used either as fixative or stain. Staining shows that the adrenaline is secreted by the cytoplasm as a mass of very fine granules. In fish, where these cells form the “suprarenal” tissue, they in most tetrapods. Heavy arrows indicate the presence of portal veins between primary venous plexus in median eminence and secondary plexus in pars distalis (cf. Fig. 2-12) in fully terrestrial forms. O.C., optic chiasma; S.V., saccus vasculosus ; V.L.,. ventral lobe (from Green, 1951). 38 SOURCES OF KINETIC AND METABOLIC HORMONES remain in their original paired positions and are innervated by the preganglionic fibres of the visceral motor or sympathetic system. They secrete mainly noradrenaline. From Amphibia to Mammalia this tissue, having migrated towards the anterior ends of the kidneys, becomes progressively enveloped in the interrenal or cortical tissue, derived from coelomic epithelium (§ 2.311). The chromaffin tissue forms the adrenal medulla, or core, and secretes mostly adrenaline in mammals. Together, the cortex and medulla form the complex adrenal gland.* The cells of the medulla are innervated in. the same way as in fish. Some unmodified ganglionic cells may be seen among them; but the chief characteristic of the tissue is the net- work of blood spaces with which every cell is in contact and into which their secretion can be passed with great rapidity in response to nervous stimulation in an emergency (§ 3.11). 2.12 ENDOCRINE GLANDS DERIVED FROM ECTODERMAL EPITHELIUM A number of endocrine glands are derived from ectodermal epithelium, without having any apparent connection with the nervous system. These seem to be more frequent in invertebrates than in vertebrates, and include the Y-organ and the prothoracic glands, which are the main sources of the morphogenetic moulting hormones of Arthropoda (§ 4.21 and Part II). The salivary glands of Cephalopoda may be included here (§ 2.121), although it is doubtful if their secretion is a true hormone. The corpora allata of Insecta (§ 2.122) and the adenohypophysis of Vertebrata (§ 2.123) are both important ectodermal sources of kinetic and metabolic hormones. 2.121 Salivary glands of Cephalopoda Salivary glands of cephalopods are primarily used for the external secretion of tyramine, a poison for immobilizing the prey; * Unfortunately, in medical terminology, this compound gland is usually referred to as the “‘suprarenal’’, from its position ‘“‘above’’ the kidney in the upright posture. It must not be confused with the supra- renal of fish, which is homologous with the medulla only. § 2.122 ENDOCRINE GLANDS FROM ECTODERMAL EPITHELIUM 39 but this substance also passes into the blood, to affect the chrom tophore muscles (§ 3.21), probably by an indirect action. The glands, of which there are two in decapod and four in octopod cephalopods, arise from the stomodaeal ectoderm, with which they retain their connection as a duct to the mouth. It is the posterior (or dorsal) pair which secretes tyramine in Eledone moschata and in the two species of Octopus which have been investigated (Bacq and Ghiretti, 1951). a- 2.122 Corpora allata of Insecta The CORPORA ALLATA are endocrine glands, the cells of which arise in development as a pair of small ventrolateral invaginations near the base of the first maxilla (Fig. 2-8). Thence the tissue migrates inwards to lie between the oesophagus and the aorta; it may remain paired, or the two parts may fuse more or less com- pletely. Each part is supplied with neurosecretory axons passing on from the corpora cardiaca, which lie just in front of them (Fig. 2-9). There is evidence that these axons must remain intact if they are to control secretion by the corpora allata, as though their action were either nervous or due to a neurohormone diffusing from cell to cell rather than being carried in the circulation. The corpora allata are also connected with the stomatogastric system by nerves from the hypocerebral ganglion. The densely packed cells of the corpora allata show cyclic phases of secretion and multiplication, coinciding with their cyclic activity in controlling the “juvenile” character of nymphal and larval moults (Part II, § 3). A new phase of activity starts very soon after each moult is completed: at first the gland is small and the cells are all alike; then there is a burst of mitotic activity followed by an increase in size of some cells, leading to increase in gland size. The larger cells, which have much larger nuclei than the undiffer- entiated cells, then begin to form their secretion. This appears first as granules in the cytoplasm; it then accumulates in vacuoles, which can later be seen to lie outside the cells in intercellular spaces. From this position the secretion, or the hormone released from it, presumably passes into the blood stream at the critical period for controlling the next moult. Some of the secreting cells 40 SOURCES OF KINETIC AND METABOLIC HORMONES become polyploid by division of their nuclei without cell division. They form giant cells (g.s.c., Fig. 2-9). When secretion is com- pleted, the gland returns to the initial undifferentiated state shortly before the actual moult (Mendes, 1948). The glands also secrete metabolic (§ 5.11) and morphogenetic hormones (Part II, § 3) during the adult life of the insect, when the histology of secretion appears to be the same as that in younger stages, and gives no indication of the hormones being different at different ages. 2.123 Adenohypophysis of Vertebrata The adenohypophysis, or the anterior lobe of the pituitary body, is the only endocrine gland which arises from non-nervous ecto- derm in vertebrates. Its origin is the hypophysis, an upgrowth from the roof of the stomodaeum. It meets the brain and induces a downgrowth from the hypothalamus or floor of the diencephalon; together they form the pituitary body (Fig. 2-13). Throughout the vertebrates the contribution from the nervous system forms the neurohypophysis which has already been described (§ 2.114). The subdivisions of the ectodermal ADENOHYPOPHYSIS are difficult to homologize in different groups. It has recently been recommended (Pickford and Atz, 1957) that the noncommittal terms pro-, meso- and meta-adenohypophysis should be used for fish and that these should only be very tentatively homologized with the three parts recognizable in most tetrapods, namely the PARS TUBERALIS, the PARS DISTALIS and the PARS INTERMEDIA (Fig. 2-10).* Of these, the pars distalis is usually the largest and appears to be the main source of hormones. It lies in front of the original hypophysial cavity, or cleft, if this persists, whereas the pars intermedia forms behind the cleft and often comes into close contact with the adjacent neural lobe. The pars tuberalis surrounds the infundibular stalk. The shapes and relative sizes of these parts * Unfortunately, the old nomenclature is still to be found in many books; in this, the pars intermedia, although formed from the same tissue as the rest of the adenohypophysis or “‘anterior lobe of the pituitary’’, is included with the pars nervosa in the so-called “posterior lobe of the pituitary”; this is due to its morphological position in some mammals. Other writers use posterior lobe as synonymous with the neural lobe, or pars nervosa, only (Fig. 2-10). The names used here follow those recom- mended by the International Congress of Anatomists (Woerdeman, 1957). § 2.123 ENDOCRINE GLANDS FROM ECTODERMAL EPITHELIUM 41 vary greatly in different animals (Fig. 2-11). The pars distalis becomes increasingly important in land animals, and especially in mammals, whereas the pars intermedia decreases in size and may even be wholly lacking, as in the chick and whale. The blood supply of the pituitary also shows important differ- ences in different classes of vertebrates (Fig. 2-11). In fish, hypophysial arteries from the internal carotids break up into a vascular plexus, which penetrates the whole organ and eventually drains into venous sinuses under the skull. (The saccus vasculosus may have a separate supply; but it is not part of the endocrine gland.) In Urodela, the arterioles pass first to the pars tuberalis and thence to the capillary plexus of the pars distalis and pars nervosa. The pars intermedia hardly has any share in the blood supply. In the Anura and amniotes, the plexus in the pars tuberalis penetrates into the median eminence and then the vessels join up to form a number of portal veins which break up into a secondary venous plexus in the pars distalis (Fig. 2-11 g and h). It might be thought that this portal system was less important in mammals than in lower forms, since in them alone the pars distalis acquires a direct arterial blood supply from the internal carotids (Fig. 2-12); but it may allow the passage of CRF (§ 4.323). In all land animals the newly developed neural lobe of their neurohypophysis also acquires an independent arterial supply from the same source. These vessels from the neural lobe, together with all those from the pars distalis, drain into the same venous sinus and eventually join the internal jugular veins. Nerve axons to the pituitary fall into two distinct categories: those of neurosecretory cells, and those of sympathetic nerves. The neurosecretory cells are confined to the neurohypophysis, and most of them end in the neural lobe; but some of them make contact with the primary venous plexus in the median eminence. In this way, secretions from the latter can pass into the portal circulation and so to the pars distalis. How much they do so, and whether they affect the rates of hormone secretion from the adeno- hypophysis, is still a matter for discussion. The vasomotor fibres of the sympathetic nerves follow the course of the hypophysial arteries (Fig. 2-12); they can therefore 42 SOURCES OF KINETIC AND METABOLIC HORMONES | | i} | | Fic. 2-12. Diagrammatic sagittal section of the pituitary gland and hypothalamus of a rabbit, Oryctolagus, with the circulation super- imposed and simplified. The neurosecretory cells (NS! and NS?) have been disproportionately magnified to show their axons con- necting with the primary plexus of veins (VP) in the median eminence (ME) and (VI) in the neural lobe (NL). The only other nerves are sympathetic vasomotor fibres (S!, S? and S%). Blood comes from the internal carotids (IC) by hypophysial arteries; the upper right (HY?) takes an independent supply to the NEURAL LOBE (NL), which is drained by the main vein (V). The PARs INTERMEDIA (PIN) has practically no circulation. The upper left hypophysial artery (HY*) supplies the primary venous plexus in § 2.123 ENDOCRINE GLANDS FROM ECTODERMAL EPITHELIUM 43 control the blood supply to the neural lobe and the pars distalis, but do not appear to make any contact with the secretory cells. There are no nerves to the pars intermedia in mammals, though secretions from this part appear to be under nerve control in fish and amphibia (§ 4.3). The PARS DISTALIS of the adenohypophysis contains at least three distinct types of cells: chromophobe (gamma) cells which have no stainable granules in their cytoplasm and seem to have no secretory function, though they may give rise to one or both of the secreting types; basophil cells, which contain secretory granules of glycoprotein that stain blue by the Mallory or Azan trichrome methods, and acidophil cells, which contain phospholipid granules that stain selectively by an acid haematin method. The last two types can also be distinguished by staining with safranine-acid violet (cf. Maximow and Bloom, 1942, Fig. 261.-2) and show changes which are clearly associated with the secretory activity of the gland. For some time it was claimed that only these two types of secreting cells could be identified histologically although the gland was known to secrete six or seven distinct hormones; but recently more sensitive tests have been applied and tentative subdivisions of the two types have been proposed (‘Table 4); further details have recently been summarized by Pickford and tzZ{ 1957). The PARS TUBERALIs has not been studied so fully but appears to consist of columns of cells separated by blood spaces, and to be closely similar to the adenohypophysis in appearance. A secretory activity has only been claimed for it in some fish and amphibians [§5-23). The PARS INTERMEDIA may consist of cells with basophil granules and non-granular cells that form follicles filled with a colloid, similar in appearance to that in the thyroid gland but containing the PARS TUBERALIS (PT), from which loops penetrate deeply into the median eminence and can receive neurosecretions. ‘These vessels join again to give the hypophysial portal system (HP) which breaks up into the secondary venous plexus (VS) in the PARS DISTALIS (PD) of the adenohypophysis. ‘This part also receives blood directly from the lower hypophysial arteries (HY*). (Original, based on Scharrer and Scharrer, 1954a, and Harris, 1955). 44 SOURCES OF KINETIC AND METABOLIC HORMONES TABLE 4. CELLS IN THE PARS DISTALIS OF THE ADENOHYPOPHYSIS | | SECTION CELLS STAIN SECRETION NO. Basophil cells beta PAS} and aldehyde fuchsin ‘TSH 4.221 delta PAS only, peripheral ? FSH 4.232 rs luge sacentral ? LA 4.232 ? — TCS 7? 4.232 Peden ceils alpha Orange G | STH 4.223 epsilon Fuchsin LSH 4.232 ? a ACTH 4.231 * See glossary. + PAS = Periodic acid—Schiff method, with which these cells give a positive reaction. no iodine. This region is well known to secrete a melanophore dispersing hormone, intermedin, in many fish and most amphi- bians, but not in Agnatha. It has also been claimed that this lobe may act as a storage-and-release centre for adrenocorticotrophin, ACTH (Miualhe-Voloss, 1955). The claim may be due to the chemical identity of intermedin with a large part of the chain molecule that makes up ACTH. 2.2 ENDODERMAL SOURCES IN VERTEBRATA Hormones secreted from endodermal sources have so far only been recognized in vertebrates. They fall into two categories: isolated cells in the stomach and intestinal mucosa (§ 2.21) and well-developed endocrine glands in the pharynx and pancreas (§ 2.22). The former secrete kinetic hormones which control gut muscles and the secretion of digestive enzymes from exocrine glands; the latter secrete various metabolic hormones (Table 5). 2321. ISOLATED: CELRLSMN, THEIEUT A large number of hormones can be located in the stomach and intestine of mammals; some of these also occur in birds, but they are not known to be active in cold-blooded vertebrates. § 2:21 ISOLATED CELLS IN THE GUT 45 Since no recognizable endocrine glands or groups of secreting cells have been found in the GuT mucosa in those regions from which the gastrointestinal hormones, such as GASTRIN and sKc- RETIN, can be extracted (§ 4.11), it must be concluded that all these hormones arise from isolated cells (Table 5). So far, however, these cells have not been found, despite intensive search; it is possible that they may not be histologically distinct from other cells in the gut lining, such as those secreting mucus, but it seems more likely that they will eventually be revealed by more sensitive or selective staining techniques. It has been suggested that SECRETIN may be produced in certain ‘‘argentaffine”’ cells, which can be stained with silver; but this seems unlikely since similar cells are abundant in the vermiform appendix from which no secretin can be extracted (Grossman, 1950). The origin of the secretory cells certainly deserves further investigation. The clues that would seem to be the most worth following are the facts that (1) the action of this whole group of hormones is kinetic, (2) all other kinetic hormones come either from modified nerve cells or at least from the ectoderm and (3) the action of these hormones in stimulating the secretion of stomach, pancreas and intestinal glands is carried out in lower vertebrates by the parasympathetic nerves in the vagus. The parasympathetic nerves form part of the general visceral motor system and like the sympathetic nerves have peripheral ganglia, connected to the brain by preganglionic fibres. The latter are of central nervous origin; but there is considerable doubt as to the origin of the peripheral cells, though the neural crest has been plausibly postulated. If so, crest cells might be expected to migrate to the intestine from the cranial region to form ganglia; and it seems possible that some of them might also form argentaffine cells, and others secretory cells. Although there is evidence that the argentaffine cells of the gut can differentiate in grafts of chick intestinal epithelium, even if this is separated from the embryo before any trunk neural crest material is formed (Van Campenhout, 1946), it is not so clear that cranial neural crest cells were excluded in these experiments. On the other hand, there is one other curious feature about all the hormones secreted by the gut wall in mammals: unlike kinetic hormones derived from neurosecretory cells, they are not secreted 46 SOURCES OF KINETIC AND METABOLIC HORMONES in response to nervous stimulation, but always depend upon a direct stimulus, either mechanical or chemical (§ 4.321). The positions from which the various hormones have been found to be secreted in the gut are summarized in Table 5. | TABLE 5. ENDODERMAL SOURCES OF KINETIC AND METABOLIC CLASS HORMONES IN VERTEBRATA SOURCE OF HORMONE 2.21 ISOLATED ‘CELLS IN “THE ‘CUT Mammalia Stomach Duodenum ” > J 2 ” | Intestine ”” 2.22 ENDODERMAL ENDOCRINE GLANDS 2.221 Glands of the pharynx Agnatha to Mammalia Teleostet Tetrapoda ) 2.222 Gland Agnatha to Mammalia Aves and some Mammalia * K = kinetic. Thyroid Ultimobranchial body Parathyroid > cells in the pancreas Islets of Langerhans NAME OF HORMONE Gastrin Cholecystokinin Secretin Pancreozymin Duocrinin Enterocrinin Enterogastrone Thyroxine Parathormone bP) ” Insulin Glucagon M = metabolic. TYPE OF ACTION AAAAAAAM 2.22 ENDODERMAL ENDOCRINE GLANDS J | Recognizable endocrine gland cells are derived from the gut epithelium in two regions: the pharynx, where there are at least ect ENDODERMAL ENDOCRINE GLANDS 47 two groups (§ 2.221), and the pancreas, where they form the islets of Langerhans (§ 2.222). These glands all secrete metabolic hormones, and their structure has long been known; it is well described in most text-books of vertebrate anatomy, embryology and histology, and little need be said here, except to emphasize the fact that they can be identified in most classes of vertebrates and are not confined to the warm-blooded forms, like the sources of the gastrointestinal hormones. 2.221 Glands of the pharynx These are the THYROID and PARATHYROID GLANDS and the ULTIMOBRANCHIAL BODIES, which are the homologues of the para- thyroids (‘Table 5). ‘The thymus glands also arise here (Fig. 2-13); but their endocrine nature is uncertain. It will be considered in relation to growth (Part II, § 3). Thyroid glands In Agnatha, the endostyle in the floor of the larval pharynx can accumulate iodine, and even synthesize THYROXINE, albeit in small quantities, even before its transformation to the adult thyroid gland. The endostyle of the amphioxus, Branchiostoma, also accumulates iodine; but it does not appear to synthesize thyroxine (Barrington, 1958). It is then possible to argue either that the endostyles of the Protochordates and the Agnatha are homologous with each other and with the thyroid gland, because the former are structurally similar although only the two latter have acquired the ability to synthesize thyroxine, or that a true homology should be marked by similar chemical activities, and is in this case limited to the vertebrates. The thyroid gland retains its position as a single median organ in the floor of the pharynx in lower vertebrates (Fig. 2-13) where the gland itself may be diffuse, as in many teleost fish, or relatively compact and enclosed in a capsule of connective tissue, as in Elasmobranchii and a very few Teleostei, such as Pseudoscarus, from which it can therefore be relatively easily removed. In tetrapods, in which the gills are lost, the gland often becomes paired and may even be further subdivided; but each part is 48 compact and encapsuled. It receives a rich blood supply from the SOURCES OF KINETIC AND METABOLIC HORMONES fn, GAL LLL Fic. 2-13. Diagrammatic sagittal half of the head and pharynx of a tadpole, Rana. An early stage in development of the brain and stomodaeum (St) shows the ADENOHYPOPHYSIS (Hyp) growing up to meet the infundibulum (Inf), or NEUROHYPOPHYSIS, from the floor of the fore brain (F); together they form the pituitary body. Optic chiasma (OC), pineal organ (Pin), spinal cord (CNS) and notochord (Nch). The pharynx, leading to oesophagus (Oes), is shown at a later stage; I to VI, visceral arches; III to VI with branchial arteries running behind gill slits to dorsal aorta (DA); THYROID (Th) is mid-ventral; a series of ventrolateral epithelial thickenings form the carotid gland (C) on III, the PARATHYROIDS (P) on IV and V, and the ULTIMOBRANCHIAL BODY (U) on VI. Dorsolateral thickenings on II and III (and also on IV and V in other animals) form the THYMUS GLAND (‘Thym). carotids, and its nerve supply is mainly vasomotor. In most vertebrates the histological character of the gland is fairly constant. Its cells form a cubical epithelium surrounding spaces, which become filled with a colloidal material secreted § 2.222 ENDODERMAL ENDOCRINE GLANDS 49 into it as droplets from the cells (Figs. 2-14a and 4-7). This appears to be a precursor of the hormone, usually in the form of diiodo- tyrosine; it is later reabsorbed into the cells, converted to thyroxine, and passed into the blood as the hormone (Fig. 4-8). This process is described more fully in relation to its control by the thyrotrophic hormone, TSH (§ 4.221). Ultimobranchial bodies — These structures arise ventrolaterally from the epithelium of the last gill slit and may be seen in development to give rise to a pair of small glands. In fish they appear to replace the parathyroid glands; but in tetrapods they may be present in addition to them (Fig. 2-13). In function they appear to be similar to the para- thyroids (§ 5.4). Parathyroid glands These glands are serially homologous with the ultimobranchial bodies behind (and with the so-called carotid gland of Amphibia in front). They arise ventrolaterally from the epithelium lining the gill slits on the anterior surfaces of the fourth and fifth visceral arches (Fig. 2-13). The individual cells of the parathyroid glands form a densely packed mass amid ramifying blood vessels, and are not arranged in follicles. They may be of two kinds: the more numerous have clear cytoplasm and relatively large nuclei; the rest are acidophil with granular cytoplasm. The former are thought to be the main source of PARATHORMONE (§ 5.4). 2.222 Gland cells in the pancreas In most vertebrates, groups of endocrine cells occur among the exocrine cells (secreting enzymes and alkali) in the pancreas; they form the ISLETS OF LANGERHANS. In larval lampreys these endo- crine cells lie adjacent to the rest of the pancreatic tissue, and only become embedded within it in the adult. The endocrine cells are distinguishable histologically because they stain much less readily than the surrounding exocrine gland cells. This 1s due to the constant presence of so-called beta (8) cells that secrete the its diabetogenic hormone, INSULIN (§ 5.212). These cells have very E 50 SOURCES OF KINETIC AND METABOLIC HORMONES fine granules only, and have little affinity for any cytoplasmic stains. In at least some of the higher vertebrates, and especially in such mammals as the cat and dog, two other types of cell can also be distinguished in the islet tissue. Of these, the alpha (a) cells secrete the diabetogenic hormone GLUCAGON (Table 5); they contain large granules that stain bright red with Mallory-azan stain, but their cytoplasm also has little afhnity for any stains. The D type of cell in the islets stains blue in the same preparation, but its function is unknown (Fig. 2-14); Maximow and Bloom, 1972). 2.3 MESODERMAL SOURCES IN VERTEBRATA Metabolic hormones secreted by mesodermal sources have so far only been found in vertebrates. These come from the adrenal cortex and its homologue, the interrenal tissue. Other hormones from the mesoderm are all morphogenetic in their actions, or predominantly so; their sources, including the source of pro- gesterone (despite the possible kinetic activities that have been attributed to this hormone in § 4.12), will be described in Part II. 2.31 ENDOCRINE GLAND CELLS DERIVED FROM COELOMIC EPITHELIUM In all vertebrates the coelomic epithelium in the region of the kidneys gives rise to characteristic yellow gland cells, filled with fat and secreting cortical sterolic hormones. The cells are homolo- gous in all classes; but they are differently named according to their positions. In fish they often retain their median position between the kidneys, where they form interrenal tissue, or they may become paired, as in the perirenal organs of Dipnoi. In the tetrapods the tissue forms the adrenal cortex; it is always paired, lies in front of the kidneys, and becomes closely associated with the adrenal medulla during development. The cortex finally encloses the medulla completely in mammals (Table 6). 2.311 Interrenal tissue In Elasmobranchii, cortical gland tissue occurs as one or more median yellow masses, the interrenal tissue, so-called from its position between the kidneys. It is widely separated from the paired $ 2.311 ENDOCRINE CELLS FROM COELOM 51 groups of suprarenal cells which represent the adrenal medulla of higher forms. In Teleostei and related bony fish, the homologous tissue is called the anterior interrenal body, to distinguish it from the corpuscles of Stannius, or posterior interrenal bodies. The former resembles the adrenal gland of tetrapods, in containing a mixture of typical cortical and medullary cells. Of these, the cortical cells TABLE 6. MESODERMAL SOURCES OF METABOLIC HORMONES IN VERTEBRATA TYPE SOURCE OF NA SECT. pene ME OF OF | SECT HORMONE HORMONE ACT-| NO. ION* Elasmobranchii Interrenal tissue Cortical hormone MoT Sstt Teleostet Anterior ,,_,, me . MES 311 (& corpuscles of Stannius ?) me a 2.31 ENDOCRINE GLANDS FROM COELOMIC EPITHELIUM Tetrapoda Adrenal cortex - = SSSS58588 Aldosterone-like Hydrocortisone-like | M | 5.321 * M = metabolic. respond to stimulation by the adrenocorticotrophic hormone, ACTH, from the pituitary. Like the interrenal cells of the elasmobranchs, this tissue yields an active extract which is inter- changeable with that of any tetrapod adrenal cortex. Injection of any of the extracts can keep alive a mammal or bird from which the cortex has been removed. The corpuscles of Stannius are small, paired globules of tissue, embedded in the mesonephric kidneys; there are numerous pairs 52 SOURCES OF KINETIC AND METABOLIC HORMONES in Amaia, but most teleosts have only one pair. Histologically their cells look like cortical tissue; but they do not seem to respond to ACTH, and their function is uncertain. Their homology with true interrenal tissue is also in doubt, since they appear to arise from the pro- and meso-nephric ducts, and not from coelomic epithe- lium (cf. Pickford and Atz, 1957). 2.312 Adrenal cortex Like the interrenal tissue, the adrenal cortex is a small mass of conspicuous yellow tissue; but it is paired and situated in front of, or below, the kidneys, whether these are mesonephric or meta- nephric. The colour is due to large amounts of fat enclosed in the cells and associated with the formation of the sterolic hormones secreted by the gland. The presence of stored hormone in the gland is also related to the presence of ascorbic acid, which becomes depleted when the hormone is passed into the blood stream in response to some form of stress (§ 4.231 and Fig. 4-9), and to release of ACTH. Three layers can usually be recognised in the cortex: an outer layer (just within the connective tissue capsule enclosing the whole gland), where active cell multiplication follows the frequent nuclear mitoses ; a thicker middle region of actively secreting cells; and the innermost layer, next to the medulla, where the cells become degenerate and are eventually consumed by macrophages from the blood. It seems that throughout the life of the gland, cells are formed near the outer surface, migrate inwards during their secretory phase and are then destroyed as they reach the inner surface. The main secreting cells of the cortex form a compact mass or continuum, in which the individual cells tend to be polyhedral, from contact with adjacent cells (Fig. 2-14c). In the rat, the mass is tunnelled through by a network of blood sinusoids, so abundant that every cell has a facet in contact with a blood vessel into which its secretion can be passed (Pauly, 1957; Fig. 2-15). The structural details of the gland, and the proportion of connective tissue to gland cells, varies from species to species; but this has not been shown to have any effect upon the functional activity of the gland. No nerves have been detected. § 2.4 REFERENCES 53 Fic. 2-15. Stereogram of part of the ADRENAL CORTEX of Rattus; the cells form a continuum through which blood capillaries tunnel to make contact with each cell. The fibres on the outer (upper) surface represent the connective tissue capsule surrounding the gland; below this, the blood vessels branch freely and then run directly inwards through the main secretory region (zona fascicu- lata) of the gland and anastomose at the inner (lower) surface, next to the medulla (which is not shown). (From Pauly, 1957). 2.4 REFERENCES ALEXANDROWICZ, J. S. (1953). Nervous organs in the pericardial cavity of the decapod Crustacea. ¥. mar. biol. Ass. U.K. 31: 563-580. Amag, R. (1948). Un organe endocrine chez Jdotea (Crustacea isopoda). GCG. R. Acad. Sci., Paris, 227: 301-303. Arvy, L. (1957). In discussion of I. Chester Jones. Colston Pap. 8: 273-274. 54 SOURCES OF KINETIC AND METABOLIC HORMONES Arvy, L. and Gase, M. (1953). Données histophysiologiques sur la neurosécrétion chez quelques Ephéméroptéres. La Cellule, 55 (2): 203-222. Baca, Z. M. and GurretTTI, F. (1951). La sécrétion externe et interne des glandes salivaires postérieures des Céphalopodes Octopodes. Arch. int. Physiol. 59: 288-314. BARGMANN, W. (1958). Elektronenmikroskopische Untersuchungen an der Neurohypophyse. In Zweites Internationales Symposium iiber Neuro- sekretion, edited by W. BARGMANN, B. HANSTROM and B. SCHARRER. Berlin: Springer-Verlag. 4-12. BARRINGTON, E. J. W. (1958). The localization of organically bound iodine in the endostyle of Amphioxus. ¥. mar. biol. Ass. U.K. 37: 117-126. Buss, D. and WELSH, J. H. (1952). The neurosecretory system of brachy- uran Crustacea. Biol. Bull. Wood’s Hole, 103: 157-169. CAMPENHOUT, E. VAN, (1946). The epithelioneural bodies. Quart. Rev. Biol. 21: 327-347. CARLISLE, D. B. (1953). Studies on Lysmata seticaudata Risso (Crustacea Decapoda) VI. Notes on the structure of the neurosecretory system of the eyestalk. Pubbl. Staz. zool. Napoli, 24: 435-447. CARLISLE, D. B. and KNow_egs, F. G. W. (1953). Neurohaemal organs in crustaceans. Nature, Lond. 172: 404-405. CARLISLE, D. B. and Passano, L. M. (1953). The X-organ of Crustacea. Nature, Lond. 171: 1070-1071. Der Lerma, B. (1956). Corpora cardiaca et neurosécrétion protocérébrale chez le Coléoptére Hydrous piceus L. Ann. Sci.nat.(b) Zool. 18: 235-250. De Rosertis, E. (1949). Cytological and cytochemical bases of thyroid function. Ann. N.Y. Acad. Sci. 50: 317-335. EnamI, M. (1951). The sources and activities of two chromatophoro- tropic hormones in crabs of the genus Sesarma. II. Histology of incretory elements. Biol. Bull. Wood’s Hole, 101: 241-258. GaBE, M. (1954). La neuro-sécrétion chez les invertébrés. Année biol. Ser. 3, 30: 5-62. GREEN, J. D. (1951). The comparative anatomy of the hypophysis, with special reference to its blood supply and innervation. Amer. F. Anat. 88: 225-312. GrossMaN, M. I. (1950). Gastrointestinal hormones. Physiol. Rev. 30: 33-90. HANstTROM, B. (1939). Hormones in Invertebrates. Oxford: Clarendon Press. Hanstr6M, B. (1940). Inkretorische Organe, Sinus Organe und Nerven- system des Kopfes einiger niederer Insektenordnungen. Kungl. Svenska Vetenskap. Handl. Ser. 3, 18, No. 8: 3-266. Harris, G. W. (1955). Neural Control of the Pituitary Gland. Monogr. Physiol. Soc. (3). London: Edward Arnold. § 2.4 REFERENCES 55 KLEINHOLZ, L. H. (1947). A method for the removal of the sinus gland from the eyestalk of crustaceans. Biol. Bull. Wood’s H. ole, 93: 52-55. KNow Les, F. G. W. (1953). Endocrine activity in the crustacean nervous system. Proc. roy. Soc. B. 141: 248-267. KNowLEs, F. G. W. (1954). Neurosecretion in the tritocerebral com- plex of crustaceans. Pubbl. Staz. zool. Napoli, 24, Supplemento: 74-78. KNow _es, F. G. W. (1955). Crustacean colour change and neurosecre- tion. Endeavour, 14: 95-104. KNOWLES, F. G. W. and Car iste, D. B. (1956). Endocrine control in the Crustacea. Biol. Rev. 31: 396-473. Maximow, A. A. and BLoom, W. (1942). A Textbook of Histology. Phila- delphia and London: W. B. Saunders Company. MENDEs, M. V. (1948). Histology of the corpora allata of Melanoplus differentialis (Orthoptera: Saltatoria). Biol. Bull. Wood’s Hole, 94: 194-207. MIALHE-Vo Loss, C. (1955). Activité corticotrope des lobes antérieur et postérieur de l’hypophyse, chez le rat et le canard. ¥. Physiol., Paris, 47: 251-254. Pauty, J. E. (1957). Morphological observations on the adrenal cortex of the laboratory rat. Endocrinology, 60: 247-264. PIcKFoRD, G. E.and Atz, J. W. (1957). The Physiology of the Pituitary Gland of Fishes. New York: New York Zoological Society. Potter, D. D. (1954). Histology of the neurosecretory system of the blue crab Callinectes sapidus. Anat. Rec. 120: 716. SCHARRER, B. (1952). Neurosecretion. XI. The effects of nerve section on the intercerebralis—cardiacum—allatum system of the insect Leucophaea maderae. Biol. Bull. Wood’s Hole, 102: 261-272. SCHARRER, E. and ScHARRER, B. (1954a). Hormones produced by neuro- secretory cells. Rec. Prog. Horm. Res. 10: 183-240. SCHARRER, E. and ScHARRER, B (19545). Neurosekretion. In Handbuch der mikroskopischen Anatomie des Menschen, edited by W. VON MOLLEN- DORFF and W. BARGMANN. Berlin: Springer-Verlag. 6 (5): 953-1066. TuHomsken, E. (1954). Darkfield microscopy of living neurosecretory cells. Experientia, 10: 206-207 VAN Dyke, H. B., ADAmsons, K. Jr. and ENGEL, S. L. (1957). The storage and liberation of neurohypophysial hormones. Colston Pap. 8: 65-76. Weser, H. (1949). Grundriss der Insektenkunde. Jena: Gustav Fischer- Verlag. WEtsH, J. H. (1955). Neurohormones. In The Hormones, edited by G. Pincus and K. V. Turmann. New York: Academic Press Inc. 3: 97-151. WoerpDeman, M. W. (1957). Nomina anatomica parisiensa (1955) et B.N.A. (1895). Utrecht: A. Oosthoek Publishing Co. Youne, J. Z. (1936). The giant nerve fibres and epistellar body of cephalopods. Quart. J. micr. Sci. 78: 367-386. CHAPTER =3 KINETIC HORMONES I. CONTROL OF MUSCLES AND PIGMENTARY EFFECTORS THE TERM “kinetic” (§ 1.51) brings together a large group of hormones, which act upon certain effectors in the organism in ways which often resemble the effects of nerve stimulation. Kinetic hormones acting upon muscles and pigmentary effectors are considered in this chapter, and those causing the secretion of glands, both exocrine and endocrine, in the next (§§ 4.1 and 4.2). The similarity in action between these kinetic hormones and some nerves is not entirely accidental, since many of the kinetic hormones are neurosecretions from modified nerve cells (§ 2.111), and some are chemically akin to the acetylcholine or noradrenaline secreted by cholinergic and adrenergic nerves respectively. An important difference lies in their means of distribution; the hor- mone reaches the effector through the blood circulation and is therefore widespread in its effect, whereas the nerve cell releases its chemical in contact with a single effector only. There are, however, other kinetic hormones that are not derived directly from nervous tissue. Some of these, like those from the corpus allatum of insects or the adenohypophysis of vertebrates, are likewise derived from the ectoderm. A few others, such as secretin, appear to be derived from neither nerve cells nor ectoderm, but from the endoderm (§ 2.22). The kinetic hormones therefore form a wider group than either ‘‘neurohormones” (Welsh, 1955) or the secretions of ‘“‘neurohaemal organs”’ (Carlisle and Knowles, 1953); yet they show quite sufficient functional similarity among them- selves to warrant their inclusion in one group. The means of stimulating their secretion may be mechanical, as 56 93.111 CONTROL OF MUSCLES 57 in the case of gastrin, or chemical, as in that of secretin (§ 4.111) but it is usually nervous (§ 4.32). Few are under the control of any other hormones, and this is probably a question of speed. Hormone action is slower than nerve action; and, whereas the delay due to one hormone may not be significant for the effectors in question, a chain of two hormones might well be so. The hormones dealt with in the following sections are shown in a series of tables, where they are arranged according to their actions. The hormones of vertebrates have names which are widely accepted and are known to occur in a variety of animals; the example quoted is usually the one described in the text, but in no way implies that the hormone is limited to the genus named. Hormones of invertebrates for the most part have no names and can therefore only be referred to by the organ from which they are secreted. If they have a name, or an abbreviation that is used in the text, this is given in a separate column. Each invertebrate example in the tables is also referred to in the text; it is often the only example from which the hormone has so far been identified. . > 3.1 CONTROL OF MUSCLES Muscles can be grouped according to their functions or their histology, and there is some evidence that all types can be influ- enced by hormones. It will be convenient to take the involuntary muscles of the viscera and heart first, because these are the most commonly subject to hormone control and react similarly, although most visceral muscle of vertebrates is smooth, or unstriated, and that of arthropods and of the hearts in both phyla is striated. 3.11 VISCERAL MUSCLE In nearly all cases the action of hormones is to stimulate both the rate and amplitude of the contraction of visceral muscle; only rarely is a hormone known to inhibit muscle action (‘Table 7). 3.111 Heart muscle CRUSTACEA. Control of heart muscle by a hormone has been shown experimentally in the crab, Cancer pagurus, and the lobster, Homarus vulgaris. Extracts of the neurosecretory ‘PERICARDIAL ORGANS” (§ 2.112) added to fluid perfusing isolated hearts, I KINETIC HORMONES 58 snavquip’) yeisoAq uol[sues pjyauniqisag jeoseydoss0qng auopayy Apog Jeyjoystdq vjaunj quad umovipied snd10d DAIDADT SNO vjaun] diag umnovipivs sndi0d DAIDLDT pur], snus 4aIUD‘) ULSIC) [eIpsVoTI9g FTdWVxa ANOWYOH YO NVOUO ALVUGALYAANI snnby 9U0II}SO}SI T, sng é UssOISIO OuLoy] dUI[eUDIPY SYA sUT[eUdIPY SNSDIOJINAC) (HAY) uUlssoidose A YTL@) UID0}AXO SNSVIOJIAAC) SN]JOY UIDO0JAXO oY 9UOI}SeSOINUY > dUT[eUDIPY syay oad DUD urluTyoysAD9T0Y7) DUDY oUI[eUudIpPY sya * SNYJDY UII}Sey) SYA dUT[eUudIpPY FIdINVXd ANOWUOH ALVUGALYAA wounguyun UOUDINWUWIYS :sajasnu Kpog AIOSAW OILYNOS ZI'¢ stat fo sajasniu IPO OL T"¢ ‘94a ‘sajayjof WET OLL'E $]9SSAQ-poolg SLs $1]29 oyayndaon yy $L1'€ JInp [DINAH ELLE sIs[eqstiod “fo uowiquyut ce Joppeyq [es sioyouryds sIs[ejsiiod ‘fo uoypjnwiys NH ZILVE Cah be ha “ce UO1DAIJIIID “JAVAFT TIVE ATOSOW 'TIVHROSIA [T'S SHO LOFAAT SHTOSQW ONITIOWLNOOD SHNOWYOH OLLANIS *Z ATAV,T, § 3.111 VISCERAL MUSCLE 59 increase the amplitude and frequency of the heart beat (Fig. 3-1) This action is believed to be due to an ortho-dihydroxytryptamine and is closely similar to the action of adrenaline and noradrenaline, which are chemically not widely dissimilar. Results on other species are rather contradictory, and different doses are needed at different times of year to get similar results. This may be compared with the effects that adrenaline and noradrenaline have on vertebrates, where contraction or relaxa- tion can be obtained, according to conditions in the organs resulting from previous treatment, size of dose, etc. (Welsh, 1955). “It is assumed that the function of the pericardial organs in these Crustacea consists in liberating, through fine neuropile-like terminations of the nerve fibres, some hormone passing with the blood into the heart and producing on it a stimulating effect”’ (Alexandrowicz and Carlisle, 1953). Blood taken from the peri- cardial cavity before reaching the heart gave the same reaction as the extracts; but that taken from the leg arteries after leaving the heart did not, presumably because the chemical was destroyed before it reached the legs. Earlier statements (e.g. Welsh, 1937) that the sinus glands of Crustacea provided a heart-accelerating extract might have been unreliable, because insufficient care was taken to avoid the presence of histamines in the extracts (the same is probably true of extracts of sub-neural glands of ascidians credited with similar activity). A heart-accelerating hormone from SINUS GLAND extracts has, however, now been obtained from the freshwater shrimp, Paratya, and also an inhibiting extract from the brain (Hara, 1952). They are thought to be distinct from the chromactivators from the same sources (§ 3.223). INSECTA. The frequency of beat of the isolated heart of the cock- roach, Periplaneta americana, perfused with a suitable Ringer solution, can be increased some 50 per cent above normal, and the amplitude of the muscle contraction increased also, if an aqueous extract of one pair of CORPORA CARDIACA (§ 2.111) of the same species is added to each 10 ml of the perfusate (Cameron, 1953). At concentrations one-tenth as strong, the amplitude is still increased, but not the frequency. If the extract is separated by paper chromatography, one spot at a time can be eluted and added 60 KINETIC HORMONES—I m= " ith MN i} Hie Fic. 3-1. Kymograph record of the heart beat of the crab, Cancer. Changes in frequency are shown as abscissae in relation to the time scale in minutes (below); changes in amplitude are shown as ordinates. The record reads from left to right. The arrows indicate the times at which different reagents reach the heart, which is the § 3.111 VISCERAL MUSCLE 61 to the perfusate. Only one of these shows the same action as the crude extract; this active constituent is thought to be an ortho- diphenol (and therefore related to adrenaline). Beautiful as this technique is, it only demonstrates, like most experiments on organ extracts, the pharmacological fact that the animal produces a chemical which has an effect upon the heart. It would be another matter to show conclusively that in life the heart beat is physio- logically controlled in any way by this substance, or that in the absence of the corpora cardiaca the animal is unable to control its heart beat to suit the conditions under which it is living. The stimulus which might cause the secretion of the chemical is unknown. The corpus cardiacum acts as a storage organ for a neurosecretion from the brain, as can be seen in Leucophaea, where severing the nerve on one side prevents the passage of the secretion (Fig. 3-2). Yet if a similar operation is performed on Periplaneta and separate extracts of the corpora cardiaca of the two sides are tested after 5 and 17 days, that from the severed side is still as active as the other. This eliminates the neurosecretion as a source of the heart- accelerating hormone, which must be the intrinsic secretion of the corpus cardiacum itself. These cells are of ectodermal, rather than nervous, origin (§ 2.112), so that their secretion is one of the few kinetic hormones that is not a vascular “‘neurohormone”’ (Welsh, 1955), although its action is akin to that of adrenaline, secreted from cells of the vertebrate sympathetic nervous system (Mendes, 1953). Adrenaline itself has a similar stimulating effect upon the heartbeat of many invertebrates, whether it is neurogenic or myogenic; but there is as yet no clear evidence of its being secreted by any gland in an invertebrate. In only a few inverte- brates is adrenaline used as a chemical transmitter at any nerve ending. VERTEBRATA. ADRENALINE (§ 2.111) increases the amplitude same specimen throughout the series: (4) extract of PERICARDIAL oRGAN of Cancer; (B) adrenaline (1 :10°); (C) noradrenaline (121 02)3 (D) the same extract as in (A). Between exposures to these sub- stances the heart is restored to seawater and the beat slows down with a reduced amplitude (from Alexandrowicz and Carlisle, 1953). 62 KINETIC HORMONES—I Pors intercerebralis of brain Accumulation of neurosecretory moteriol proximal to site of nerve section Nervus corp. cardiaci with unobstructed flow of neurosecretory material Depletion of neurosecretory Boe: i; Storage of neurosecretory material in corp.cardiacum STE ee evn MOtEriO! in Corp. cardiacum ie i ' eH) of operated side ; of operated side Larger corp.allatum of operated side Fic. 3-2. Diagram of the dorsal aspect of the brain, corpora car- diaca, corpora allata and the nerves connecting them in the cock- roach, Leucophaea maderae. On the left side the nerve from the neurosecretory cells in the brain has been cut, so that the neuro- secretion accumulates in the proximal part of the nerve and does not reach the corpus cardiacum of that side (from Scharrer, 1952). and frequency of the heart beat in Vertebrata. This can only be demonstrated with difficulty in normal mammals, although as little as 1 part of adrenaline in 1,400,000,000 (Turner, 1955) can increase the beat of a denervated heart, freed from the ‘“‘depressor”’ action of parasympathetic fibres of the vagus nerve. It is likely, however, that the action of this drug on the heart is usually through the sympathetic nerve, rather than through the circula- tion, as the drug, or hormone, is quickly destroyed in the tissues by an enzyme. 3.112 Gut muscle Insecta. The striated muscles in the gut of Periplaneta (Cameron, 1953) and Locusta will record peristaltic movements for at least Fic. 3-3. Rhythmic peristaltic contractions of intestinal muscle of the cat, Felis; frequency is recorded as abscissae from left to right and amplitude as ordinates. From a to 6, the muscle is in Ringer’s solution; then at 4, and again at f, blood from an “‘excited”’ cat, that had been barked at by a dog for some time, is added and the contained ADRENALINE causes almost immediate and quite prolonged inhibition of peristalsis. At d, peristalsis is restored by changing the perfusing fluid to one containing blood from a “quiet” cat, in which adrenaline secretion had not been stimulated (from Cannon, 1915). Fic. 3-4. Smooth muscle contraction in isolated rat uterus, showing frequency as abscissae, and amplitude as ordinates. The fresh extract of the neurohypophysis, containing OXYTOCIN, injected at the time indicated by the arrow on the left, induces strong rhythmic contractions. The marked time interval is 10 min. (after Trendelenburg in Bud denbrock, 1950) § 3.112 VISCERAL MUSCLE 63 24 hours, if perfused with a suitable Ringer solution. In Peri- planeta, an extract of one pair of its own CORPORA CARDIACA to 10 ml Ringer solution causes the rate of peristalsis in the hind-gut to be doubled, and stronger solutions have more marked effects; but the peristalsis of the fore-gut seems to be inhibited. It is certain, at least in Periplaneta and in the locust, that increase in peristalsis of the Malpighian tubules can also be induced by a substance from the corpus cardiacum, as well as possibly one from the BRAIN. The latter observation requires confirmation, in view of Cameron’s finding that the neurosecretion from the brain does not contain the active principle causing heart muscle contraction. VERTEBRATA. The visceral muscle of the vertebrate gut is mainly under the dual control of the nerves of the sympathetic and parasympathetic systems working in opposition; but the effect of the former can also be brought about in emergency by adrenaline from the adrenal medulla. COLD-BLOODED VERTEBRATES. The effects of ADRENALINE appear to depend upon dosage. In the frog, Rana, small doses stimulate the contraction of gut muscles; but larger doses inhibit it (Budden- brock, 1950). . MAMMALIA. ADRENALINE, Secreted in response to excitement or fear, contracts the sphincter muscles of the gut and inhibits peristalsis, as can be shown by adding blood from an excited cat to the Ringer solution in which isolated gut muscle is contracting rhythmically (Fig. 3-3). Both reactions tend to stop digestion and fit in with the emergency mobilization of the blood supply in the somatic muscles. As its name implies, CHOLECYSTOKININ contracts the muscles of the gall-bladder in frogs and in most mammals. It may also relax the sphincter muscle. The hormone therefore causes dis- charge of the bile down the cystic duct, and appears to be the only means of stimulating this reaction, for which there 1s no nerve control (Grossman, 1950). It can be extracted from the duodenal mucosa, and in nature is secreted in response to the presence of fat, or fatty and other acids, in the duodenum. Its histological origin has not been determined, but it appears to be derived from endodermal cells, like secretin, and therefore not 64 KINETIC HORMONES—I to be a neuro-secretory product. It is absent in the horse, Equus, which has no gall-bladder. Gastrin, secreted by the stomach, can induce the contraction of stomach muscles, forcing food into the duodenum as the gastric phase of digestion is completed; but this action of GASTRIN follows, and is subsidiary to, that of stimulating the acid-secreting gland cells of the stomach (§ 4.111). The contraction is said to be inhibited by enterogastrone; but this action is not purely hor- monal, since it is more effective if the vagus nerves are intact (Grossman, 1950). The nervous inhibition can be stimulated by acid in the duodenum. 3.113 Muscles of the genital ducts No cases seem to have been recorded so far in which the muscles of any part of the genital ducts of an invertebrate are controlled by hormones. Mamma tla. Isolated uterine muscle of many mammals, such as the rat, Rattus, reacts to OXYTOCIN from the neurohypophysis by strong rhythmic contraction (Fig. 3-4). At the end of pregnancy these contractions force the embryo out of the uterus. The fact that this does not normally occur until the embryo is fully devel- oped seems to be due, not to a lack of oxytocin during earlier stages of pregnancy but to changes in the level of sensitivity of the uterus. In the rabbit, Oryctolagus, the relative insensitivity of the uterus to oxytocin during the 30 days of pregnancy, as compared with variability at other times, is due to the abundant presence of progesterone. As this decreases and oestrogen increases in the circulation towards the end of pregnancy, the uterus becomes increasingly sensitive to the oxytocin until finally it reacts sufficiently strongly to bring about parturition (Fig. 3-5). Earlier experiments, in which it was found that rats from which the hypophysis had been removed were still able to produce their litters successfully, have now been explained on the grounds that the neurohypophysis is only a storage organ for hormones secreted in the hypothalamus of the brain (§ 2.111), and the experimental technique therefore failed to remove the source of the oxytocin. In those cases where the source was also destroyed by hypothala- § 3.114 VISCERAL MUSCLE 65 mic lesions, parturition was not achieved, both the mother and the litter dying in the attempt. It has also been suggested that oxytocin may play some part in driving sperm upwards into the fallopian tubes after copulation, 0-002 P & 5 b 0:03 fo} ‘S ee 5 2 a o oe 0:5 >2:0 0 10 20 30 40 50 60 70 Days after mating Fic. 3-5. Graph showing schematically the sensitivity to OXYTOCIN of strips of rabbit uterus tested at various stages of pregnancy. Days after mating are shown as abscissae, with the normal term for parturition at P; ordinates (on a logarithmic scale) show reactivity expressed as the minimal number; of units of oxytocin which causes a motor effect when added to 100 ml of Ringer-Locke solution (from Robson, 1933). by affecting the tonus differentially in different parts of the oviduct (Fitzpatrick, 1957). 3.114 Myoepithelial cells of mammary glands There is no known case of a hormone activating the duct muscles or myoepithelial cells of a gland in invertebrates. Mammatta. Branched myoepithelial cells (Fig. 3-6), which surround the alveoliand smaller ducts of the mammary glands, can, by their contraction, drive the milk from the ductules of the gland F 66 KINETIC HORMONES—I down into the teats, achieving what is known to farmers as “‘let- down”’ of the milk. Figure 3-7 shows that injection of OXYTOCIN into an anaesthetized bitch, Canis, stimulates the contraction of these cells, and increases the amount of milk available to the puppies. In the normal state, the act of suckling (or milking) stimu- lates the brain to induce secretion of oxytocin for this purpose. Fic. 3-6. Diagram of a generalized exocrine gland to show the location of factors which may influence the flow of secretion. Duct activity influenced by: 1, smooth muscle sphincters; 2, longi- tudinal smooth muscle shortening ducts or producing peristalsis; 3, myoepithelium; 4, reservoirs in large ducts, or cisterns; 5, vasodilatation pressing on ducts or reservoirs; 6, vasoconstriction shortening inter-lobular vessels and squeezing adjacent ducts; 7, secretion from duct epithelium. Lobule activity influenced by: 8, smooth muscle bundles in inter- lobular septa squeezing lobules as a whole; 9, smooth muscle interspersed between alveoli; 10, vasodilatation or vasoconstriction affecting alveoli mechanically; 11, myoepithelium; 12, elastic fibre recoil in stroma, when pressure in distended alveoli is released; 13, nervous stimuli to secretory epithelium and smooth muscle; 14, hormonal stimuli to epithelium. In mammary glands the myoepithelial cells, 11, round the lobules, contract in response to OXYTOCIN. The secretory epithelium, 14, is stimulated by PROLACTIN (§ 4.13). (From Richardson, 1949). $3115 VISCERAL MUSCLE 67 3.115 Muscles of blood vessels No case of hormone control of muscular contraction in the vascular system (vasoconstriction), apart from the contraction of heart muscle, has been reported for any invertebrate except, possibly, in the cephalopoda (p.416). VERTEBRATA. The arteries of vertebrates react to a number of hormones: for instance, to VASOPRESSIN*, ADH, and to a lesser extent to OXYTOCIN, both from the neurohypophysis (§ 2.114). This effect, which may be merely pharmacological, can best be shown by intravenous injection of vasopressin into any tetrapod after the peripheral blood vessels have been expanded by hypophysectomy (Buddenbrock, 1950). Certain doses of pure ADRENALINE dilate the peripheral blood vessels when first injected into the rabbit’s ear, whereas subsequent injections of similar doses cause contractions (Fig. 3-8). This seems to be unexplained. 3.116 Other visceral muscles VERTEBRATA. Visceral muscles attached to the hair follicles in mammalian skin cause erection of the hair (as on a dog’s neck or a cat’s tail) in response either to sympathetic nerve stimulation or tO ADRENALINE secretion. These also cause the radial muscles of the iris of the eye to contract, thereby greatly enlarging the pupil. The release of adrenaline into the circulation, as a result of fear, shock or rage (cf. secretion in response to excitement, Fig. 3-3), is therefore accompanied by at least the sensation of the hair standing on end, and by the appearance of staring eyes with expanded pupils, and blanching of the face, due to the contraction of the peripheral blood vessels (§ 3.115). More useful features of this “emergency” syndrome due to adrenaline are the increased rate of heart beat (§ 3.111), and the enlarged blood flow bringing a greater supply of sugars to the body muscles (§ 5.211), which enable the animal to achieve a very high output of muscular effort for a time—probably long enough to effect an escape from the predicament causing the original fright. * This hormone would be better named antidiuretin from its main action (§ 5.32). 68 of or iS KINETIC HORMONES—I 100 9 o Oo Milk , Oxytocin Gt 2-3 45 36-7 8.9 0 WV 284 See Time, min Fic. 3-7. Effect of OXYTOCIN on milk ‘‘let-down”’ in the nursing bitch, Canis. The time in minutes is given as abscissae and the amount of the milk yield (as indicated by the increase in weight of the pups) as ordinates. In the normal case, the pups get all the milk that the mammary glands can yield in about 8 min, there being an initial latent period before milk becomes available to them and a falling off as the gland becomes exhausted. In the right-hand curve, starting again at time 0, the pups were put to the anaesthetized mother and failed to obtain more than 30 per cent of the milk, after a very slow start. After 7 min an injection of 0.5 ml Pituitrin containing oxytocin enabled the pups to obtain almost all the rest of the milk, presumably by causing contraction of the myoepithelial cells round the alveoli. The injection of oxytocin has no effect upon the amount of milk secreted, as can be seen by the lack of effect of similar injections made when the curves had already exceeded 90 g (from Gaines, 1915). In the vertebrates, most types of visceral muscle which are innervated by the sympathetic fibres of the autonomic nervous system are thus seen to react also to secretion from the adrenal medulla; but the nerves of mammals are now known to secrete mostly noradrenaline at their end-plates, whereas the greater part the gland secretion (derived from neurosecretory cells which iginated in sympathetic ganglia) is adrenaline itself, although it admixed with larger amounts of noradrenaline in the lower vertebrates, e.g. in the secretion of the suprarenal bodies of the elasmobranchs. Fic. 3-8. Changes in volume, increasing upwards as ordinates, of the perfused ear of a rabbit, Oryctolagus, with time in 30 sec intervals as abscissae. In the upper tracing increase in volume is due to dilation of the blood capillaries following the first injection of adrenaline. The lower tracing shows the opposite effect of two later injections of the same dose. This is an example of reversal from a dilator response of the capillaries to a constrictor response with repeated doses (from Burn and Robinson, in Welsh, 1955). $3.12 SOMATIC MUSCLES 69 3.12 SOMATIC MUSCLES CEePHALOPODA. Ablation of the EPISTELLAR Bopy of the octopod, Eledone moschata (§ 2.111, Fig. 2-4), is followed by general loss of muscle tone, with even the tentacles hanging limply ; but recovery starts about a week after the operation (Young, 1936). In one case the tone gradually returned to normal during the 186 days for which the animal survived, although there was no trace of regene- rated epistellar tissue. No attempt seems to have been made to restore the tone by injecting extracts during the first post-operative week; but control experiments made it clear that the effect cannot have been due to shock, which is an important factor in these very sensitive animals with their highly developed nervous systems. The loss of muscle tone extended to the chromatophores, so that animals without the epistellar body became abnormally pale (cf. § 3.21): CrusTACcEA. Well-controlled experiments (Roberts, 1944) show that exposure of the crayfish, Cambarus virilis, to relatively bright light releases an unidentified EYESTALK HORMONE that reduces loco- motion. It is possible that this hormone may act either by raising the stimulation threshold of the skeletal muscles of the legs or by lowering the strength of the nerve impulses from the brain; but it seems more probable that the result is due to an indirect metabolic effect (§ 5.1). The reaction may have adaptive value because the animals normally feed by night and escape from predators by remaining hidden by day. Insecta. The normal rhythm of nocturnal activity of the cock- roach, Periplaneta, is stimulated by a HORMONE, from the SUBOESO- PHAGEAL GANGLIA, the secretion of which is controlled through the ocelli. If the illumination is kept constant, or the ocelli are painted over, the rhythm disappears. The action of the hormone has been shown by implants and in parabiotic experiments, in which two cockroaches are so joined that blood can flow from one to the other. If, for instance, a specimen in which the ocelli have been occluded has another specimen, with no legs, joined to its back, then normal diurnal changes in illumination acting upon the upper specimen cause a correlated rhythm of locomotor activity in the lower (Harker, 1956). 70 KINETIC HORMONES—I VERTEBRATA. There are no specific hormones in vertebrates to control somatic muscles, although these are noticeably affected by the hormones from the gonads and also to some extent by thyroxine and adrenaline. For instance, the loss of TESTOSTERONE in castrated mammals, such as cart-horses, results in lowered spontaneous activity and muscle tone, and is shown by the whole stance of the animals, as compared with a stallion, Equus. In females, the pres- ence of oestrogen in the circulation during oestrus (§ 4.234 and Part II, § 4) is accompanied by a great increase in activity, as has been shown by attaching a pedometer to a sow. Similar variations in activity are shown during the 4-day oestrus cycle of rats (Beach, 1948); a loss of 82 per cent in muscular activity can follow ovari- ectomy. It seems probable that all these effects and those of thyrox- ine may be the result of metabolic changes caused by changes of hormone balance, rather than of any direct kinetic action of the hormones on the muscle contractions, or even on the tonus. The action of adrenaline is slightly to prolong the active state of the muscle fibres and to increase the tension accompanying a twitch initiated by the nervous system; it does not itself cause contraction of skeletal muscle as it does in the case of visceral muscle (Goffart and Ritchie, 1952). 3.2 CONTROL OF PIGMENTARY EFFECTORS Pigmentary effectors of a variety of animals bring about colour changes mainly in two distinct ways: one in which extrinsic muscle fibres change the shape of the colour-containing cells, and the other in which the pigment granules themselves are moved within the confines of a stationary cell. Rarely, the cells change shape and may even move their position. These processes cause so-called physiological colour changes (‘Tables 8 and 9). ‘“Morpho- logical colour change”’ (Sumner, 1940, and Dawes, 1941) produces similar effects by slow alteration in the total amount of pigment present, rather than by its redistribution. This reinforces the adaptive physiological changes when the external conditions remain relatively constant for days or weeks. Together they play an important part in providing “protective coloration” for the animal. 71 CONTROL OF PIGMENTARY EFFECTORS § 3.2 (s}ooy9 yOoIIpuy) x DIN sajauowuanj vg DIN) SAJOUOMIDID «e snsvquv;) sé snisnpav’y «ce auopalq HIdNVXY é so.INssTuUIUOd ce puvjs snus é ypeisoA é uol[sues [eeseydossoqng ZoUlIejIg gOUTUURIA J, ANOWUYOH YO NVOUO ALVUEALYAANI ysiep url uoNexeyar JYSIT UT UOTIORIZUOD ‘saagyf I]UYIDAJUOD Y}1M2 $7199 JOUIZAT == yiep UI UOT}eI}UIDU09 yYSI] ut Testodsip /8]]29 [DUNad JoOISIP KADUOUIDIS' T77'E u0l}e.1}U99DU09 [esiodsip /s]]a9 pousapida ajduugy 177'¢ SHTIANVUD LNINDId AIGVAOW HLIM STTAD 7Z'E SUIUd}Y SI] *9°2 UOTeXLISI V[ISNIA[ SuUIUDyIEp *a°2 UOT}NRIZUOD VfOSN]A[ SHIOSAW HLIM SHUOHdOLVYINOUHD § [7'¢ ATdHNVXA ANOWYOH ALVUEFALYHA SYOLOFATaA SYOLOGIAT AUVINANDId DNITIOWLNOOD SHNOWYOH OILANIS °8 ATAV T, Ta KINETIC HORMONES—I 3.21 CHROMATOPHORES WITH MUSCLES CEPHALOPODA. This type of chromatophore occurs only in cephalopods, and forms a convenient link between those muscular structures which have been considered in the previous section, and the chromatophores with movable granules which follow. SZ Fic. 3-9. Chromatophores with muscles from a _ cephalopod, Loligo; on the left the muscles are relaxed and the chromatophore cell is elastically contracted so that it looks pale, with the pigment in a small mass at the centre; on the right the muscles have con- tracted and stretched the cell body to which they are attached so that the chromatophore shows the maximum amount of colour (from Bozler, 1928). The cephalopod chromatophore (Fig. 3-9) consists of a central pigment-containing cell with a highly elastic wall, and from 4-24 single muscle fibres, attached radially around the circumference; when the fibres all contract, they increase the area of exposed pigment. Contraction of muscles, therefore, corresponds to ex- pansion of the pigment and a darkening in appearance of the animal. Although each muscle fibre is under direct nerve control, the fibres to any one chromatophore usually contract together; but adjacent chromatophores can be separately stimulated to produce the very rapid and varied patterns of colour change which are characteristic of cephalopods and appear to be connected with their emotional states, as well as related to the colour of their environment. § 3.22 EFFECTORS WITH MOVABLE PIGMENT GRANULES 73 If this were the whole story, cephalopod chromatophores would deserve no place in the present context; but overall changes of colour can also be produced by chemicals in the blood. TyramMINE from the posterior salivary glands is normally used as poison for paralysing the prey, but is also associated with pigment dispersion, so that animals usually become much darker in colour when the salivary glands are active; BETAINE in the blood is associated with pigment concentration. Moreover, of the three Mediterranean octopuses, Eledone moschata and Octopus macropus are normally well-coloured species, but Octopus vulgaris is pale and habitually has less tyramine in its blood. Transfusion of blood from either of the first two into the last of these species results in darkening. Denervated chromatophores, on the other hand, are completely insensitive to these chemicals (Bacq and Ghiretti, 1951). Tyramine and betaine can therefore be better compared to “‘para-activators”’ (§ 1.2) than to true hormones, in that they seem to act through the nervous system and not directly upen the effectors. Their mode of action is still uncertain; Sereni (1930) postulated that they might act directly upon the inhibitory and excitatory centres in the brain, but this has not been fully established. 3.22 PIGMENTARY EFFECTORS WITH MOVABLE PIGMENT GRANULES Although varying in form and situation, these effectors all contain pigment granules which move to and fro within them, dispersing widely to give a large coloured area, or concentrating into a limited space to give only a small spot. Dispersal here gives the same effect as contraction of the muscles around a cephalopod chromatophore. The most plausible explanation of how this granule movement is brought about is that given by Marsland (1944) for the branched chromatophores of fish; but it seems probable that the same principle underlies all cases. The pigment granules are attached to a partially gelated system of long protein molecules in the cyto- plasm of the cell, and as the cytoplasm gelates fully the molecules contract, drawing the granules “‘as on a string bobbing through the current” towards the centre of the cell, while squeezing the 74 KINETIC HORMONES—I ‘‘plasmosol phase”’ out into the tips of the cell branches. Dispersal may then be a re-stretching of these proteins on solation, and not a matter of Brownian movement. High pressures (up to 8000 lb /in?) can be shown to cause solation, and to inhibit the concen- tration of the granules. This is on a par with amoeboid movement and particularly with muscle contraction; but the problem is still unsolved of whether either nerve stimulation or hormone action can affect the state of the chromatophore proteins in the same way as in the muscles, or whether changes in osmotic pressure (Abramowitz and Abramowitz, 1938) or in permeability of the cell membrane play a part. As a rule chromatophores react much more slowly than muscles; but there is a great difference in the reaction speeds of the similar-looking chromatophores of Amphibia, Reptilia and various Crustacea. This may be a question of the strength of stimulation. The effectors with movable pigment fall into three types. (1) The epidermal cells of certain Insecta are relatively un- specialized, except for the presence of pigment granules, which may be of more than one colour and may move in different direc- tions (Fig. 3-10, § 3.221). (2) ‘The retinal pigment cells of certain Crustacea and Insecta occur in two positions, proximal and distal, round the ommatidia of the compound eyes (Fig. 3-12, § 3.222), and contain moving granules of black pigment. Movement of reflecting, or white pigment, granules in cells round the retinulae also occurs. (3) ‘The branched and specialized chromatophores of Crustacea and some other invertebrates, as well as of lower Vertebrata, may be epidermal, but are usually mesodermal, and may contain more than one pigment; but if so, each pigment remains in its own branch of the cell (Plate 3-1, § 3.223). 3.221 Pigment movement in epidermal cells InsecTA. The cell-boundaries are reputed to disappear between moults, and the pigment granules can then migrate to an extent comparable with that in chromatophores. In green specimens of the stick insect, Carausius, the colour change is obscured by stationary yellow-green pigment; but in brown specimens the red pigment moves parallel to the surface of the epidermis, and the § 3.221 EFFECTORS WITH MOVABLE PIGMENT GRANULES 75 black melanin, which has been more fully observed, moves almost at right angles to it, to cause darkening in appearance (F ig. 3-10). The upward, or dispersing, movement of the melanin depends on both moisture and light, the effects of which are transmitted by the nervous system to the brain, and thence, either by nervous stimulation or by neurosecretion through the circumoesophageal connectives, to the SUBOESOPHAGEAL GANGLION (§ 2.111). This releases a Carausius-DARKENING HORMONE which passes in the blood to disperse the melanin. The ganglion cells only release the hormone if the connectives from the brain are intact (Dupont- Raabe, 1956). If moisture is maintained constant, the melanin granules con- centrate in the light and disperse in the dark, and tend to maintain Fic. 3-10. Two diagrammatic sections through the skin of the stick-insect, Carausius, to show the movement of pigment granules in epidermal cells. The clear space above indicates the position of the cuticle. The pigment in the upper section is concentrated in the light-adapted position, and in the lower dispersed, in the dark- adapted position. The green pigment (1) remains stationary; the red pigment (2) disperses laterally above the nuclei; the dark, melanin-like pigment (3) disperses mainly outwards (from Giers- berg, 1928). 76 KINETIC HORMONES—I this diurnal rhythm for a time under constant conditions. They show a limited background response, becoming darker on an illuminated dark background than they are on a white or yellow background. In constant light, the melanin disperses in response to moisture, under the control of what is probably the same darkening hormone, since its secretion is also stimulated through the nervous system (Giersberg, 1928). This was established by putting the insect into a humid box, with its head projecting through a diaphragm into the dry air outside (Fig. 3-11); this induces darkening of the whole animal from head to tail in about half an hour. If a ligature is put round the body to prevent the circulation of the blood and hormone to the tail end, the darkening only affects the part in front of the ligature. If the ventral nerve cord is cut at a level just outside the humid box, no darkening takes ee | Fic. 3-11. The stick-insect, Carausius, with the hinder part of the body in a moist chamber and the head and thorax projecting through a membrane. The pigment dispersion, caused by the moisture, is transmitted by a DARKENING HORMONE from the suboesophageal ganglion. This acts only upon the head and pro- thorax because of the ligature just behind them (from Giersberg 1928). ~ place, because the stimulus from the damp skin is not conducted to the brain. But if the same animal, with the nerve cord cut but the body unligatured, is then reversed, with its head in the box and the tail left out, the whole body darkens, because the stimulus from the skin of the head can reach the brain, which therefore stimulates the secretion of the hormone. This then circulates freely to all parts of the body. Secretion of the darkening hormone has been located histologically in the suboesophageal ganglia and confirmed experimentally by injection of extracts into animals from which the brain had been removed. Headless animals, lacking the source of the darkening hormone, have their pigment fully con- centrated, as in the normal light-adapted animals; they therefore § 3.222 EFFECTORS WITH MOVABLE PIGMENT GRANULES 77 form good test subjects for extracts containing the darkening hormone, and show that extracts from other parts of the brain are also active, but those from the corpora allata and cardiaca are inactive (Dupont-Raabe, 1954). A lightening hormone has not yet been located, partly because of the lack of a suitable preparation for testing extracts. It is not clear why damp-adapted insects with dispersed pigment are not used. One source, at least, of the concentrating hormone must be situated in the body, since it is present in headless animals from which the corpora cardiaca are almost certainly absent (Dupont-Raabe, 1956). Otherwise, it might perhaps have been supposed that the corpora cardiaca would yield a Carausius- lightening hormone, since they yield a neurosecretory substance (Cameron, 1953) which concentrates the melanophores of Crago (Knowles, Carlisle and Dupont-Raabe, 1955, § 3.223), and some, at least, of the erythrophores of Leander. It seems significant that the concentration of both these crustacean chromatophores are light-adaptations, and it is a light-adaptation hormone that appears to be missing from Carausius. 3.222 Pigment movement in retinal cells CrusTAcEA. The compound eyes of most Crustacea have three groups of pigment-containing cells round each ommatidium; distal and proximal retinal cells that surround the cone (Fig. 3-12), and reflecting cells that extend below the basement membrane but are not shown in the figure. These last contain white pigment, which behaves like the pigment in white chromatophores (§ 3.223) and is probably under similar hormone control. The proximal retinal cells contain dark pigment granules, which may be fixed in appositional eyes, such as those of Eupagurus, but which in many species can be dispersed outwards to isolate the sensitive thabdomes and convert superpositional eyes into a temporarily appositional type for accurate vision in bright light (Bruin and Crisp, 1957). The movement of the granules in these cells is sometimes a direct effect of light, as in some chromatophores (§ 3.223); but in others it is controlled by hormones, that appear to be the same as those controlling the distal cells (Knowles and Carlisle, 1956). 78 KINETIC HORMONES— I Fic. 3-12. Three ommatidia from the compound eye of the cray- fish, Cambarus bartoni; a—c in longitudinal section and d-—f in surface view. (a) Ommatidium of a typical dark-adapted eye with pigment concentrated at two levels: round the cone (con.) in the distal retinal cells (d.r.c.), and below the basement membrane (6.m.) in the proximal retinal cells (p.7.c.). The corresponding surface view (d) § 3.222 EFFECTORS WITH MOVABLE PIGMENT GRANULES 79 The distal retinal cells contain dark, melanin-like pigment granules. The pigment movement can be observed in the intact eye (Fig. 3-12, df), and is accurately adapted to the light intensity, especially over the lower ranges, such as inshore, under- water animals are most likely to encounter. The adaptive move- ment is, however, achieved in two different ways: (i) by the usual migration of pigment granules in stationary cells (Fig. 3-12), and (11) by contractile fibres which change the shape of the cells, thereby causing redistribution of the pigment in relation to the thabdome (Fig. 3-13). This latter process overrides any sign of pigment migration (Parker, 1932). Migration of pigment granules in stationary distal retinal cells In the crayfish Astacus and Cambarus and the crabs Dromia and Maza, the pigment granules move inwards to meet the out- ward movement of the proximal pigment, as they become light- adapted; they move outwards, towards the surface of the eye, to reach the dark-adapted position, which occurs in dim light, rather than in complete darkness. The latter movement appears to be comparable to concentration of pigment in the cell body of a chromatophore; the former inward movement is like dispersal of the pigment into the stationary but outlying cell-processes (Fig. 3-12c). In Cambarus, inward migration, or dispersal, of retinal pigments to their light-adapted position is induced by injection of an eye- stalk extract, the extent of the migration being dependent on the quantity of the RETINAL-PIGMENT-DISPERSING HORMONE, RPDH, used. The threshold for response of the distal pigment is lower than that for the proximal pigment (Fig. 3-12, b and c). No shows a bright orange centre to the cornea (crn), when seen by reflected light, because the rhabdome (rh) is unscreened by the proximal pigment. (b) and (e) In response to the injection into a dark-adapted animal of retinal-pigment-dispersing hormone extracted from one eyestalk, the distal pigment has partially dis- persed inwards but the proximal has not moved towards the light- adapted position. (c) Extract from two eyestalks has been injected and pigment in both sets of retinal cells has dispersed to the fully light-adapted position. In surface view (f) this eye appears black all over, as in a naturally light-adapted animal (from Welsh, 1939). 80 KINETIC HORMONES—I evidence of the action of a concentrating hormone has been reported (Welsh, 1939). Movement of pigment due to change in shape of distal retinal cells Change in cell shape in the prawns Palaemonetes and Leander and in a Bermudan shrimp Anchistioides is brought about by con- tractile fibres (Welsh, 1936). ‘The distal retinal cells here lie in the same position round the cones of the ommatidia, as in Cam- barus; but if the usual pigment migration occurs, it is obscured by such a surging inwards or outwards of the protoplasm that.even the nucleus is moved as well as the pigment granules, and the whole cell appears to change shape (Fig. 3-13a). If the pigment is dissolved away, fibres in these cells can be seen to cause the inward pigment movement by their contraction (Welsh, 1930; Fig. 3-13). It may be noted that physiologically the same effect 1s produced by this fibre contraction as by pigment dispersal in Cambarus (Fig. 3-12c), yet these would appear to be opposite reactions in terms of contraction of protein molecules. Much work on Palaemonetes and Leander has confirmed the action of a RETINAL-LIGHT-ADAPTING HORMONE, from the sINUS GLAND. This has been found to be similar to that in such Brachyura as Cancer and Uca (Kleinholz, 1936); but it is not clear if it is the same as RPDH of Cambarus. Evidence for an antagonistic, RETINAL-DARK-ADAPTING HORMONE has been found in Palae- monetes, and the richest extracts have been obtained from the TRITOCEREBRAL COMMISSURES (§ 2.112; Brown, Hines and Finger- man, 1952), A persistent diurnal rhythm of movement of distal retinal cells has been shown, at least in continuous darkness, for Anchistioides (Welsh, 1936), Palaemonetes (Webb and Brown, 1953), and Leander, Praunus and Pandalus (Bruin and Crisp, 1957); for this a hormonal control has been postulated. It is not inhibited by sinus gland removal; but it is probable that the sinus gland is, as in other cases, only the storage organ for the hormone, of which sufficient is still secreted from the source in the ganglionic-X- organ (§ 2.112) to maintain the rhythm. Changes in distal retinal pigment of grapsoid crabs (R. I. Smith, 1948) are very similar to those in prawns, with a marked § 3.222 EFFECTORS WITH MOVABLE PIGMENT GRANULES 81 Fic. 3-13. Two ommatidia from the compound eye of the prawn, Palaemonetes, in longitudinal section to show pigment cells which change shape. (a) In D, the ommatidium is shown with the pigment cells in the extreme dark-adapted position; in L, in the extreme light-adapted position. As in Fig. 3-12, the pigment in the distal retinal cells (d.r.c.) surrounds the cone (con.) in the dark position, and the rhabdome (rh.) in the light. In L, the pigment in the proximal retinal cells (p.7.c., not shown in D) has dispersed outwards from below the basement membrane (just out of the figure). (b) The same ommatidia with the pigment removed to show contractile fibres (c.f.) in the distal retinal cells (d.r.c.). They are attached to a mass of accessory pigment (ac.p.). In D, the fibres are relaxed in the dark-adapted position; in L, the fibres are contracted in the light. Most of the cell protoplasm and the nuclei (d.r.n.) have moved inwards with the pigment, leaving only an attenuated distal cell process (d.p.). The nuclei of the proximal or retinula cells (7.t.7.) remain stationary (from Welsh, 1930). diurnal rhythm that persists in constant darkness; but this is reduced or absent in continuous light. The rhythm can be induced experimentally to become out of phase with the time of solar daylight. About one-third of the dark-adapting hormone of the crab eyestalk is in the sinus gland, and two-thirds in the ‘‘optic G 82 KINETIC HORMONES—I ganglion”, which presumably indicates the ganglionic-X-organ as its source. The light-adapting hormone for the distal retinal pigment comes from the same source as that which concentrates the red chromato- phores of Palaemonetes (PLH § 3.223), and the two may be the same, since they both appear to cause contraction of protein molecules. Yet it requires 20 times as much crude extract to be effective on the retinal cells as on the chromatophores (Kleinholz, 1942). Insecta. The distal retinal cells in the compound eyes of in- sects can be divided into the same two types as in Crustacea; but the means of controlling their pigment migration is not known. 3.223 Pigment movement in chromatophores There is an embarrassing wealth of detail about the so-called chromatophorotrophic or chromactivating hormones from which it is difficult to select representative examples. They are interesting because they have similar physiological functions in both Crustacea and the cold-blooded vertebrates, and because similar methods of investigation have been applied to both groups, so that they can be directly compared. Chromatophores are usually elaborately branched cells which apparently remain stationary in the tissues, although they become PiaTE 3/1. Coloured photographs of the prawn, Leander serratus. (a) Dorsal view of the cephalothorax to show the pattern formed by the differentiation of the chromatophores into large red ones form- ing the stripes, with small red and white ones between ( x 4). (6) Part of the same, enlarged to show that there is more than one pigment in each chromatophore; the yellow component in the red chromato- phores can be seen faintly and the central red component of the reflecting, white chromatophore is clear. All are fully dispersed (x 50). (c) Two eyestalkless specimens kept on a white background, on which red pigment becomes fully dispersed. Half an hour before the photographs were taken each was injected with a different fraction of an extract of the sinus gland. That given to the upper specimen caused strong red pigment concentration; that given to the lower specimen had no effect. (d) Part of the tail fan of a specimen like the last, in which the maximum dispersion of red pigment is shown (from Knowles, 1955). (c) Fic. 3-14. Photomicrographs of melanophores from the web of the foot of the clawed toad, Xenopus laevis, with pigment in progressive stages of dispersion corresponding to the melanophore indices 1 to 5. Values such as 1-5 are sometimes recorded directly ; but they usually represent the mean of several readings (from Hogben and Slome, 1931) (b) (a) Fic. 3-18. Photomicrographs of chromatophores exposed by remov- ing a scale mid-dorsally from the killifish, Fundulus. (a) Black-adapted with melanophores dispersed and showing iridosomes at the centre of some; guanophores are concentrated and xanthophores appear grey. (b) The same after injection of ADRENALINE. The melanophores are concentrated, and the guanophores widely dispersed as they would be in a light-adapted specimen (from Odiorne, 1933). Fic. 3-19. Photographs of the male fiddler crab, Uca pugilator, in the light. The normal specimen on the left is dark; that on the right had the eyestalks removed 2 hours previously and the pigment has concentrated in the legs; the pallor shows best in the large asymmetric chela. The carapace is too thick to allow the colour change to be seen through it (from Carlson, 1936). Fic. 3-20. Photographs of the head of the sea-slater, Ligia oceanica, with the antennae cut off short. (a) Face-view; (b) lateral view of the head and eyes, differential illumination of which controls the release of chromactivating hormones (from Smith, 1938). 8 SS ee § 3.223 EFFECTORS WITH MOVABLE PIGMENT GRANULES 83 difficult to see (Plate 3-1) when their pigment granules ‘“‘con- centrate’” at the centre. The branches reappear as the pigment “disperses” to their extreme tips. The cause of this pigment migration has already been referred to (§ 3.22). The cells are sometimes named according to their pigments: “erythrophores” for those with red pigment; “‘guanophores”’ with white guanin; “melanophores” with black melanin, and ‘‘xantho- phores” with yellow pigment. Some cells have black pigments other than melanin. All these colours are to be found in vertebrates ; the Crustacea, especially the Malacostraca, have other lipophores, or coloured cells, with a blue pigment as well. Both may have iridosomes, with a movable reflecting material giving a blueish- white appearance (Fig. 3-18). The control of chromatophores in Crustacea is by hormones only, but can be either by nerves or hormones or a combination of the two in vertebrates. The type of control affects the response somewhat, since hormones, which reach all the chromatophores through the blood, tend to produce a slow and similar response in all parts of the animal, whereas nerve control can quickly stimulate individual chromatophores to produce a colour pattern that may match the background closely, as in Pleuronectidae and cham- eleons. Such an effect can only be simulated in those Crustacea in which the chromatophores themselves are much differentiated and each type responds to distinct hormones. ‘This seems to be the case in the prawn, Leander, in which some chromatophores are large and together form dark bands, while others supply a stippled and variable body colour. They include as least four colours and can be adapted to a variety of backgrounds (Plate 3/1), but their control is not yet fully elucidated (Knowles and Carlisle, 1956), and is too complicated to use as an example here. Among vertebrates, it is the chromatophores of all Agnatha, Elasmobranchii and Amphibia, but of only some species of Teleostii and Reptilia, that are under hormone control (Fig. 3-24). The chromatophores of the leeches, Hirudinea, concentrate in light and disperse in the dark, but show no background response. They are probably all under nerve control, and need not be con- sidered here. A typical background (or albedo) response is related to the 84 KINETIC HORMONES—I amount of light reflected from the background to the eyes. It helps to afford protective coloration to the animal, so that on a light background the result is a pale appearance; but to achieve this the dark pigments must be concentrated and the white dis- persed. It is a wide-spread phenomenon in both Crustacea and Vertebrata, and more often than not it is controlled by a pair of antagonistic hormones, which between them can maintain the pigment in the chromatophores at any position between full concentration and full dispersion. Observation on the behaviour of chromatophores has been facilitated by the introduction of a chromatophore or melanophore index, by which five stages of pigment dispersion are defined, from 1, fully concentrated, to 5, maximally dispersed (Fig. 3-14). Half stages can be used; but it is important to use chromatophores in the same region of the body for successive measurements, as different groups of cells can show considerable differences (Hogben and Slome, 1931). It must also be remembered that intervals on the chromatophore index scale are not quantitatively equal nor exactly related to the dosage of hormone needed to shift the pattern from one stage of dispersion to the next. Their representation by equal intervals on a graph can, therefore, be somewhat misleading. Before considering hormonal control of responses in more detail, it must be emphasized that several extraneous factors, other than “‘morphological colour change”’ (§ 3.2), can also affect chromatophores. The direct effect of light on chromatophores usually causes | dispersion (rarely concentration) in the absence of either nerve connections with a light receptor, or of any hormones in the circulation. Stephenson (1932) showed this clearly in the hermit crab, Eupagurus prideauxi, which has functional red and yellow chromatophores, not only on the exposed limbs, but also on the abdomen, where they are usually hidden within the whelk shell that it carries. The direct effect of bright light causes pigment dispersion, whereas the secondary effect, due to hormones stimu- lated by the eye, is to cause concentration of pigment. On an illuminated light background in a dull light, only this secondary response is elicited, and the animals are pale all over; but in bright light the pigment on the limbs disperses considerably. That this § 3.223 EFFECTORS WITH MOVABLE PIGMENT GRANULES 85 is due to antagonism between the direct dispersion and the second- ary hormonal concentration can be seen by quickly removing the crab from its shelter; at first, the newly exposed abdominal chromatophores are concentrated by the hormone effect only, but rapidly darken to match those on the limbs, as a result of exposure to the direct effect of the light. A similar direct effect occurs among reptiles. When a climbing lizard, Anolis, is blinded, its chromatophores show dispersal in the light and concentration in darkness (Brown, 1950a). The degree of dispersion usually in- creases with increase in the light intensity. ‘Temperature increase can concentrate the pigment in the white chromatophores of Palaemonetes (Fingerman and Tinkle, 1956) and thereby counteract its dispersion in response to a bright white background. This may help to keep the state of dispersion under control in very bright, hot conditions. In reptiles, such as Phryno- soma, concentration of dark pigments at high temperatures may help to prevent excessive heat absorption by the body. Moisture disperses some pigments, like those of Carausius (§ 3.221) and that in the melanophores of the frog, Rana, where the concentration due to a light background may be enhanced by dry conditions or almost completely overcome by total immersion. Other degrees of moisture give intermediate results. A rhythm of colour change related to the tides is said to persist for sometime, under still water conditions in the laboratory, in inter- tidal forms such as the fiddler crab, Uca pugnax. A diurnal rhythm of colour change persists under constant conditions of either light or darkness among other Crustacea, Insecta and Amphibia, and may be mediated by hormones in some cases, such as Uca pugilator. In experiments designed to show the action of hormones in controlling physiological colour change, care must be taken to control or eliminate these extraneous factors, affecting the reac- tions of the chromatophores. Once this has been done, two main lines of attack on the problem can be followed, though a combina- tion of the two is needed for its full elucidation. The first method is that due to Hogben and Slome (1931 and 1936), who studied the behaviour of the chromatophores in relation to changing environmental factors in intact Xenopus. This, which may be called the physiological method, they later 86 KINETIC HORMONES—I extended by the second, or pharmacological, form of investigation, using injections of organ-extracts and purified hormones. The second method has been that chiefly exploited in the search for crustacean hormones by Brown and his co-workers in America; they inject extracts of suspected endocrine organs into variously prepared animals, for which environmental changes are as far as possible ruled out. This method can eventually lead to the location of the source of the hormones, and since about 1942 it has been shown (Brown, 1944), for a steadily increasing variety of pigment cells with moving granules, that two hormones are in control (Table 9), although, for technical reasons, one is usually easier to demonstrate than the other. This, perhaps, accounts in part for the controversy which centred for so long around the problem of whether or not a second antagonistic hormone was necessary to account for crustacean colour change, or whether all the observed changes in any given animal could be accounted for by quantitative differences in a single hormone (Parker, 1948). The following account of the reactions of chromatophores and their control by hormones in Arthropoda and Vertebrata is only limited to red, white and black pigments in order to simplify the picture (Table 9); this is perhaps to oversimplify it, but it should suffice to give a frame of reference within which to consider further data, such as those relating to the complex system in Leander (Carlisle and Knowles, 1959). Much more information is still needed to extend the knowledge of most species from the realm of mere pharmacology towards an understanding of their natural physiology. Red pigment in chromatophores Crustacea. Physiological colour change in Crustacea is rela- tively slow in reaching equilibrium; but it can bring about an almost complete reversal of the state of some chromatophores in two or three hours. For instance, the following observations can be made on the red chromatophores of Palaemonetes, or of the fiddler crab, Uca (Figs. 3-15 and 3-19), if in the latter case it is remembered that the animals show a diurnal rhythm of colour change in daylight, so that observations must all be made over the same few hours of the day. One group of animals can be adapted 87 NT GRANULES — 4 4 EFFECTORS WITH MOVABLE PIGME § 3.223 sn4ogovy) DIN Dist'T OSDA) DIN DIsIT OSD’) DIN saqauomav Dg DIN saqauowmlavjDg DIN snansodnyq sajauowmavjvg DIN snansodnyq saqauowmavjvg TIdNVXa Hdl mG 2 Hd) H’ITfl me a | H’Td)O H’ILLO HdMf HdMd HOMN HOMd Hawn Hada Hdd Homan HTH *H'Id ANOWYOH ulvlg purj[s snutg é se SOINSSIUTUIOTD é — puvys snug ulelg puv[s snulg d — pues snutg é — SdINSSIWUUIOZ) purys snus Z 6c SoINSSTUIUIOT) SINSSTUIUIOZ) Z “cc puvjs snus NVOUO (VdOdOUH LUV) ALVUGALYAANI *AIBSSO[S 99S y DUDY ~—. sndouay — af (aArIou SNUIXOYT eta). SnANIAUL ah ae snurysoyars ch 2y (uIpoul vAJaquy'T -19}UT) ..&,, esiodsiq, vyinsupy dUT[eUsIPY sndouay — i (9Ar1oU SNUIXOYT snd) a (se snuiysoudagy | -I9qn J) ..M,, 93¥1}U99U07) (39v]q) sasoydounjajy “cc a3 M cS “ce snjnpuny | IO sUuI;VUSIPY osiodsiqi «e snjnpunsy 2 9}¥1]U99U07) (a314M) sasoydounnyy ce SNAJUAIOIO FT (AJUO 9AION) a SNUIXOYT UIPoUI9}UT osiodsiq “ce $N4£1UII0]0 FT (AJUO 9AION) eS SNUIXOY T sUI[eUsIPY 93¥1}U99U07) (por) sasoydosyz Gay 7 € AId NVXA ANOWYOH eqon amit ALVUGaALYAA LNAWNDId ATAVAOW HLIM SHYOHAOLVINOYHO ONITIOULNOOD SHNOWYOH OILANTS °6 TIAV T, 88 KINETIC HORMONES—I to an illuminated black background, on which the chromatophores will become fully dispersed (index 4.5), and another group to an illuminated white background, on which they will become fully concentrated (index 1). If the backgrounds are then reversed and observations are made on the state of the chromatophores on a chosen part of a leg or chela (the carapace of Uca is too thick to allow of satisfactory observation) at short time intervals, graphs of the average values for each group can be plotted to show the gradual dispersion of the chromatophores after the background change from white to black (dotted curve) and the concentration in the reverse case (full curve, Fig. 3-15a; Brown, 19500). The first conclusive evidence that such changes in the chromato- phores were controlled by a hormone was obtained by Perkins (1928), who showed for the prawn, Palaemonetes, that, although cutting the nerve supply to any part of the body does not interfere with its colour responses to a change of background, a ligature occluding the blood supply to that part stops the response, as in the stick insect Carausius (§ 3.221). Release of the ligature restores the response. Concentration of red pigment in Palaemonetes. 'The source and action of the red-concentrating hormone that causes the response to a white background has been identified in Palaemonetes by the pharmacological method. In the first place it is found that, in prawns and other Decapoda, except the Brachyura, removal of the eyestalks destroys the background response and leads to permanent dispersion of the pigment in the red chromatophores; extracts of the eyestalks can overcome this dispersion and lead to a temporary concentration of the pigment. Within the eyestalk the most potent source of the red-concentrating or Palaemonetes- LIGHTENING HORMONE, PLH, is in the sinus gland. Moreover eyestalk extracts from most Decapoda, except crabs, produce this pigment concentration, if injected into a dark, eyestalkless prawn; PLH is therefore not specific to any particular genus (Brown, 1950a). Much contradiction in earlier work was due to the fact that crude eyestalk extracts, containing PLH, were admixed with varying amounts of a second type of hormone that has a darkening effect on crabs. These two kinds of hormones were first separated § 3.223 EFFECTORS WITH MOVABLE PIGMENT GRANULES 89 by fractionation in alcohol (Brown and Scudamore, 1940); but much greater elegance and precision in separating pure substances from the extracts is now possible by paper electrophoresis (Knowles, Carlisle and Dupont-Raabe, 1955). So far the substances eluted Background changes Connective injection 3 = —* Intact 4 De e- Intact VA 3 See yf s ¥ Intact \_ fen -e—o = ow i Eyestalkless 5 (a) | () | l l S : arer © : Sinus _gland eee Sinus gland fractions = Comeioeeretd 5 i c a ae Intact (a + insol. e ( 7p gb CE ope Ts a, i a ° ee ean (ae ai eg mes Oe, 0: COS o Sol i / “4 ag ae a E seinen a8 ris 7 / y iv yr | -o-9? och (b) (d) O | (Za S fe) | 2 3 Time, hr Fic. 3-15. Reactions of the red chromatophores of the fiddler crab, Uca. (a) The natural background response: the rising curve (dotted line) shows the rate of change of red chromatophores after a light-adapted specimen is transferred from a white to a black background; the falling curve (full line) shows the converse change from black to white, in constant illumination. All measure- ments were made over the same time of day to avoid the influence of diurnal rhythmic changes. (6) The response of intact and eye- stalkless specimens from black backgrounds to injections of Uca- RED-DISPERSING HORMONE, URDH, from the sinus gland. Prompt dispersal occurs in the eyestalkless specimen; the dark intact specimen shows pallor first, presumably due to operative shock. (c) The response of similar specimens to injections of an extract of circumoesophageal connectives. The result indicates a short-lived action of Uca-RED-CONCENTRATING HORMONE, URCH, but is ambiguous. (d) The responses of eyestalkless specimens to in- jections of alcohol insoluble (above) and alcohol soluble (below) fractions of a sinus gland extract, to show that the former contains most of the dispersing hormone, URDH (all from Brown, 1950). 90 KINETIC HORMONES—I from different spots appearing on the paper have only been tested upon eyestalkless prawns kept on a white background; in these the red chromatophores are fully dispersed owing to the lack of PLH, reinforced by the dispersing effect of direct light. It follows that only light-adapting substances causing concentration of red pig- ment can be satisfactorily identified, though by using sufficiently concentrated extracts relatively rapid reactions can be obtained (Fig. 3-16). Other parts of the nervous system yield extracts with an action similar to that of PLH, but it is more than likely that chromatophore index time (min.) Fic. 3-16. Effect of extracts of the tritocerebral commissures on the dispersed red pigment in the chromatophores of eyestalkless prawns, Penaeus braziliensis. 'The effect of SINUS GLAND extracts would be similar. The abscissae represent time in minutes after the injections; the ordinates, the state of dispersion of the pigment on the same scale as the melanophore index. The dotted lines show that readings taken at night are closely similar to those by day, shown in full lines. The extracts from the post-commissure region (p.c.) are rather more active than those from the main commissure (c.) in yielding a PIGMENT-CONCENTRATING HORMONE, the effect of which is to cause rapid pigment concentration. This wears off as the hormone is destroyed in the tissues and the pigment returns to its original state of dispersion (from Knowles, 1953). § 3.223 EFFECTORS WITH MOVABLE PIGMENT GRANULES 91 these are precursor substances, rather than the natural hormone, since they often have mixed effects or need to be “‘activated”’ in some way, such as boiling or extraction in alcohol, before becoming effective (Figs. 3-15-17). Dispersal of red pigment in Palaemonetes. The red-dispersing or Palaemonetes-DARKENING-HORMONE, PDH, that acts antagonistic- ally to PLH, has been difficult to locate because it requires a test animal that is pale, yet contains no source of an overriding, concentrating hormone. This has been achieved by starting with eyestalkless specimens of Palaemonetes in which the red pigment is fully dispersed (index 5); then at time 0 (Fig. 3-17) an extract containing PLH from either sinus gland or tritocerebral commis- sures is injected, and pigment concentration becomes almost complete in 15 min (index 2 or even 1.5). After this, the effect wears off slowly as the hormone is destroyed in the tissues and the chromatophores gradually return towards full dispersion. The rate of this return is unaffected by an injection of sea water at 30 min (Fig. 3-17, curve i). If, instead of sea water, an injection of an extract of the darkening hormone, PDH, from either abdomi- nal ganglia or circumoesophageal connectives is given, the rate of dispersion is greatly increased (index 4 is reached in 15 min), and may continue until full dispersion is achieved, within an hour from the start of the experiment (Fig. 3-17, curve ii). This shows conclusively the presence of PDH in the extract, whereas testing this extract on eyestalkless animals which had not been pre- treated and were therefore dark, though consistent with this interpretation, does not by itself prove the activity of the extract (Fig. 3-17, curve iii). If PDH is injected into normal, eyed animals with pigment concentrated in the light, only a slight dispersion is produced and this is quickly followed by a return to full concen- tration, showing that the action of PDH is unable to overcome the naturally secreted PLH (Fig. 3-17, curve iv). To allow for the natural reversal of the chromatophore response within a relatively short time, a further elaboration of the two- hormone hypothesis has been postulated, namely, that in response to a black-to-white background change, a large amount of PLH is ‘discharged suddenly into the blood, but that ‘‘as adaptation becomes complete there would be a reduction of the hormone 92 KINETIC HORMONES—I level to some lower maintenance one’’. Conversely, a white-to- black change would result in an abrupt discharge of PDH, which would become similarly reduced to some lower level as adaptation became complete. ‘The introduction of large amounts of either factor, in the presence of a somewhat lower titre of the second, yr ARS | Eyestalkless Injections: o = Tritocerebral (PLH) 0 = Abdo: gang: (PDH) e = Seawater Eyestalkless Av. chromatophore index iv Intact @) | 2 =) Time, hr Fic. 3-17. Effects of extracts containing Palaemonetes-LIGHTENING and -DARKENING HORMONES, PLH and PDH, on the red chromato- phores of the prawn, Palaemonetes. Pigment dispersion is shown with time in hours, on a much less extended scale than Fig. 3-16; the extracts are less concentrated and act more slowly. Curves 1, ii and iii show effects in eyestalkless specimens with initially dis- persed pigment: curve i: the effect of PLH injected at time 0 wears off and is unaltered by subsequent injection of seawater at the time marked by the arrow; curve ii: PLH injected at time 0 followed by PDH, instead of sea water, injected at the arrow, shows the rapid dispersing effect of PDH; curve iii: PDH injected at time 0 pro- duces a slight concentration due to shock and a rapid return to full dispersion. This last is compatible with a dispersing effect of PDH, but does not prove it alone. Curve iv shows that injection of PDH only partly overcomes the natural supply of PLH in a normal specimen exposed to light on a white background. PLH was obtained from sinus glands and tritocerebral commissures. PDH was obtained in least contaminated form from the abdominal ganglia (from Brown, Webb and Sandeen, 1952). § 3.223 EFFECTORS WITH MOVABLE PIGMENT GRANULES _-93 effects an initial response which is practically that which would be achieved in the complete absence of the second. Such a mechanism would assure rapid response in either direction, provided the time elapsing between the responses is adequate for the normal reduc- tion in titre after the initial secretory burst”—i.e. about 15-50 min in the black-to-white change, and 30-60 min in white-to- black (Brown, Webb and Sandeen, 1952). Eupagurus. 'The red pigment in the hermit crab, Eupagurus, (Table 11) behaves like that in Palaemonetes ; it is concentrated in bright light on a pale background, and dispersed on a dark back- ground. It is clear that both reactions in Eupagurus are under hormone control, but the actual substances and their exact sources have not been determined, although removal of the eyestalks again results in persistent darkening. Uca. The background responses of the red pigment in the chromatophores of the fiddler crab, Uca, show well and have already been referred to (Fig. 3-15a). Both red-concentrating and -dispersing substances can be extracted from nearly all parts of the nervous system; but, as in most brachyuran crabs, the sinus gland is the richest source of Uca-RED-DISPERSING HORMONE, URDH (Fig. 3-155 and d). As the same is true of the hormone dispersing the melanophores, removal of the eyestalks of crabs results in permanent pallor (Brown, 1950). This is unlike the situation in prawns and most other Malacostraca that have been examined, where the sinus gland provides a concentrating hor- mone, like PLH, and removal of the eyestalks therefore results in permanent darkening. The main source of the Uca-RED-CONCEN- TRATING HORMONE, URCH, is in the circumoesophageal connec- tives (Fig. 3-15c) and this is again the opposite site from that in the prawns (Tables 9 and 11). The sinus gland also yields small quantities of URCH. On a constant white background Uca also shows a diurnal rhythm of colour changes that are comparable to the direct effects of light and darkness on most dark chromatophores. The crabs become dark by extensive pigment dispersal by day, and pale at night; but these changes in Uca are not merely direct effects (p. 84), since their rhythm persists for four days in constant dark- ness, and it is not altered, although the amplitude of the pigment 94 KINETIC HORMONES—I dispersion is reduced, by reversal of day and night illumination (Webb, Bennett and Brown, 1954). VERTEBRATA. Relatively little is known of the control of red pigment in vertebrates. TELEOSTEI. The red (and also the yellow) chromatophores of the minnow, Phoxinus, appear to be under purely hormonal con- trol and to show the usual background responses. The dispersing action of INTERMEDIN, B, from the pars intermedia (§ 2.123), is well established (Giersberg, 1930 and 1932), but the source of a red-concentrating hormone has only been rather uncertainly located in the epiphysis or pineal organ. Adrenaline has no effect. Other red pigment cells of fish, such as Holocentrus, are controlled by nerves only. RepTILia. The erythrophores of the chameleon, Lophosaura, are probably under nerve control, like their melanophores, but they do not seem to have been fully investigated (Brown, 1950a). White pigment in chromatophores Crustacea. The hormonal control of other pigments in chromatophores is essentially similar to that already described for red pigments, though many minor variations have been found (such as those between Palaemonetes and Crago), and much remains unknown (Tables 10 and 11). In bright light on a white background the response of guano- phores is usually dispersion of their white pigments, which reinforces the pallor produced by the concentration of dark pig- ments in the other chromatophores. In all the Decapoda exam- ined (except the Brachyura), the richest source of extracts which induce these light-adaptations is the SINUS GLAND, and this is almost certainly the organ where the natural hormones are liber- ated into the blood; for eyestalk (and therefore sinus gland) removal causes the opposite effect. Like the natural response to an illuminated black background, it results in the white pigment becoming concentrated and the dark pigments being dispersed. Extracts causing the latter effects can usually be obtained from the commissures (which include circumoesophageal connectives and tritocerebral commissures); but it is not known for certain at what point the natural hormone passes into the blood. § 3.223 EFFECTORS WITH MOVABLE PIGMENT GRANULES 95 Palaemonetes. The white chromatophores (Table 10) reinforce the protective colour change provided by the red pigments of this prawn (Brown, 1950a). The concentration of white pigment, which occurs naturally on a dark background, can be induced by injecting an extract from the commissures. This contains a Palaemonetes-WHITE-CONCENTRATING HORMONE, PWCH. The dis- persion, that occurs on a white background, can be induced by a Palaemonetes-WHITE-DISPERSING HORMONE, PWDH, from. the sinus glands. The direct effect of light on the skin is to cause pigment disper- sion in these white chromatophores, as it does in erythrophores; but here it reinforces the hormone reaction controlled by the eyes, instead of counteracting it (‘Tables 10 and 11). Crago*. 'The hormonal control of white chromatophores in the shrimp follows the same pattern as that in Palaemonetes. The sources of the two hormones, CWCH and CWDH, are probably similar, but this is not certain. There appears to be no direct effect of light upon the guanophores in the skin, for they remain con- centrated, whatever the light intensity. Uca. In Brachyura, the control of the white chromatophores, like that of the red and black cells of the same animals, follows a pattern distinct from that of the other Decapoda. Although the background response is the same as in the prawns, the source of the Uca-wHITE-DISPERSING HORMONE, UWDH, cannot be mainly in the sinus gland, as theirs is, since the pigment remains perma- nently dispersed in eyestalkless animals. The latter observation suggests that, like the Uca-red-dispersing hormone, the source of the Uca-wHITE-CONCENTRATING HORMONE, UWCH, might be in the sinus gland; but extracts have not so far given any concen- trating effect. There is apparently no direct effect of light upon the white chromatophores of Uca, which remain permanently dispersed, even in the dark, in the absence of hormonal control. TELEOSTEI. The white chromatophores, or guanophores, of the killifish, Fundulus, show the same adaptive reaction to background colour as do those of the Crustacea, dispersing on a white back- ground and concentrating on black (Fig. 3-18 a and 6). ‘Their * This spelling of Crangon is commonly used in this context in America, and is retained here for simplicity. KINETIC HORMONES—I 96 Poyeorpul st pueys snulg { * %& * % * * * * * * * % tHOMN HdM/!) * * * % das dss Loge LOTdIa GASNOdSaeY SSH TA TV.LSAA HOMO ¢é HAMO * * * * “Aiessojs 990G | 4HdMd * * * * ASNOdSdy GANNOWOMOVE TWUNLVN DIN) OsDL sajauowmavjvg SHNOWYOH AGNV SHYOHdATOLVNOYHOD *po}B1}UIDUOD = §¢ § uMOUYUL) So.INSSTUIWIOZ) uoT}JeIZUIIUOT) uMOU UL) puv]s snutg uorsiodsiq ANOWYOH AO AONNOS wo NOILOV * * pesiodsip = Jy 1432] 11M 1y8Yy 1YysUg uoypununyzt Aupy 7481) 1]Np 40 ‘YID]q uo 148y JYysiUg aq1ya2 uo JY48Y IYSUg GQNNOWOMOVEA GNV NOILYNINOATII LHOIT OL NOILVYTHY NI LNANDId FLIHM ONITIOULNOO SHNOWYOH NVAXOVLSNY) “OT ATAV T, § 3.223 EFFECTORS WITH MOVABLE PIGMENT GRANULES 97 normal control may be by nerves, as is that of the melanophores of this fish; but both types of chromatophores react rapidly to injections of either ADRENALINE, or Antuitrin, that supplies a MELANOPHORE-CONCENTRATING HORMONE, MCH (Odiorne, 1933) to give the “light reaction” in which melanophores concentrate, and guanophores disperse (Fig. 3-185). Although denervated melanophores of Fundulus are known to disperse in response to injections of B (or MSH), there is only slight evidence that this causes the expected concentration of the guanophores. Black pigment in chromatophores Crustacea. The shrimp, Crago, is the only member of the Decapoda Natantia so far investigated that has black pigment akin to melanin in cells that may be called melanophores. These melano- phores can produce the beginnings of a pattern, because they are differentiated into two sizes, larger on the body and smaller on the tail. ‘These both react to one Crago-DARKENING HORMONE, CDH, from the commissures, but are concentrated independently (Brown, 1946 and 1950a); those on the tail by Crago-TaiL- LIGHTENING HORMONE, CTLH (alcohol-insoluble extract from the SINUS GLAND), and on the body by Crago-BoDY-LIGHTENING HORMONE, CBLH (water-soluble extract from the BRAIN), both of which are dominant to CDH. In nature, this means that the shrimp can go wholly dark if only CDH is acting, or the tail or body only may be lightened, while the rest remains dark, accord- ing to which of the concentrating hormones are present with the CDH. Finally, if both CTLH and CBLH are present, the shrimp becomes completely pale. An eyestalkless specimen lacks CT'LH, and is at first pale with a dark tail, but after a time this effect dis- appears. The natural stimulation of pattern forming changes in the shrimp is not known; but in prawns, like Leander, the exten- sion of a similar system to a large number of different types of chromatophores, apparently each controlled by separate hormones, must give them adaptive advantages by increasing their ability to match a variety of backgrounds (Plate 3-1). Uca. The background responses of Uca melanophores (Fig. 3-19) are more difficult to demonstrate than those of the red chromatophores, because they are almost completely overridden H “(9S6] ‘HISITUVD pUuv SHIMONY]) 4apuvaT jo y oq Avur : vinAyovig ur jUaseid osje = »v ‘soroydojewosys ut ‘passodsip Ayjezed = (* *) ‘posiodsip = * * ‘pazerjyua0u0s = ¢ ¢ 3k * “epodos] 94} 03 1877 pur ‘einkyoeig epodesaq 94} 0} v2-9 ‘eInwiouy epodessq 94} 03 snansodng ‘eryueye Ny epodesoq sy} 0} Osp4D pur sajauomanjog :edVs\SOoR|LJA, 2YI 0} Buojoq TT qk], ur sajduexe |TV re a ee ey ee i Sale a4 é he -' 1484 11NC ({1e3) * (* *) ate * * * ok 1Y48Y 1YsuUg * * % hs * * ok * LOddda LOTIG 7 (1123) ie “ie — ae atin ws yg uolnoUuuunzye Kupy 7, * ok * * (e) Ls ASNOdSau SSATHIV.LSAAT m o) 2 gHdanh a = Hawn = = puvjs snuig ) = = — a Hdd nae umouyuyl) fs —_— ¢ Hal Hdo — — eHdd SoINSSTUTWUOZ) rea) a * * * Ok * * * d ae a La * SI ‘49019 UO 4 * %* 1k * * OK * ok eel aiigee © OE EL ee 1 faye = == HOUN ae a uMmouyuys) — — »H'1dO =— = or pretal — HT Hit: | (oun) oer oH’Id pues snurg ( =) Her ous ee Ria6 ey U01}81]UI9U0Z a414%2 UO 481) JYsiUAg ASNOdSaud ANNOYDMOVA IVHALYN Ss Fs RN ae De AE I ee I Ree Ae) Op or aS | eee fore) DI) DIBVT osv4) DI{) ‘soqdny “Mav ANOWUOH JO FOUNOS GNNOUDNOVA ANY s sOVIa am uO NOLLOV NOLLVNIWOATI a LHOIT OL NOILWTHY NI SHHOHAOLVINOUHO NI SLNAINDId MOVId GNV Ga ONITIOWULNOO SHNOWYOH NVAOVLSNUT) “II ATAV § 3.223 EFFECTORS WITH MOVABLE PIGMENT GRANULES 99 by diurnal changes, which operate in the opposite sense. If, however, allowance is made for the diurnal changes, it can just be seen that, for any given intensity of light incident upon the background, the degree of dispersion of the black pigment is greater on a black background than on a white background (Brown and Sandeen, 1948). The Uca-DARKENING HORMONE, UDH, controlling dispersion of the melanophore pigment, like the Uca-red-dispersing hormone, URDH, is secreted from the sinus gland; eyestalk removal therefore results in relative pallor. The dispersing activity of UDH can be neatly demonstrated by using the two sexes of these crabs (Fingerman and Fitzpatrick, 1956). Melanophores in the female are normally more dispersed than in males in the same situation. If the large, hollow, asym- metrical chela, which distinguishes the male from the female, is removed before the crabs are exposed to light, the operated male becomes as dark as the female. If more legs are removed he be- comes even darker. ‘This can best be interpreted by assuming that the same amount of dispersing hormone, UDH, is released in each animal in response to similar stimuli; but that the degree of disper- sion of the chromatophores depends upon the concentration of the hormone in the blood. This is increased as the blood volume is decreased by removing successive appendages. The natural secretion of Uca-LIGHTENING HORMONE, ULH, has been demonstrated by using the blood of crabs, in which the melanophores were maximally concentrated, to perfuse isolated limbs on which the melanophores were dispersed. ‘These show slow pigment concentration even if perfused with sea water; but the rate of concentration is increased if perfused with blood containing ULH (Fingerman, 1956). The source of ULH has not been located; but, unlike CTLH, it cannot be in the eyestalk. Ligia. Apart from the Decapoda so far described, the only crustacean that has had its melanophores investigated in detail is the sea slater, Ligia, among the Isopoda. In nature it changes colour from black, when lurking in damp and shady crevices, to pale mottled grey, when exposed to light. On any black background in bright light they show the usual response, by which the melano- phores become fully dispersed (index 5). On an experimental white background the pigment can become concentrated to an unnatural 100 KINETIC HORMONES—I pallor (index 1.5, Figs. 3-14 and 3-20). In darkness the melano- phores assume an intermediate condition (index 2.7). These reactions are similar to those of the red pigment of Palaemonetes (Table 11). | Blinded animals show a direct effect by which the melanophores are more expanded the brighter the light, but never as much as in normal animals on a black background in the same light (‘Table 12). If different groups of the ommatidia in the sessile compound eyes of these animals are illuminated separately, either by painting over part of the eye with opaque varnish (which affords a. good class demonstration) or by exposing the animals in specially constructed boxes which admit light only to certain narrowly delimited retinal areas (D, L, and V, Figs. 3-20 and 3-21), the effects shown in Table 13 can be obtained. It is concluded from these and other observations that two antagonistic hormones are involved: a Ligia-DARKENING HORMONE, LDH, normally stimulated by direct light on area D to cause dispersion; and a Ligia-LIGHTENING HORMONE, LLH, stimulated by reflected light on areas L and V to cause full concentration. When the whole eye is illuminated both are secreted, but LLH overrides LDH to produce almost complete concentration. A TABLE 12. CHANGES IN MELANOPHORE INDEX IN LIGIA All index values are an average of measurements made on the posterior part of the body of 24 specimens (from H. G. Smith, 1938). ANIMAL BACKGROUND BRIGHT LIGHT DIM LIGHT DARKNESS Normal Black 50 +0 46 +008 2:7 + 0-1 Normal White 1-7 + 0:08 144+ 0:06 2:7 + 0:1 Blinded White 4-2 + 0:06 3:9 + 0:09 2:7 + 0-1 balance between the two hormones produces an intermediate effect (Fig. 3-21b). The sources of the two hormones have not been fully determined, but Kleinholz (1937) showed that removal of the whole head is followed by dispersion, and extracts of the head cause concentration of the melanophores. The source of § 3.223 EFFECTORS WITH MOVABLE PIGMENT GRANULES 10] Light activating tDH No LLH secretion Reflected light activating some LDH Light activating LLH No LDH - secretion Light activating LLH only Fic. 3-21. Diagram of Ligia, in face view (cf. Fig. 3-20a), to show a method of illuminating separately the dorsal (D), lateral (L) and ventral (V) areas of the compound eyes. Arrows indicate the direc- tion of light falling on the eyes, which are exposed by cutting short the antennae (An.ii). The animals are confined in shallow cells allowing horizontal movement only. Screens, indicated by black- ened areas, prevent the entry of light. Stippling represents the chromatophore reaction to secretion of LLH, Ligia-lightening hormone and LDH, Ligia-darkening-hormone. (a) Illumination from above only. (%) Illumination by direct light from below, with reflected light from above. (c) Illumination from below only (Original diagram, based on Smith, 1938). 102 KINETIC HORMONES—I LLH is therefore probably in the organ equivalent to the sinus gland, since isopods lack eyestalks and the gland lies within the head capsule, in the vicinity of the optic centres, where it is associated with a blood sinus (Hanstrém, 1939). The source of the darkening hormone is unknown, but must be, in part at least, behind the head. TABLE 13. ILLUMINATION OF DIFFERENT AREAS OF THE EYES OF LIGIA Blue light is most effective in eliciting these responses in Ligia. The index values are averages (from H. G. Smith, 1938). AREA MELANOPHORE | CONVERSE AREA | MELANOPHORE ILLUMINATED INDEX COVERED INDEX D 4-7 en oe 4-6 Da 2:5 (dark) 27 DY 2°4 — — Ve 1-8 a —- ee bes 1-4 * 1-5 DV A<7 none Ley (normal) * illuminated from below. InsEcTA. The black pigment cells of the phantom midge larva of Chaoborus (= Corethra) differ from those of Carausius in being mesodermal. They normally cover the surface of the two pairs of air sacs and show a background response of the usual protective type; but concentration of pigment on a white background is brought about by an amoeboid change in shape of the cells which become spherical instead of elongated. The cells also tend to aggregate in small groups instead of being evenly spread out to show the maximum amount of colour, as they do on a dark back- ground. Extracts of the brain cause pigment dispersion, and therefore darkening, as in Carausius (Dupont-Raabe, 1956). VERTEBRATA. Melanophore control varies in different fish; among Teleostei it is only in some species that it is wholly under hormone control or even partially under hormones and partially under nerves. Amphibia, where the control is purely hormonal, will be considered first. § 3.223. EFFECTORS WITH MOVABLE PIGMENT GRANULES 103 AMPHIBIA. ‘The background responses of Amphibia are similar to those of the Crustacea in so far as the animals become pale on an illuminated white background and dark by melanophore dispersion on an illuminated black background. Early investiga- tions of these colour changes were made on the African clawed toad, Xenopus and attempted to use the different rates of change of the chromatophores, following changes of background or changes to and from darkness (Hogben and Slome, 1931), to indicate the number of hormones that were necessary to control the changes. ‘The observations were good in that they did not interfere with the integrity of the animals, but recorded their natural physiological reactions. But the changes were so slow and were interfered with by extraneous factors like diurnal rhythms and unsuitable temperatures, so that the results were unsatis- factory. Recourse to the pharmacological method of injecting extracts into variously prepared specimens was therefore necessary to obtain conclusive evidence for the presence of two hormones. The dispersing or MELANOPHORE-STIMULATING HORMONE, MSH, (known as B to the earlier writers) can be easily established by injection of extracts of the posterior lobe of the frog hypophysis, although the early experiments made use of mammalian extracts. The source is the pars intermedia (§ 2.123). It has not been possible to prepare active extracts of the antago- nistic CONCENTRATING HORMONE, known as W and believed to be secreted from, or controlled by, the PARS TUBERALIS (§ 2.123). The best evidence for its presence is obtained by injecting equivalent extracts of active B substance into variously prepared test toads, and comparing results on a white background (Hogben and Slome, 1936). Injecting a standard dose of B into the normal pale animal produces a temporary increase in melanophore dispersion that wears off in about 10 hr, when the toad’s normal response again takes charge (Fig. 3-22, curve B). If the whole hypophysis is removed, including both the known source of B and the postulated source of W, a much greater effect (Fig. 3-22, curve A) is produced for the given dose of B than in the intact animal. The effect also persists longer, just as if the natural supply of W were absent. Melanophore index 104 KINETIC HORMONES—I If the posterior lobe of the pituitary is removed (taking with it the pars intermedia, which is the source of B, while the pars tuberalis is left intact) a similar dose of B results in the least and shortest response of the three (Fig. 3-22, curve C). If there were no ats D Total removal : Pars tuberalis é : 3) If Posterior lobe f Ves \* Normal (white background) | : ms E Posterior lobe C only removed If | 0 2 4 6 8 10 12 14 16 IS Hours Fic. 3-22. Effects upon the melanophores of Xenopus of extracts, containing equal amounts of the melanin dispersing hormone, B (intermedin). (A) Six toads from which the whole hypophysis, including sources of both B and W, has been removed. (B) Six normal intact toads, in which production of both hormones is stimulated by the white background. (C) Six toads from which the posterior lobe and the pars intermedia have been removed, taking with them the source of B only. For further explanation see text. B was obtained from extracts of ox “‘posterior pituitary”, freed from pressor and oxytocic activity, but including intermedin from the pars intermedia to provide a dispersing effect. Although this is not the natural amphibian hormone, the facts of the dose being the same in each case and producing slow dispersion, and the fact that the magnitude of the effect is altered by prior operations on the toads make the results appear significant. The specimens used for the experiments were “‘selected for the same degree of pallor’, and all were exposed to an illuminated white background (from Hogben and Slome, 1936). active hormone secreted by the pars tuberalis, this curve should be the same as curve A, since the source of B is absent in both 2) § 3.223 EFFECTORS WITH MOVABLE PIGMENT GRANULES 105 cases. The difference may therefore be attributed to the secretion of a large amount of the concentrating hormone, W, by the pars tuberalis in response to the white background (Hogben and Slome, 1936). The toads always remain pale on a black background after removal of the source of B in the posterior lobe of the hypophysis. If the whole adenohypophysis, including the pars tuberalis, is removed, the toads become permanently dark. That this is due to the loss of the source of W is claimed from finding that, after slightly incomplete hypophysectomy of Xenopus, the pars tuberalis may regenerate, in which case the lost response to a white back- ground is regained (Waggener, 1930). As in Ligia, it is claimed that the adaptive colour changes are controlled by the secretion of the two hormones in different pro- portions in response to illumination of different parts of the retina of the eye by direct and reflected light (Fig. 3-23). The eyes of Xenopus are on the dorsal surface of the head and direct light stimulates the “floor” of the retina; on a black background no other light reaches the eye and the secretion of the DISPERSING HORMONE, B, is induced. Scattered light from a white background stimulates the ‘“‘peripheral”’ part of the retina and induces secretion of the CONCENTRATING HORMONE, W, whatever position the toad adopts (Hogben and Slome, 1936). In the eel, Anguilla, where the eyes are lateral, the ventral and dorsal parts of the retina are as- sumed to play the same réle as floor and peripheral parts in the toad. These results seem to be conclusive; but they lack control injections of saline or of some other non-active substance, and they have not been confirmed by more recent work using better techniques. It is possible, therefore, that there is no real difference between Xenopus and the frog, Rana pipiens, in which there appears only to be the one melanophore-dispersing hormone, B (Parker and Scatterty, 1937), concentration of melanin resulting merely from absence of B. ELASMOBRANCHII. The background responses of the melano- phores of the rough dogfish, Scyliorhinus, and other elasmo- branchs are similar to those of the Amphibia, but they seem to be even slower (Waring, 1938). There is no doubt that the melano- phores are dispersed by B, since this can be extracted from their 106 KINETIC HORMONES—I own pars intermedia and can also be detected in effective quantities in the blood of dogfish that have been kept ona black background. There is still uncertainty about the presence of the antagonistic, concentrating hormone, W, which is always more difficult to Light Light CNS. Eye BLACK Melanophore Dispersed ‘Concentrated Fic. 3-23. Diagram of the nerves and hormones concerned in mixed control of melanophores in the eel, Anguilla. Similar hormones act alone in an amphibian such as Xenopus. An illuminated black background is represented on the left and an illuminated white background on the right. On black, only the floor of the retina is stimulated by light and causes pigment dispersion, either by stimulating the secretion of B (MSH) from the pars intermedia (p.in.), or by adrenergic nerves (a.n.). On white, both the floor and the upper part of the retina are stimulated and pigment is concen- trated, either by secretion of B as before and of overriding W (MCH) from the pars tuberalis (p.t.), or by cholinergic nerves (c.n.). In either case nerves from the eye pass through the brain (C.N.S.) to transmit stimuli from the retina to the chromactivating hormone or nerve (from Parker, 1943). detect. A full review of the literature relating to the chromatophores of these and other fish has been given recently by Pickford and Atz (1957). TELEOSTEI. The control of melanophores in the teleosts is more § 3.224 EFFECTORS WITH MOVABLE PIGMENT GRANULES 107 complicated and varied than in the other groups considered, because there appear to be at least two distinct types of response to hormonal stimulation, as well as the differences in reaction introduced by mixed control by both hormones and nerves. In some of the more primitive forms, such as the pike, Esox, and the carps, eels and Ameiurus, the control appears to be purely hor- monal. ‘The melanophores disperse in response to the MELANO- PHORE-STIMULATING HORMONE, MSH, from the pars intermedia, when it is injected into pale specimens, whether the source of the hormone is from the same fish, from amphibia or even from mammals. ‘The majority of other teleosts like Fundulus, are in- sensitive to MSH, at least until the melanophores are denervated. This second group is, however, found to be more sensitive to a MELANOPHORE-CONCENTRATING HORMONE, MCH, which is present in most extracts of fish adenohypophyses. It is present in Antui- trin and has no action on Amphibia. A few teleosts, like Phoxinus, although having their melano- phores normally under nerve control, can be induced to respond appropriately to both MSH and MCH under suitable conditions. Hormonal control in Phoxinus apparently serves to maintain the melanophore reactions over long periods when the nerve control becomes fatigued (Healey, 1948). Many other teleost fish have melanophores which are controlled only by nerves (for further details see Pickford and Atz, 1957). RepTILia. Hormones, especially INTERMEDIN, B, play a part in the dispersal of the melanophores of certain of the older genera of lizards, such as Phrynosoma, Hemidactylus and Anolis; but con- centration appears to be controlled by ADRENALINE and not by a pituitary secretion. Dual nerve control replaces this in the more highly evolved chameleons and others, producing varied patterns of colour change. 3.224 Discussion of the hormonal control of pigmentary effectors Looking back over the pigmentary effectors just considered (Tables 9, 10, 11), it seems possible to make some generaliza- tions, although the more complex cases have been left out, and the evidence is not always complete for those that have been included. Two hormones are concerned in the control of nearly all types of 108 KINETIC HORMONES—I pigmentary effectors with movable granules, including some which have nerve control as well. But in nearly all cases the actions and sources of one hormone are much more fully established, than those of the other; the more certain of the two is the light-adapting hormone for Crustacea, and the fishes Phoxinus and Fundulus, whereas it is the dark-adapting hormone for the Amphibia, Elasmobranchii and some Teleostei, such as Ameiurus. It seems plausible to expect that in the stick insect, where so far only the Carausius-darkening hormone has been found (§ 3.221), an antagonistic pigment-concentrating hormone will be revealed in due course. In Crustacea there is evidence from Isopoda and from many Decapoda, other than Brachyura, that the hormones producing the light background response are normally stored in, and pre- sumably discharged from, the sinus gland or its equivalent; this is true for the concentration of dark pigments such as black and red, and also for the converse dispersion of the white pigment (Tables 10, 11). Similar converse reactions of the pigment in coloured and white chromatophores in fish may also be due to one and the same hormone, as in Fundulus. The situation in the Brachyura is peculiar. There is a background response, which though slight is similar to that in prawns; but, instead of the concentrating hormone, it is the dispersing hormone which is released from the sinus gland in the eyestalk. In many crabs, such as Uca, the background response is overridden by an opposing reaction causing darkening in bright light. It is tempting to interpret this as being either (1) a morphological accident the result of which is that, when light stimulates the sinus gland via the eye, the gland releases its hormone, as in other decapods (but this produces the opposite effect in crabs because the secretion is a dispersing instead of a concentrating hormone); or (ii) the effect of a single hormone controlling dispersion of pigment in both the melanophores and the retinal cells; for dispersion of retinal pigment, as in Cambarus, is the adaptive response to light and the daytime dispersion of the crabs’ melanophores would then be but following in the wake of their retinal pigment. It must be emphasized, however, that none of the cases consid- ered (except Crago) have more than one colour of pigment in § 3.224 EFFECTORS WITH MOVABLE PIGMENT GRANULES 109 the same chromatophore, unlike the complex pattern-forming chromatophores of Leander and Penaeus (Knowles, 1955; Knowles, Carlisle and Dupont-Raabe, 1955, and Knowles and Carlisle, 1956), in which there may be as many as four. It is to be hoped that the refined technique developed by these authors for separa- ting pure substances from tissue extracts by paper electrophoresis will soon be extended to the circulating blood of these and other prawns, with their chromatophores in different states of dispersion in response to different states of illumination or background colour. When extracts of tissues yield the same substance as that found to be active in the blood, it should be possible to identify the sources of hormones actually used by the animal, and to distinguish these from other substances, like acetylcholine, pro- duced at nerve endings in the central nervous system; for these can react upon effector systems experimentally, and yet never reach them through the circulation in the living animal (Knowles, 1955). Modern techniques might also throw light upon the rate of chromatophore reaction, which seems normally to be so much slower in the vertebrates than in the Crustacea, and to be controlled to a considerable extent in the latter by the concentration of the hormone reaching the cells. The rates could be compared with the high speed of reaction of nerve-controlled chromatophores, to see if the differences were due to the more concentrated dose of the chemical which can be supplied at a nerve ending, rather than to differences in sensitivity of the chromatophores (Waring, 1942). Nevertheless, Waring’s idea, that an evolution in either the sensitivity of the chromatophore or in its speed of reaction was a necessary precursor of the evolution of nerve control with adaptive value, is interesting. It appears to be borne out to some extent by his table showing that nerve control is only of importance in vertebrates of more recently evolved families, and has not been achieved in the Elasmobranchii and Amphibia, which are both classes with a much longer fossil history than either the Teleostet or the Reptilia (Fig. 3-24). The persistence of purely hormonal control in Crustacea would then be expected, and could be looked upon as having evolved along lines of chromatophore differenti- ation with multiple hormone control, instead of being replaced by nerve control. 11 an) KINETIC HORMONES—I Ww @ @ a id — Oo a3 : ree : nt Gi = g 3 2 rt Ss Oo ee es a Mic “— _ = _ =! w 2p = 2 2.9 oie 2 aed 3.0 Ee ¢ ee eS oS 5 re 22936 oa So~e® x a oS et lS Ej Be i ee) Boas ave tie ® o@ad2 pete — aS 5. 8 8 ue SeegEB2 88 z 8 “Hef 2 = SO Le ea, GO SS c 2] o a co: 5 = Bo] ° = & Oi o Cc = Sy ie eS Mie See eee 2 ate AOnne qnton OSs stimulates CMM inhibits jejunum enterocrinin ileum Fic. 4-1. Diagram to show the actions of hormones in the mam- malian gut. Heavy arrows extend from the source of each hormone to the effector which it controls, either by stimulation or inhibition. SECRETIN stimulates the secretion of bicarbonate; PANCREOZYMIN, DUOCRININ and ENTEROCRININ the secretion of enzymes. In addi- tion to their effects upon the secretion of HCl, GASTRIN and ENTEROGASTRONE stimulate and inhibit respectively the muscles of the stomach. CHOLECYSTOKININ only stimulates the muscles of the gall bladder and relaxes its sphincter (amplified from ‘Turner, 1955). in an increased flow of acid, but in little change in the pepsin content of the gastric juice. The secretion of GASTRIN is therefore stimulated either by food or by mechanical distension; it is secreted from the mucosa of the § 4.111 DIGESTIVE GLANDS 121 stomach, chiefly in the pyloric region, and stimulates the exocrine glands of the mucosa in the fundus to secrete HCI into the lumen of the stomach. There is some evidence that as acid increases in the stomach the secretion of gastrin is inhibited. Nerves of the ik Il (a) (b) Vessels ligated, — Vessels Ir (b) ligated Fic. 4-2. Diagrams showing the two operative stages by which a pouch from the fundus of the stomach of a dog, Canis, is isolated from the rest of the gut and grafted into the mammary gland, where it acquires a new blood supply (I, IIb and IIIc), and an opening to the exterior at 4. The remaining pyloric part of the stomach is stitched up and later brought to the exterior by an open- ing at 9, 10. (I, Ila and IIIb). The oesophagus at 5, 6 is joined to the duodenum at 13, 14, so that food by-passes the stomach (Ila and IIIa). This arrangement was used to demonstrate the presence and action of gastrin (from Grossman, Robertson and Ivy, 1948). vagus can also stimulate the flow of gastric secretion; but it seems clear from experiments like the above that they are not necessary for the release of either gastrin or of acid. How far they may in- crease the natural secretion of the juices and their enzyme content is not clear. 122 KINETIC HORMONES—II COLD-BLOODED VERTEBRATA. There is as yet no certain evidence of the natural action of gastrin in any class of cold-blooded verte- brates. Frogs have been found to respond to injection of mammalian GASTRIN by an increased gastric secretion; however, the natural physiological stimulus seems to be brought about by the sympathetic nerves. In Elasmobranchs, on the other hand, it has been claimed that both adrenaline and sympathetic stimulation serve only to inhibit the flow of the gastric secretion, while histamine or acetylcholine, but not vagal stimulation, increases it (Prosser, 1950). ‘This rather contradictory state of affairs clearly merits further investigation. ahs Distention stopped cm? _ Distention of balloon In main stomach pouch with 150 cm of air --—@ s Volume, Fic. 4-3. Result of an experiment on a dog, with two isolated stomach pouches, as in Fig. 4-2. ‘Time in hours is plotted as abscissae, and output of acid from the fundic pouch as ordinates; the output is stimulated by gastrin, and is shown by weight of HCl (below) and by the changes in volume of the pouch (above). ‘The acid increases markedly in response to the mechanical stimulus of a balloon inserted into the pyloric pouch and inflated with air at the point indicated. When distention in the pyloric pouch stops, the secretion in the fundic pouch falls off rapidly with the fall in GASTRIN production (from Grossman, Robertson and Ivy, 1948). Aves. Birds are among the few forms, other than mammals, in which positive evidence has been found for injected GASTRIN stimulating the acid gastric secretion in the proventriculus (Keeton et al., 1920). Its physiological action has not been shown. Bicarbonate secretion by the pancreas MAMMALIA. SECRETIN was discovered in 1902 by Bayliss and Starling; but it was many years before its action was fully proved. § 4.111 DIGESTIVE GLANDS 123 It is produced from the duodenal mucosa by the action of the acid chyme entering from the stomach. Pavlov (1910) and others showed that this stage in digestion initiates a copious flow of secretion from the pancreas into the duodenum, and that this still occurs after section of both the vagus and sympathetic nerve 160,- HCl Secretin 140 120 100 Bacto- . 80 Sodium : protone oleate Amino acids 60 40 Percentage change in volume Water Sol. m =e starch Dextrose one a Saline Maltose Canin < level ees se eS ay ok SS eS. 4 ae) Nee Fic. 4-4. Diagram showing the effectiveness of various substances introduced into the lumen of the duodenum in inducing secretion of bicarbonate from a transplanted pancreas. The collected pan- creatic secretion indicates by its change in volume (ordinates) the quantity of SECRETIN released into the circulation from the duo- denum. Under each substance three vertical bars represent three consecutive 20 min collecting periods; each result is the mean of 4-10 tests, carried out on 3-5 dogs in each case. Acid and secretin are equally effective; peptones and amino acids much less so (from Wang and Grossman, 1951). supply; a completely denervated loop of the intestine of a dog, if treated with acid in the lumen, can start the pancreatic flow, so long as the circulation is intact. The stimulus for pancreatic secretion is shown, by this and other cross circulation and trans- plantation experiments, to be due to a blood-borne factor, and 124 KINETIC HORMONES—II not to be dependent upon any nerve reflex between the gut sur- face and either the cells secreting the factor or the pancreas. Direct introduction of acid into the circulation is not effective in causing the pancreatic secretion; whereas introduction of extracts of the duodenal mucosa brings on the flow of secretion within about one minute. | Small quantities of secretin can also be extracted from the gastric mucosa; but the main source is from the duodenum, and con- sequently its secretion is only brought about by placing acid in this part of the gut. Acid in the stomach or the lower part of the intestine has no effect in stimulating pancreatic secretion. The natural acid is hydrochloric from the stomach; but any buffer that keeps the duodenal contents between pH 4 and 5 stimulates the production of secretin (Grossman, 1950). Peptones and amino acids are much less effective (Fig. 4-4). The pancreatic glands which are stimulated by SECRETIN produce an alkaline solution containing NaHCOQ;; as this secretion brings the duodenal contents to neutrality, the stimulation of secretin stops and the flow of alkali from the pancreas also stops, the end point of this hormone-controlled titration having been reached. Secretin also increases the flow of bile from the liver; but enzyme secretion from the pancreas is stimulated by the vagus nerve or by pancreozymin (‘Table 14). COLD-BLOODED VERTEBRATA. Elasmobranchs, salmon, frogs, salamanders and some reptiles, such as tortoises, have all been found to yield extracts, which act like secretin in mammals; but it is not certain that all the extracts contain the same chemical as the mammalian hormone: they may owe their activity to the presence of histamine, which is a potent secretagogue for the pancreatic glands. There is no evidence for the hormone having any natural function in these animals. Aves. Claims made for the presence of secretin in birds appear to be founded on some confusion between this hormone and gastrin. They are said, however, to respond to injection of SECRETIN. Digestive enzyme secretion by the pancreas MAMMALIA. PANCREOZYMIN, like secretin, is secreted from the mammalian duodenal mucosa and acts upon the pancreas (Fig. § 4.111 DIGESTIVE GLANDS 125 4-1); but, as its name implies, it stimulates those gland cells which secrete the digestive enzymes. Vagal stimulation also facilitates the secretion of enzymes; but the action of the hormone persists after bilateral section of both the vagus and the sympathetic nerves. Experiments (Wang and Grossman, 1951) on trans- planting the pancreas, to eliminate any possibility of nerve con- 300° Bacto- 280 Protone is ‘E 260 Amino ~ 240 acids B 220 = Sodium 3 200 : oleate » 180 3 = 160 E 140+ § = “(20 HCL @ 100 1 OE = 80 fe Ci Corn - o 60 oy = oS ® ir 2 40 Bwtr 5 £ £ E 5 © A Oo Oi ea feet oO 20 ee. = Q of 5 2 Be Oa c a (@) : ob ontrol level 5 4 5 Tests 3 U 4 4 3 5 on 4 See 3 3 Dogs Fic. 4-5. Diagram, as in Fig. 4-4, showing the effectiveness of various substances in stimulating enzyme secretion from a trans- planted pancreas, for 3-5 dogs in each case. The change in enzyme secretion indicates the quantity of PANCREOZYMIN released into the circulation from the duodenum, in response to each substance. Peptones and amino acids are more effective than acid in stimu- lating pancreozymin secretion (from Wang and Grossman, 1951) nection between the duodenum and the pancreas, prove the presence and independent action of pancreozymin as a hormone. The stimuli that have been reported to cause the release of pancreozymin include soap, peptone, casein, starch, various sugars and even distilled water, introduced into the duodenum; they have now been rigorously tested on pancreatic transplants. ‘These experiments show that peptones and the amines, leucine, trypto- phane and phenylalanine, resulting from protein digestion, are more effective than acid (Fig. 4-5). 126 KINETIC HORMONES—II Digestive enzyme secretion by the duodenum Mamma.ia. There is some evidence (Grossman, 1950) for DUOCRININ being secreted like secretin, from the mucosa of the duodenum in response to acid, but it acts upon Brunner’s glands in the wall of the duodenum itself and not upon the pancreas. It appears with secretin in crude extracts of the mucosa, but 1s separable from it. Digestive enzyme secretion by the intestine MamMatia. Certain peptones in contact with the intestinal wall stimulate the secretion of ENTEROCRININ, which increases the succus entericus, both in volume and in enzyme content. The gland ells that secrete the succus entericus are in the walls of the jejunum and ileum. The cells that secrete enterocrinin are con- fined to the same region of the intestine, but have not been identi- fied histologically. 4.112 Decrease in secretion by gut glands A number of hormones have been postulated as having an inhibiting effect upon the exocrine glands of the mammalian gut; but such an action is difficult to prove because so many factors in the experimental procedures are liable to reduce either the activity of the gland or the sensitivity of the tissues at one point or another in the chain between the gut surface, the endocrine cells and the exocrine glands. Inhibition of acid secretion by stomach glands Mammatia. The most clearly established of the inhibiting hormones is ENTEROGASTRONE. Its source, like that of some stimu- lating hormones, is in the duodenal mucosa; but its action is to counteract that of gastrin and to inhibit the secretion of acid by the gastric glands, thereby bringing to an end the gastric phase of digestion and facilitating the neutralization of the gut contents as they pass into the duodenum (Fig. 4-1). ‘“‘Among the agents which inhibit gastric secretion when present in the duodenum, fat is the only one which has been proven by experiments with the transplanted pouch to act by release of § 4.112 DIGESTIVE GLANDS 127 enterogastrone. Concentrated sugar solutions, which have been shown to be capable of inhibiting gastric secretion when present in the duodenum”’, probably act in the same way. ‘Curiously, although gastric motor inhibition by acid in the duodenum depends on the integrity of the vagi, the gastric secretory inhibition apparently does not. This phenomenon deserves further study” (Grossman, 1950). To summarize the situation in mammals: when food distends the stomach, GASTRIN causes the secretion of acid in which diges- tion starts. As this acid passes on into the duodenum, aided by gastrin stimulating the stomach muscles to contract, it causes SECRETIN to stimulate the flow of neutralizing alkali from the pancreas, and bile from the liver. Acid in the duodenum also stimulates the secretion of DUOCRININ which causes a flow of enzymes from Brunner’s glands. At the same time, fats entering the duodenum cause ENTEROGASTRONE to inhibit acid secretion and reduce muscular action in the stomach. Incidentally, fats also stimulate the secretion of CHOLECYSTOKININ which controls the muscles of the gall bladder (§ 3.112). When the products of protein digestion reach the duodenum from the stomach, they cause PANCREOZYMIN to stimulate the flow of enzymes from the pancreas to continue the digestion of proteins in the duodenum. When the resulting peptones reach the ileum, they cause ENTERO- CRININ to stimulate there the secretion of further digestive enzymes in the succus entericus. It is therefore apparent that this series of hormones acts independently of any nervous stimulation to produce a co-ordinated series of changes in the gut, and especially in the sequence of secretions poured into its lumen, so that the phases of digestion follow one another in the right order and at the right time. These hormones have several peculiar features in common: they are not stimulated by nerves but by direct mechanical or chemical stimuli; they are kinetic and appear in this case to be nerve-like, in that their actions are the same as those of the parasympathetic nervous system in lower vertebrates; they are all secreted by isolated cells in the gut mucosa, since no discrete endocrine glands are to be found, but their histological origin is unknown (§ 2.132). If the cells are endodermal, they are unlike those secreting any other kinetic hormones. 128 KINETIC HORMONES—II 4.12 OVIDUCAL GLANDS GasTRopopaA. Increased formation of secretory material in the albumen gland in the reproductive ducts of the slugs, Arion and Limax, has been shown to depend on a hormone from the GONAD (Laviolette, 1956). ‘The growth of the glands is also stimulated and it seems probable that the hormone is having a morphogenetic effect rather than the strictly kinetic action of releasing the secretion into the ducts. AMPHIBIA. The oviducts of Anura and Urodela secrete jelly, which is laid down round the ovum and swells when it is laid in water. At first it serves a protective function, but may be eaten later by the newly-hatched young. In Bufo arenarum it has been shown (Smith, 1955) that injection of a variety of hormones can induce the secretion of this jelly from the oviducal glands. The most consistent results seem to be obtained with PROGESTERONE- like substances from the simple corpora lutea that form during ovulation in the ovary (Galli-Mainini, 1951), under the morpho- genetic influence of the luteinizing hormone, LH, from the adenohypophysis (§ 2.123; ‘Table 15). This secretion of jelly has also been induced by PROLACTIN, when supplied from implanted toad hypophyses; but this hormone has not been confirmed as physiologically active in nature. It is probably the same as luteotrophin, LSH, the action of which in mammals would be to stimulate the secretion of progesterone. If it is the same in Bufo, this would be one of the rare cases in which the secretion of a hormone with a kinetic action is subject to endocrinokinetic stimulation (§ 4.2); but the main function of progesterone is morphogenetic and this kinetic action seems to be subsidiary. It is claimed however that prolactin can act in the absence of the ovary (Houssay in Nalbandov, 1959). Aves. The single oviduct of young birds secretes albumen most freely in response to stimulation by PROGESTERONE in the presence of oestrogen; in mature birds the process is facilitated by the presence of the egg yolk, though this may be replaced by any smooth foreign object such as a glass bead of suitable size (Nal- bandov, 1959). Mamma tia. The uterus is a derivative of the oviduct, in which OVIDUCAL GLANDS 129 § 4.12 vOISU] é suoskpoy epodomseyy ¢ peuoryy snnby erqrydury S[PUTLUPR TAT sparg s[PUIUUR JAI spar s[BUIUUIR A] a9 ofng | ce 2 ouITeUuDIpY ud3011SIOC) Uud3801}S9Q HST ‘unovpordg HS ‘unovjoidg 9UO0II}SOBOIg HST “Unorfordg dUOII}SIsSOIg B8uT}9199S 99ND JROMG snonj[ SGNV1I9 NIMS +1['$ ALBURY do19g UO1JadAIAS UL aSVAAIBd TELL + AIvUTUIR I doi9g UOMAAIAS UL ASVAAIUT [E+ SGNV1ID ONILAMOES-ATIAL «CLP oUuTIgI-) uswINng]y SGNV1ID TVONGIAQ TIP Be a asgS0—0—_._—o—a ATH NVXo ANOWYOH YO NVYOUO ALVUPALYAANI ATH NVXA ANOWYOH ALVUEALYAA SuO LOasAa SAGNVID ANIMOOXA YAHLO ONITIOULNOOD SHUNOWUOH OILANTS “CT ATAY LT 130 KINETIC HORMONES—II glands hypertrophy and secrete at the time of implantation of the embryo and during early stages of its development. Not only the growth of the glands but also their secretion seem to be stimulated by PROGESTERONE from the corpora lutea in the ovary. This similarity to the hormones that are claimed to stimulate the secretion of oviducal glands in the toad, Bufo, may lend weight to the claim of progesterone to be concerned in the latter. It is clear, however, that there are distinct differences between the hormone pattern in the lower vertebrates and the mammals, and this line of argument must be accepted with caution. 4.13 MILK-SECRETING GLANDS Aves. In some birds, such as the pigeon, Columba, a nutritive fluid, the so-called “‘pigeon’s milk’, is produced from the thickened lining of the crop in both sexes and is regurgitated to feed the unfledged young. ‘This secretion seems definitely to be under the control of PROLACTIN from the adenohypophysis. The hormone is effective even in castrated, hypophysectomized and adrenal- ectomized birds (Turner, 1955); its action must therefore be direct. MAMMALIA. PROLACTIN is also active in most mammals, induc- ing the secretion of milk in the mammary glands; but it is only able to act upon glands that have already been stimulated to grow and to reach a certain level of size and development of the acini by the combined action of oestrone and progesterone. It is not clear whether the action of the prolactin, LSH, is direct, as in birds, or whether the effect is due to its also stimulating the secretion of progesterone. The former seems the more probable, but, if so, it is the only example so far noted of a hormone that is able to stimulate the secretion of both exocrine and endocrine glands. The action of prolactin is linked with that of other hormones in that the muscular release of the milk from the glands into the ducts, and so to the teats, is under the control of OXYTOCIN (§ 3.114). Other hormones must also have an effect upon milk production through their effects upon metabolism, and in particu- lar on the level of sugars, fluid and calcium in the blood (§§ 5.2, 5.3 and 5.4). Prolactin can apparently be inhibited by high levels of OESTRO- § 4.2 ENDOCRINE GLANDS 131 GEN in the blood, such as are present towards the end of lactation or at the renewal of the oestrus cycle (Cowie and Folley, 1955). It is possible that the nervous system plays a part in the control of prolactin secretion, but there is less evidence for a psychological control over this secretion than there is for oxytocin (Fig. 3-7) . 4.14 SKIN GLANDS AmpuiBiA. The skin of amphibians is kept moist in air by the secretion of mucus from epidermal glands, which respond to the presence of ADRENALINE, at least in pharmacological doses of a mammalian preparation. The glands have a sympathetic nerve supply, and it has not been shown that the hormone plays any part in their physiological control. Under experimental conditions it is possible that adrenaline contracts muscles round the glands rather than stimulating secretion (Wastl, 1922). The same is probably true of neurohypophysial extracts which cause an outflow of secretion from the skin of Xenopus (Bastian and Zarrow, 1954). _ Mammatta. In the cat, Felis, as in man, the sweat glands in the skin respond to the peculiar cholinergic stimulation supplied by their sympathetic nerves; they therefore do not respond to adrenaline in the blood as do other effectors, supplied by the more usual adrenergic sympathetic nerves. In the horse, Equus, and sheep, Ovis, the sweat glands respond to ADRENALINE by secretion, as the mucus glands do in amphibians (fide Winton and Bayliss, 1955). 4.2 ENDOCRINE GLANDS The endocrine glands, like their exocrine counterparts, are effector organs, although the fact that their secretion passes into the blood, instead of appearing in a duct, makes their action less apparent. The hormones, which in many cases stimulate their secretion, therefore fall within the kinetic group. Since, however, a hormone which causes the secretion of another hormone has some peculiarities, it is convenient to distinguish it as an endo- crinokinetic hormone, or one that is able to activate an endocrine gland.* These hormones have sometimes been referred to as * “Glandotrope Wirkung” has been used to express this activity in German (KarLson, 1956); but there is no accepted English term. 132 KINETIC HORMONES—II either trophic or tropic especially in such compound names as gonadotrophic; but reasons have already been given (§ 1.51) for avoiding the confusion to which the use of these terms leads. It is not all endocrine glands, the secretion of which can be stimulated by hormones; but the exceptions are clear cut, at least in vertebrates. Glands formed from modified nerve cells are never so controlled, nor are the parathyroids and the insulin-secreting cells of the islets of Langerhans in the pancreas (§ 5.52); such control is very rare for any gland secreting kinetic hormones. It is almost always the glands secreting hormones with metabolic or morphogenetic actions, or both, that are controlled by endocrino- kinetic hormones. Among invertebrates, the only cases so far known are glands derived from the ectoderm; but it seems more than likely that some will be found in the gonads. The vas deferens gland of Crustacea, with its morphogenetic hormone, could well be controlled by such a hormone; but neither this nor any other form of control has yet been found for it. Among vertebrates, all the endocrine glands in question are derived either from the endoderm or the mesoderm, and all are controlled by endocrino- kinetic hormones from the adenohypophysis. The action of an endocrinokinetic hormone is much more difficult to establish than that of a hormone which acts directly upon some kinetic or metabolic process, especially as it is so often found that the endocrinokinetic hormone stimulates a whole complex of metabolic processes, interference with any one of which may upset the balance of the rest, and interfere with the health and reactions of the test animal. An indication of the minimum number of experiments that are theoretically necessary to establish the interrelations of two hormones, one endocrino- kinetic and one metabolic, has already been given (§ 1.6). It is rare to find animals in which the nervous system does not introduce further complexities. Often the different stages in the experimental proof have been supplied by different authors, and can only be interpreted by reference to the histological and chemical results of yet other workers. Interpretation of work upon vertebrates has been particularly beset by difficulties introduced by testing the hormones from one class of animals upon those of another. In the cases given below, the main action of each endocrino- ENDOCRINE GLANDS 133 § 4.2 vaoyqyjvy é ‘OU UW UlBIG vaoyqojvA FT uin1Iga199.19} UT e193} uol[sues -dorswoydy jeaseydossoqns é uvsI0 a10d DIDUSA'T AtOSUdS S WIOISsUR;T ATd NVXA ANOIWYOH YO NVONO ALVUdALYAANI SISSB[D JSOTA[ S[BUTUIR TAT é Biqryduy s[PUTWe A SOSSBID SOJT SN]JOM ‘oo ‘STUDD sny]P) STBUILUR AT eiqrydury sty DIdNVXY HSO!I HST (¢ HSA +) HSOI HLOV HLOV H.LS HANOWYOH ALVYEALYAA 9U019}SO}SO J, 9U0I9}SaB0Ig ua8011s9Q spouoyy 9UOST}IOIOIPATT dUuO0I9}SOp[y XaqAOI [DUAAP PY SANV1D ‘IvVWuadosay uoseon{y suoysasuvTT fo sajsy piosdy J, SGNVTID TIVINGACOGN uinjo]]0 snq4o;s) ‘IJa ‘spud]s I19DLOYIOA Spun]s ]d44uUa A UDSAO- X SANV19 TIVINYACOLOY SHO LOFAAa ClO + Lec v tc ¥ EO Ice + CCV vIC + Cle v CIC Fr LIC + Ic tb SANVT19 ANIYOOGNYA ONILVTIOIWNILS SHNOIWHOH OILLANIMONINOOCNY ‘QT ATE I, 134 KINETIC HORMONES—II kinetic hormone is stimulating the secretion of an endocrine gland. It will be noticed that, although this is often accompanied by growth of the gland, recent evidence shows that the two actions may be due not to the same hormone but to two separate, though closely similar, hormones. Reduction or inhibition of the secretion of endocrine glands is usually due to an indirect “feed-back” system, whereby the accumulation of the metabolic or morpho- genetic hormone in the blood depresses the output of the endocrinokinetic hormone, which had induced that accumulation (e.g. ACH and ACTH, § 4.231). _ Since these endocrinokinetic hormones stimulate the secretion of a second hormone which nearly always has metabolic or morphogenetic effects, they will be more fully understood after a consideration of the hormones that they control. The reader with little previous knowledge of hormones is therefore recommended to read Chapter 5 on metabolic hormones, before considering their control by endocrinokinetic hormones in the present chapter. Reference could well be made to each case as it arises, to which end the section numbers are given as a guide. An account of the morphogenetic hormones, and more details of the endocrinokinetic hormones which control them, will be given in Part II. 4.21 ECTODERMAL ENDOCRINE GLANDS OF ARTHROPODA Ectodermal glands that are stimulated to secrete by endocrino- kinetic hormones are few; the only glands that have so far been postulated all arise in the heads of Arthropoda (Part I1). The glands derived from ectodermal invaginations in the anten- nary or 2nd maxillary segments all secrete hormones which have a moult-promoting action. The fact that their secretion can be stimulated by endocrinokinetic hormones has been fully estab- lished in a number of insects, but is still uncertain in crustaceans. 4.211 Y-organ possibly stimulated by a secretion from Hanstrém’s sensory pore organ Crustacea. The Y-organ has been identified in many Crustacea and is derived from either the antennary or the 2nd maxillary segment, in whichever position is unoccupied by the excretory organ. Its hormone has both metabolic and morphogenetic actions, § 4.213 ECTODERMAL ENDOCRINE GLANDS OF ARTHROPODA 135 though the former, which concern protein metabolism (§ 5.2) and calcium distribution (§ 5.4), are intimately related to its main action as a moult-promoting hormone (Part II, § 3). Without its secretion, neither moulting nor regeneration can occur (Echalier 1956). It is inhibited, or its action is overridden, by the moult- inhibiting hormone from the sinus gland; but it can secrete when its nerve supply has been severed. This last observation suggests that it may be subject to some hormonal stimulation, such as might be exerted by Hanstrém’s sensory pore organ, which secretes a MOULT-ACCELERATING HORMONE in Lysmata and Leander; but there is only presumptive evidence for naming the latter as a true endocrinokinetic hormone, capable of stimulating the secretion of the Y-organ (Knowles and Carlisle, 1956). 4.212 Ventral glands stimulated by a secretion from the suboeso- phageal ganglion Insecta. ‘The maxillary ectodermal glands which secrete the moult-promoting hormone, ecdysone, in the more primitive orders, Ephemeroptera and Odonata, retain their original ventral position in the head (Fig. 2-8). Unlike their homologues in other orders of insects, they are stimulated to secrete by neurosecretory cells in the suboesophageal ganglion, instead of in the brain. The axons of these cells connect directly with the ventral glands (Gabe, 1953) and probably release a neurohormone (§ 1.2), since they are only effective if the axons remain intact. A rich source for extracts of the neurohormone has been found in the ganglion (Arvy and Gabe, 1954), but a vascular endocrinokinetic hormone has not been shown. 4.213 Peritracheal and prothoracic glands stimulated by a secretion from the intercerebrum Insecta. Despite the differences in their final positions (Fig. 2-8), these glands appear to be homologous with each other and to be derived from ectodermal intuckings in the 2nd maxillary (or labial) segment, like the ventral glands and some of the crustacean Y-organs. The peritracheal glands form part of Weismann’s ring in Diptera, such as Calliphora. Prothoracic glands occur in most other genera, of which Rhodnius, Hyalophora 136 KINETIC HORMONES—II and Bombyx are the only ones that need to be referred to here. These glands have all been found to secrete the moult-promoting hormone, ECDYSONE, in response to an endocrinokinetic hormone, PROTHORACOTROPHIN, from the neurosecretory cells in the inter- cerebrum of the brain. | It is clearly established that no nervous connection is needed between the brain and prothoracic glands for stimulation to be effective, except possibly in the Diptera. A conclusive proof for the action of an endocrinokinetic hormone in the Lepidoptera can be given in connection with the termination of diapause (§ 5.112), and is therefore more appropriate here than an example derived from the control of moulting (Part II, § 3); but the interaction of the hormones seems to be the same in either case. In nature, diapause of the Cecropia silkworm, Hyalophora, lasts for 6 to 7 winter months before the tissues again become active and differentiation leads to the emergence of the adult in the spring (§ 5.12). Diapause can be shortened or “‘broken”’ artificially in various ways, of which the simplest is the injection of an active extract of ECDYSONE from the prothoracic glands. These glands do not secrete during diapause, but they can be induced to do so at any time by the presence of an actively secreting brain. Reactiva- tion of a diapausing brain can best be brought about by chilling it suitably (Williams, 1952). If a diapausing pupa is joined by a plastic tube to a diapausing pupal abdomen, so that their influence on each other can only be by hormones in circulating haemolymph, then a chilled brain, implanted in the hinder abdomen (Fig. 4-6), breaks diapause in the anterior specimen first, and only later in the hinder abdomen, where the implant is. It may be concluded that the brain has no direct effect in the hinder abdomen, but that its endocrinokinetic hormone, PROTHORACOTROPHIN, stimulates the prothoracic glands in the anterior specimen to secrete ECDYSONE, which may override the diapause hormone, D (see § 5.112). If the length of the tube separating the two pupae is increased, the time lapse between brain implantation and the end of diapause is also increased. The possibility that the secretion of other metabolic hormones by the prothoracic gland may also be subject to endocrinokinetic stimulation by a hormone from the brain has not, apparently, § 4.213 ECTODERMAL ENDOCRINE GLANDS OF ARTHROPODA 137 Fic. 4-6. Experimental method used to test the action of hormones in dia- pausing pupae of the Cecropia silk- worm, Hyalophora. The pupa in front is joined, by a plastic tube up to 3 cm in length, to an isolated abdomen behind. The tube passes at either end through a disc (Di) which closes the cut surface of the abdomen, being sealed in with paraf- fin wax. Windows (W) can be sealed in at either end for observing the onset of differentiation in the internal tissues. This is stimulated by im- planting a chilled, and therefore activated, brain into the hinder abdomen (Im) ; PROTHORACOTROPHIN from the implant causes secretion of ECDYSONE, the moult-promoting hor- mone, from the prothoracic glands in the front specimen (A), which reacts first. Diapause is broken later in the hinder abdomen when it is reached by the haemolymph bearing ecdysone from the front specimen (based on data from Williams, re- drawn after Hinton, 1951). been specifically investigated. Presumptive evidence in favour of such an effect is that both the metabolic and morphogenetic functions of the glands are carried out simultaneously, so that they could be due to the same prothoracic hormone under the same endocrinokinetic control. 138 KINETIC HORMONES—II 4.214 Corpus allatum of Insecta possibly stimulated by a secretion from the brain The corpora allata are ectodermal invaginations from the 1st maxillary segment (§ 2.122) and their secretion inhibits metamor- phosis in the nymphal or larval and pupal stages; but it stimulates egg-growth as well as increasing oxygen consumption in the adult. The best evidence for any hormonal stimulation of their secretion is derived from experiments on the blow-fly, Calliphora; but the _ results are not conclusive. If blow-flies, 8 hours after their emergence as adults, have their median neurosecretory cells, m.n.c., removed from the brain, it is found that the eggs in the ovary fail to mature, the result being the same as that which follows allatectomy. Reimplantation of mature corpora allata into allatectomized females can restore egg- growth to normal in 93 per cent of cases; but a similar implanta- tion (of corpora allata) into flies deprived of m.n.c. has scarcely any effect. The reimplantation of a double dose of m.n.c. into flies deprived of their own m.n.c., but retaining their corpora allata did not cause so great an increase in egg size as the reimplantation of corpora allata into flies with normal m.n.c. (Thomsen, 1952). If these results are interpreted as showing that the neurosecretion from the brain has a stimulating effect upon secretion by the corpus allatum, then they suggest that the brain cells secrete this endocrinokinetic hormone less freely in isolated implants than when they remain zm sztu in the brain. ‘There is no positive evidence that the brain has any control over the secretion of metabolic hormones from the corpora allata; the effect of removing the m.n.c. from the brain has not been tested in relation to oxygen consumption. On the other hand it is clearly established that inhibition of secretion from the corpora allata can be brought about by the brain, both during metamorphosis (Part II, § 3) and while developing eggs are present in the oviducts of the viviparous cockroach, Leucophaea maderae (Liischer and Englemann, 1955). Growth of the corpora allata, as well as their secretion, is released from this inhibition by sectioning the nerves from the brain. Since this operation not only deprives the corpora of nervous stimulation but § 4.221 ENDODERMAL ENDOCRINE GLANDS OF VERTEBRATA 139 also of a copious neurosecretion, which passes to them within the nerve axons from the brain and the corpora cardiaca, it is not clear whether the inhibition is nervous or secretory (Scharrer, 1952: cf. Fig. 3-2). 4.22 ENDODERMAL ENDOCRINE GLANDS OF VERTEBRATA 4.221 Thyroid gland stimulated by TSH The thyroid-stimulating hormone or THYROTROPHIN, TSH (Table 16), can be extracted from the adenohypophysis of most vertebrates, including teleost fish, but not the elasmobranchs; yet all too often the hormone has been tested on an animal of a differ- ent class from that of the donor (Adams, 1946). In most cases, even when the hormone comes from the same kind of animal, the test of the efficacy of the TSH has been its action in inducing growth of the THYROID GLAND rather than in stimulating its secretion of THYROXINE, which is the aspect under consideration here. TELEOSTEI. ‘The most readily observed effect of thyroid secretion is its power to increase the oxygen consumption of animals in which it occurs. This can be used as an indicator of the action of TSH on the thyroid. In most teleosts, except Pseudoscarus, it is extremely difficult to remove the thyroid glands, because they are so diffuse; but the same effect as thyroidectomy can be achieved by the injection of methylthiouracil, MTU, which destroys the thyroxine. By using this on goldfish, Cyprinus, before measuring their oxygen con- sumption for several hours under constant conditions, it could be shown that, whereas injections of TSH into normal adult fish in the summer caused an increase of up to 200 per cent in oxygen consumption, similar injections of TSH, following the use of MTU, were without effect (Miiller, 1953). This shows that, at least in active adult fish, THYROID GLANDS are responsive to the endocrinokinetic action of TSH. It has also been shown that more TSH is required at lower temperatures to maintain the same level of thyroxine secretion (cf. Pickford and Atz, 1957). Removal of the hypophysis should not affect these results. Amputsta. One of the early discoveries about endocrine glands 140 KINETIC HORMONES—II was that the THYROID GLAND of frog tadpoles, Rana, regressed and might atrophy, if the hypophysis was removed. ‘The tadpoles then failed to metamorphose; but could be induced to do so by injections of a hypophysial extract containing THYROTROPHIN, TSH. This stimulated renewed growth of the thyroid gland and the formation and release of THYROXINE, which in turn induced metamorphosis. Mamma tia. It seems clear that here (and in birds, Bates and Cornfield, 1957) the discharge of THYROXINE from the thyroid into the blood circulation occurs in response to stimulation of the gland by TSH. The action is, however, not altogether simple, since evidence is accumulating for there being either two separable actions of TSH, or possibly two hormones; one stimulating the growth of the THYROID GLAND, and the other its iodine metabolism and secretion. The cycle of iodine metabolism has been worked out in some detail, partly by means of studies on radioactive iodine (I**), which can be followed through the tissues zm vivo (Taurog et al., 1958). Iodides, derived from food and from the breakdown of disused thyroxine, circulate in the blood, whence they are trapped by the thyroid gland and oxidized to iodine. (This reaction is specific and much more efficient than that by which other halogens and related elements are picked up by the thyroid.) In the gland, iodine is attached to an organic molecule to form thyroglobulin, which is a precursor of the hormone. It is stored as a characteristic colloid in the intrafollicular lumen (Fig. 4-7 a-c). It can then be hydrolysed by a proteolytic enzyme to release thyroxine into the circulation (Fig. 4-7 d-e). This filters through the capillary walls to reach the tissue spaces, and bathes the cells before being drained into the lymphatics. The thyroxine accumulates in the peripheral tissues and, notably, in skeletal muscle, where it seems to be concerned with the activity of tissue enzymes. When the tissue hormone is broken down through use, it yields up iodides to the circulation, and the cycle starts afresh (Salter, 1949). TSH acts upon this cycle in the thyroid gland itself. Its first effect is to activate the enzyme converting the stored thyroglobulin to thyroxine, and its second is to stimulate the release of thyroxine into the circulation. This reaction is rather slow in rats, and may only be detectable histologically after 22 hr (Fig. 4-8). At the same Fic. 4-7. Camera lucida drawings of sections of thyroid glands from the guinea pig, Cavia, rapidly frozen-dried at intervals after inject- ing standard doses of thyroid stimulating hormone, TSH ( 2150). (a and 5) after half an hour and (c) after 6 hr from the time of inject- ion, to show gradual increase in cell-size and accumulation of droplets near the inner surface of the cells, through which some are passing into the intrafollicular store of colloid above; (d and e) a later stage with increase in cell height and a reversal of the direction of cell activity. Colloid is being reabsorbed from the store as clear drop- lets, and secreted towards the surface of the cell next to the blood capillaries below (from de Robertis, 1949). Fic. 4-8. Thyroid gland of Rattus, 22 hr after an injection of TSH (later stage than Fig. 4-7 d). The droplets of colloid (sc) from the intrafollicular store (above) have all moved nearer to the secreting surface, which is extended towards the lumen of the blood capillary (£). (From de Robertis, 1949). § 4.221 ENDODERMAL ENDOCRINE GLANDS OF VERTEBRATA 141 time, trapping of iodides and the formation of the intrafollicular colloid is stimulated; although the amount trapped depends upon the level already in store (Barker, 1955). In the absence of TSH, both trapping of iodine and secretion of THYROXINE may be reduced to 10 per cent of normal in 5 days; the binding of iodine to the colloidal proteins may fail or proceed only slowly, and in some cases diiodotyrosine (or even monoiodotyrosine) is formed instead of thyroxine. It has been shown that TSH is inactivated in vitro by exposure to thyroid, but not to other tissues; and that it can be re-activated by treatment with reducing agents such as thiouracil. This may be related to the fact that the latter destroys the thyroxine in the thyroid. The increase in height of the follicular epithelium in the thyroid gland, which is often used as a means of assay for TSH activity, accompanies the normal release of thyroxine from the gland (Fig. 4-7 d—e); but increase in weight of the gland is not necessarily related to its rate of secretion, though it may also follow prolonged treatment with TSH (or withthe growth-promoting fraction thereof). The rate of secretion of TSH, rather than the quantity of it that is stored in the hypophysis, seems to be increased by some form of control from the hypothalamus, since it can be disturbed and reduced by hypothalamic lesions in the brain (Bogdanove, 1957). On the other hand, its secretion can be decreased by high con- centrations of thyroxine, circulating in the blood; this feed-back reaction tends to maintain a steady level of production of the metabolic hormone. Evidence for there being two distinct, though closely similar, substances both included under the name of thyrotrophin is provided by experiments on mice, in which the growth-promoting action of TSH responds differently from its secretion-promoting action. Hypophysectomy results in reduction in size of the thyroid gland, as well as in its secretion. If the hypophysis is transplanted elsewhere in the body, the mice do not show any increase in thyroid weight, when compared with hypophysectomized controls ; but such reimplantation does restore the iodine content and thy- roxine formation and secretion by the thyroid glands to 66 per cent of normal, as compared with only 10 per cent in hypophysecto- mized controls without implants (Greer, Scow and Grobstein, 142 KINETIC HORMONES—II 1953). The growth-promoting action of TSH is only restored to any appreciable extent if the reimplantation of the hypophysis brings it into contact with the median eminence of the brain, in the position from which it was originally removed. This may be com- pared with the similar effect of CRF upon the growth-promoting actions of other hypophysial hormones (§ 4.323). 4.222 Parathyroid glands Earlier work suggested that activity of the parathyroid glands _ could be stimulated by thyroxine from the thyroids. It is now established that this is not an endocrinokinetic action, but an indirect effect, in that thyroxine tends to lower the calctum and increase the phosphate content of the blood and these changes both stimulate the parathyroid glands directly (§ 5.521). 4.223 Glucagon-secreting cells of the pancreas stimulated by STH Aves. 'The growth hormone or SOMATOTROPHIN, STH, has only been shown to induce an increase in blood-sugar in the chick, Gallus; but it seems probable that this is an indirect action brought about through its endocrinokinetic stimulation of GLUCAGON secretion, as in some mammals (§ 5.211). Mammattia. The stimulation of GLUCAGON secretion from a cells in the islets of Langerhans (§ 2.222) is not yet fully under- stood, nor is glucagon widely distributed in mammals (§ 5.2). Though some may be active in all mammals, it is only in some carnivores (cat and dog, but not ferret) that it has been found to be the main factor in raising the level of blood-sugars, in response to stimulation by hypoglycaemia (Saka, 1952) or by sSoMATO- TROPHIN or growth hormone, STH™%, from the adenohypophysis (Young, 1945). The evidence for the endocrinokinetic action of 5'TH has been somewhat obscured by the fact that its administra- tion to hypophysectomized dogs and cats gives different results at different ages. In young animals the resulting release of sugar can all be consumed in growth, and the stimulation of the latter is the only observable result; but in older animals, when growth has ceased, the stimulation of glucagon secretion and the conse- quent release of sugar into the blood is not masked, and hyper- * Glucagonotrophin would be more descriptive in this context. § 4.231 MESODERMAL ENDOCRINE GLANDS OF VERTEBRATA 143 glycaemia, or diabetes mellitus, results and may even become permanent (cf. § 5.211). 4.23 MESODERMAL ENDOCRINE GLANDS OF VERTEBRATA 4.231 Adrenal cortex stimulated by ACTH In most vertebrates there is some evidence that the secretion of CORTICAL HORMONES is stimulated by ADRENOCORTICOTROPHIN, ACTH, from the adenohypophysis. Since the actions of the corti- cal hormones are mainly metabolic, there is no doubt here, as there can be with the thyroid, that the endocrinokinetic hormone is in fact stimulating the secretion of metabolic hormones, and not only of morphogenetic hormones. The stimulation of the adrenal cortex by ACTH presents an interesting problem. Although the actions of cortical hormones connected with electrolyte and water balance all lead to concentra- tion of salts in the blood, it is clear that two distinct types of hormone are involved: the ALDOSTERONE-LIKE HORMONES which increase salt reabsorption (§ 5.311), and the HYDROCORTISONE-LIKE HORMONES which mainly cause water diuresis (§ 5.321). Neverthe- less, the tentative suggestion that there may be more than one hormone in ACTH only indicates a separate control for increasing the weight of the adrenal cortex from that inducing its secretion. There appears to be no indication of separate hormones for stimulating the secretion of the different hormones from the cortex (Munson and Briggs, 1955). Most authors, however, still support the view that adrenocorticotrophin, ACTH, is a single substance (Astwood, 1955). Secretion of aldosterone-like hormones from the adrenal cortex Aldosterone is chemically quite distinct from the other cortical hormones in having an 18-aldehyde group in the molecule. It is biologically very active in stimulating salt retention or reabsorp- tion by the kidneys, although it is secreted in relatively minute quantities. Little is known of the means by which its secretion is controlled; but there is some evidence that it is secreted in response to stimulation by ACTH in the rat, though not in man (Simpson and Tait, 1955). Elsewhere it appears to be directly stimulated by a lowering of salt concentration in the blood. 144 KINETIC HORMONES—II Secretion of hydrocortisone-like hormones from the adrenal cortex COLD-BLOODED VERTEBRATA. There is apparently not much direct evidence of ACTH stimulating secretion by the ADRENAL CORTEX in vertebrates other than mammals, although growth of the adrenal tissue, or of the anterior interrenal tissue which is its main homologue in fish, has been established in relation, not only to mammalian ACTH, but also to extracts of a similar fraction from the hypophysis of fish (Pickford and Atz, 1957). There is - some evidence (§ 5.411) that in Astyanax the secretion of cortical hormones can continue in the absence of ACTH (Rasquin and Rosenbloom, 1954); but the rate of secretion does not seem to have been measured. ACTH derived from the pituitary of the salmon, Salmo salar, causes reduction of ascorbic acid in the rat, but has not been tested on fish (Rinfret and Hane, 1955). MammMa tia. The HYDROCORTISONE-LIKE HORMONES, ACH (some- times referred to as the glucocorticoid hormones, from their diabetogenic action on carbohydrate metabolism), are the most abundant secretions from the adrenal cortex. In normal circum- stances the release of ACH from the cortex into the blood is continuous, but it has a slight diurnal rhythm of change in rate. In the human, for instance, this is lowest at night and highest in early morning. In response to any stress, or tissue damage, there is an immediate increase in the rate of secretion. Since small injections of ADRENOCORTICOTROPHIN, ACTH, from the adenohypophysis, can influence the rate of secretion, it is thought that there is also a basic rate of secretion of this endocrinokinetic hormone, but that it can very quickly be increased in response to damage or stress, and that this induces the cortical reaction (Munson and Briggs, 1955): Secretion of ACH from the cortex is accompanied by a marked loss of ascorbic acid (as well as of cholesterol and a sudanophilic substance, Fig. 4-9), which is not replaced until more hormone is synthesized. The amount of the acid present at any time can be assayed in the frozen glands of rapidly killed specimens. Its disappearance under stress provides a measure of the hormone secretion, which follows it closely (Slusher and Roberts, 1957). It can, therefore, be used as an indication of the rate of ACTH § 4.231 MESODERMAL ENDOCRINE GLANDS OF VERTEBRATA 145 secretion. The reaction varies with the exact type, duration and severity of the stress. If, for instance, anaesthetized rats are given some relatively mild noxious stimulus, such as haemorrhage or an injection of histamine, or if normal rats have a bout of severe Sudano- philic substance 50 We ee 7 Adrenal cholesterol e e Ascorbic acid —_— O I2 24 Fic. 4-9. Diagram to show the type of change that occurs in the adrenal cortex, following brief secretion of ACTH from the adeno- hypophysis, induced by non-fatal haemorrhage for 1 hr. The sectors of gland, above, indicate the change in its size and in the amount of its sudanophilic content. The ascorbic acid (O) and cholesterol (@) in the gland are graphed in percentages over a 24 hr period. These and the sudanophilic substance show a sudden drop in response to the ACTH stimulus at hour 0 until the end of stress at hour 1; this corresponds with the release of the whole store of cortical hormone. During the rest of the 24 hr the store is gradually built up again. Similar effects can be produced by a sudden bout of exercise, or an injection of histamine or of ACTH itself (from Sayers and Sayers, 1949). exercise, ACTH is released and causes a rapid increase in the amount of circulating adrenocortical hormone, ACH, revealed by a reduction in adrenal ascorbic acid (Fig. 4-9). In these cases recovery occurs within 24 hr. In more severe stress, depletion of ACH may be so complete that recovery may not be possible, and death results. The release of ACTH is also related to the amount of ACH already in circulation; when the latter is high it inhibits further JL, 146 KINETIC HORMONES—II ACTH secretion by the usual type of “feed-back” reaction, tending to maintain a steady level. If, therefore, an injection of ACH precedes a histamine injection, it is able to block the secretion of ACTH. There is then no reduction in ascorbic acid, as compared with the control (Fig. 4-10). This result can be achieved, even though the ACH injection precedes the histamine by as little as 500 400 Mg Ascorbic acid /10Og Adrenal 300 200 — Control A.G.E.25ec . A.C.E. Histamine before S3sec 5sec lOsec alone Histamine after Histamine Fic. 4-10. Ascorbic acid remaining in the adrenal cortex, of six groups of rats, killed shortly after receiving injections. Control rats were given saline injections; the others had all been given similar injections of histamine, sufficient to produce a noxious, but not a fatal, stimulus. The marked lowering of ascorbic acid due to histamine alone, which elicits full secretion of ACTH from the hypophysis, is shown on the right. Additional injection of an adrenal cortical extract, A.C.E., 3 to 10 seconds after the histamine has practically no effect upon this lowering of ascorbic acid. A similar injection of A.C.E., given 2 seconds before the histamine, is able to block ACTH secretion, so that the histamine injection produces practically no more response than the saline given to the controls. As in Fig. 4-9, the loss of ascorbic acid corresponds to release of ACH from the adrenal cortex (from Munson and Briggs, 1955). 2 sec, or just sufficient time for the ACH to reach and affect the source of ACTH in the adenohypophysis. By reversing the order of the intravenous injection of the § 4.231 MESODERMAL ENDOCRINE GLANDS OF VERTEBRATA 147 histamine and the cortical hormone, and varying the time interval between the two injections, it can be shown that in 10 sec the histamine has already exerted almost as great an effect upon the release of ACTH and the reduction of ascorbic acid, as it does in the absence of any cortical injection (histamine alone, Fig. 4-10). After only 3 sec there is practically no blocking effect by the cortical hormone, so that it can be concluded that the histamine has already produced some effect upon the hypophysis. The action is in fact almost as rapid as that of the cortical hormone itself, and takes little longer time than the blood needs to circulate from the point of injection to the hypophysis. This does not, however, solve the question of whether these substances act upon the hypothalamus of the brain, rather than upon the hypophysis direct. It is interesting to note that the time required for ADRENALINE to produce a similar reaction in the hypophysis is considerably longer than that for histamine (Fig. 4-11). Adrenaline cannot therefore be held to act as a necessary intermediary between the noxious stimulus and the adenohypophysis from which it calls forth the ACTH secretion (Munson and Briggs, 1955). The rate of secretion of both ACTH and of the cortical hormone can also be measured directly in the blood. ACTH can be detected in circulation soon after the application of any noxious stimulus; ACH can be detected in the adrenal vein within a few seconds of the injection of ACTH. But this method of assay is not wholly satisfactory, since both hormones are destroyed in the tissues within a few minutes of their release. It has been estimated that ACTH is reduced to about half its previous concentration in 2 to 5 min in the blood; but its fate is somewhat uncertain. It is not excreted in the urine, but accumulates in even larger quantities in the kidneys than in the adrenal cortex. It is inactivated after incubation with kidney or liver tissue in vitro; yet the removal of both kidneys and most of the liver does not render an ACTH injection any more effective than before, in reducing the ascorbic acid contents of the adrenal cortex, possibly because even small doses can produce a maximal effect (Astwood, 1955). In the dog, the action of ACTH, in increasing ACH secretion, persists for about 15 min after injection; and even an hour after removal of 148 KINETIC HORMONES—II the hypophysis and the source of ACTH, the naturally induced secretion of ACH is only reduced to one half. 4.232 Gonads stimulated by gonadotrophic hormones Morphogenetic hormones, secreted by the gonads, have a variety of effects related to reproduction, and have been reported from 500 Saline 400 Epinephrine 300 Histamine Mg Ascorbic acid/ 100g Adrenal 200 O I5 30 45 60 Minutes Fic. 4-11. Effects of injections of saline, epinephrine (adrenaline) and histamine on the ascorbic acid content of the adrenal cortex of rats, in mg/100 g gland (ordinates). The saline causes no change in 60 min (abscissae); the reduction due to adrenaline and histamine is by then the same, showing the two doses to be equivalent. The effect of histamine is much the more rapid, and must therefore evoke secretion of ACTH directly and not through any inter- mediary action of adrenaline (cf. Fig. 4-10, based on similar experiments). (From Munson and Briggs, 1955). many animals, including an unusually wide range of invertebrates as well as vertebrates. So far, it is only in the vertebrates that the secretion of these gonadial hormones is known to be controlled by endocrinokinetic hormones. Like the other hormones of this type. in vertebrates these are all secreted from the adenohypophysis ; they are often referred to as gonadotrophins, in common with § 4.232 MESODERMAL ENDOCRINE GLANDS OF VERTEBRATA 149 other hormones from the same source which have marked morphogenetic effects upon the growth of their target organs. Only those causing secretion of endocrine glands need be treated in the present section; although all the gonadotrophic hormones play an important role in relating breeding cycles to seasonal stimuli from the environment (§ 4.232 and Part II, § 4). The gonadial hormones themselves are mainly morphogenetic in action (§ 1.53); but they are peculiar in having some subsidiary kinetic effects (§§ 3.12, 4.12 and 4.324). In their latter capacity they pro- vide the only examples of kinetic hormones which are controlled by endocrinokinetic hormones. Information regarding the gonadotrophic hormones of the cold- blooded vertebrates is scanty, compared with that concerning birds and mammals; but in general they appear to have similar effects upon the secretion of the gonadial hormones (Pickford and Atz, 1957). Further reference will be made to them in relation to the latter (Part II, § 4). Interstitial cells in the testis stimulated by ICSH In birds and mammals, and probably in most other vertebrates, the release of testosterone, or other androgenic hormones, from the testis is brought about by an endocrinokinetic INTERSTITIAL- CELL-STIMULATING HORMONE, ICSH, secreted from the adeno- hypophysis. The control of its secretion, like that of TSH, comes from the hypothalamus of the brain and can reflect the effects of environmental changes, transmitted to the brain either directly or by the eyes. The secretion of ICSH, by its release of testosterone, fires off the whole chain of events associated with the breeding cycle in the male. In birds these can include migration, courtship and nest-building and the appearance of secondary sexual characters of plumage and wattles, as well as the essential develop- ment of the genital ducts. In mammals corresponding changes in behaviour and structure are brought about by similar hormones; in those with a limited breeding season there is a slow feed-back mechanism, whereby the accumulation of testosterone inhibits further ICSH secretion after a few months. In those species with continuous breeding this inhibition is not effective, at least until extreme old age. 150 KINETIC HORMONES—II Follicle cells in the ovary stimulated by ICSH The release of oestrone, or other oestrogenic hormones, from the vertebrate ovary appears also to be stimulated primarily by the INTERSTITIAL-CELL-STIMULATING HORMONE, ICSH. Its action may depend in some cases on the synergic effect of the FOLLICLE-STIMULATING HORMONE, FSH; but the most important action of the latter is to cause the growth of the follicle cells, along with that of the ovum that they enclose. There are differences in the relative importance and effects of these hormones between different species and even more between different classes of vertebrates. The release of oestrogens by ICSH results in the development of the female genital ducts, but not asa rule in that of any secondary sexual characters; the external appearance of the female is usually distinguishable from the male by the absence of male characters, rather than by any positive features due to oestrogenic hormones. In the female mammal, whether the breeding season is short or continuous, there is always an alternation, or cycle, of phases within the breeding period and of these it is the oestrus phase which results from the secretion of oestrogens (§ 4.323). ‘The feed-back reaction, whereby the accumulation of oestrogens decreases ICSH secretion, is relatively rapid in the female and makes way for the alternate phase of the cycle, whether this is dioestrus, pregnancy or pseudopregnancy. In most species oestrus only lasts for a few days each time. Corpora lutea in the ovary stimulated by LSH MammMatiA. The endocrinokinetic hormone LUTEOTROPHIN, LSH, from the adenohypophysis, stimulates the secretion of progesterone from the corpora lutea that form in the ovarian follicles after the shedding of the ova. LSH is probably the same as PROLACTIN, which causes the secretion of milk from crop and mammary glands (§ 4.13); if it is the same it is the only one of the endocrinokinetic hormones which has the power to stimulate any exocrine glands as well as an endocrine gland (§ 4.323). It can only affect the corpora lutea after their growth has been stimulated by the morphogenetic LUTEINIZING HORMONE, LH, also from the § 4.232 MESODERMAL ENDOCRINE GLANDS OF VERTEBRATA 151 adenohypophysis. Incidentally, LH was for many years confused with ICSH; but there is now good reason to believe that they are distinct and that while ICSH occurs in both sexes, LH is normally present in the female only. Its occurrence throughout life can be completely inhibited if a testis, either natural or implanted, is present during the development of an animal of either sex (Witschi, $955). The action of LSH, by inducing the secretion of progesterone, initiates the alternate, or dioestrus, phase of the female cycle, a phase which makes pregnancy possible and for which there is no hormonal counterpart in the male. This may not have been the primary function of progesterone, for it occurs in association with viviparous development in many vertebrates besides mammals; it is probably under the same endo- crinokinetic control throughout (Matthews, 1955). Interaction of gonadial and endocrinokinetic hormones in repro- duction - In the sexual reproduction of all vertebrates there is need for the gametes to ripen simultaneously in males and females of the same species, and for the young of most land forms to be pro- duced at a suitable season of the year for their early growth. The endocrinokinetic hormones play a vital réle in bringing this about. Mamma tia. In placental mammals the viviparous development of the young embryo in the maternal uterus and its subsequent nourishment from the mammary glands increase the complexity of the situation, so that several more hormones are needed to control reproduction in the female than in the male. In the male, the secretion of TESTOSTERONE is induced by ICSH, and maintains the reproductive ducts and the accessory organs in a steady, active state, as well as developing secondary sexual characters, such as antlers. At the same time the morphogenetic hormone, FSH, ensures the continuous ripening of sperm. In species with a limited breeding season, like deer, the onset and ending of these hormone-controlled changes appears to be related by the brain to environmental factors, such as temperature and length of daylight. 152 KINETIC HORMONES—OH CH20H CH20H D- Glucose D-Galactose Le Arabinose | Gor ees =| S ieee ata ae 7 | CHO! CH20H | CHO ! rie | Fae HO—-C—H C—O HO——-C——H moor rear Me emer ine n ge HO——-C——H gee HO——C——H H—C——OH Eee ae ee + eww ie wom a ———— 1 8 a ee ad H—C—OH Ho=g—-0H ae asic H—C——OH Ba eH CH20H H>OH CH20H D-Mannose D-Fructose D-Arabinose Fic. 5-10. The structure of some of the sugar molecules which have been found to be responsive to insulin and to resemble glucose in having the same side chains on the three terminal carbon atoms; and (below) three molecules of sugars which differ, however slightly, from glucose, and are found to be unresponsive to insulin (from Levine and Goldstein in Stetten and Bloom, 1955). of these facts to the known chemical structure of the insulin, which is a protein, is not yet within sight. The rate of secretion of insulin is self-regulating, in that a high level of blood-sugar increases insulin; and, as this lowers the level, so the insulin secretion itself is reduced. § 5.22 PROTEIN METABOLISM 199 There is no known hormone that stimulates the secretion of insulin, unless secretion is found to be related to the increased growth of the gland, which is said to occur as a result of injecting extracts of the posterior pituitary (Staszyc, 1956). This is as yet unconfirmed. The claim that STH increases insulin secretion is based on an indirect effect, since it only occurs in those species where STH stimulates glucagon to raise the level of blood-sugars, and this in turn stimulates the insulin secretion (§§ 4.222 and 5.521). 5.22 PROTEIN METABOLISM Hormones play a part in protein metabolism in Arthropoda and Vertebrata, but the results so far reported are not always clear. Noble (1955) points out that, in vertebrates, most attempts to assess nitrogen, which is a characteristic element in proteins, have been based on measurements of the over-all balance in the body, whereas a more realistic picture might be obtained by following reactions in different organs of the body separately. Increased protein catabolism in one organ may be accompanied by an increase in nitrogen excretion (§ 5.222), or it may be mainly responsible for supplying materials for protein anabolism in some other organ. ARTHROPODA. The same difficulties of interpretation are prob- ably true for Arthropoda. Perhaps the best that can be suggested at present is that an eyestalk hormone in Crustacea tends to re- strain protein breakdown and nitrogen excretion and that the Y-organ hormone may inhibit them (Table 24). In Insecta a brain hormone appears to be associated with an actual increase in new protein formation. No hormones are yet known to stimulate catabolism in either Crustacea or Insecta. 5.221 Restraint of protein catabolism Crustacea. It has been claimed that an EYESTALK HORMONE restrains protein catabolism and that removal of the sinus gland, which is the main storage organ of the hormone, results in protein breakdown and loss of nitrogen; but the evidence is ambiguous, partly because the results seem to vary with stages in the moult and intermoult cycle. Positive evidence shows that sinus gland removal causes a loss 200 METABOLIC HORMONES of nitrogen in the crab, Hemigrapsus. Sinus gland removal also leads eventually to moulting, because of the absence of the moult- inhibiting hormone (Part II, § 3); but it is probable that the time required for moulting would be more than the 23 days of the experiments summarized in Table 23 (Neiland and Scheer, 1953). TABLE 23. CHANGES IN BODY COMPOSITION OF CRABS (HEMIGRAPSUS NUDUS) FOLLOWING STARVATION AND SINUS GLAND REMOVAL All values are given on a wet weight basis and are means of measure- ments on two to four individuals (cf. Table 20). The changes in protein content, following the operation, are significant and are shown in italics (from Neiland and Scheer, 1953). | STARVED WITH STARVED NORMAL SINUS GLAND e 23 DAYS REMOVED Bex i anne MF sh Wi ee Body weight, g 10:5 9-3 9:5 7-4 10:3 7-4 Glycogen, mg/g 0-69 1:24 0:8 0:8 0:76 1:02 Protein, mg N/g 12-71 14:96 | 12:08 12-08 10:39 10-40 Chitin, mg glucose equivalent/g 3:91 4-23 3:89 4-09 3:49 4-14 The results may be taken to represent the intermoult situation. The effects of fasting and the technique of SINUS GLAND removal in Hemigrapsus have been referred to already in relation to reduc- tion of fat (§ 5.112). Their effect upon glycogen “which might be regarded as the most logical source of glucose and of chitin’’, is not appreciable, and supports the suggestion that the crabs were not near moulting, when new chitin is formed. There appears to be a certain weight loss, which is reasonable in starved animals; but it would not occur in those from which removal of eyestalks was inducing moulting, as this is accompanied by increased water uptake (§ 5.321). It is claimed that in these conditions the reduction in total nitrogen is significant, and represents the effect of removing an eyestalk hormone that normally restrains protein catabolism. § 5.221 PROTEIN METABOLISM 201 This hormone may be the same as the moult-inhibiting hormone. Increased loss of proteins following eyestalk removal is also claimed for Carcinus from measurement of nitrogen excretion. This is partly due to the stress of wounding, since a similar effect, but of only half the magnitude, results from leg amputation (Needham, 1955). As no hormone injections were used to counter- act this effect, the evidence is not conclusive; but it again suggests that in non-moulting crabs an eyestalk hormone restrains protein breakdown. | During moulting the situation is different. Although the protein content of the plasma is unusually high because of resorption from the old skin, nitrogen excretion is particularly low. Needham (1957) concluded that ‘“‘within limits the animal is able to control its nitrogen output, whatever the external conditions”. As moulting proceeds most of the protein from the plasma is presumably deflected into certain anabolic processes, such as the formation of the new integument. These protein transfers would not of them- selves alter the total proteins in the body. Nor does starvation affect the issue, since most crabs do not feed for some days before, during or after moulting. Koch (1952) confirms the view that the nitrogen and protein content of the body remains remarkably constant during moulting, and is not affected by the absence of any eyestalk hormone. Incidentally, the moult-inhibiting hormone is not released from the eyestalk at this stage (Part II, § 3). Koch examined the nitrogen content of the mitten crab, Eviocheir sinensis, and found that the eyestalks have no effect upon the nitrogen metabolism, at least during the first experimentally induced moult. There is no difference in the ratio of nitrogen content to size, in the cast skins of crabs moulting normally and those induced to do so by eyestalk removal. The ratio of total nitrogen content to carapace width measured before moulting is the same, within limits, for moulting and non-moulting control crabs, and for operated crabs (between curves, Fig. 5-11). After an induced moult, the ratio of nitrogen in body and cast skin to carapace width, measured after moulting in the operated crabs, is significantly lower (VI, Fig. 5-11) than before, by an amount that seems to be proportional to the increase in water content that follows from the absence of the diuretic 202 METABOLIC HORMONES hormone of the eyestalk (§ 5.321); but this is not shown quantita- tively. Excessive water uptake cannot afford an explanation for the contrary findings in Hemigrapsus, where there is no weight in- 800 600 Total nitrogen, 400 30 SO 60 70 Carapace breadth Fic. 5-11. Relation of the total nitrogen content of the crab, Erio- cheir sinensis, to its size before and after moulting. The recorded values fall into three categories. (1) Total nitrogen in body (ordin- ates) measured before moulting and plotted against carapace breadth (abscissae): @ for non-moulting control crabs. (ii) Total nitrogen in body plus that in cast skin measured immediately after moulting and plotted against carapace breadth of old shell: © for control crabs moulting naturally; [_] for eyestalkless crabs after forced moult. (111) Same total nitrogen as in (ii) plotted against carapace breadth of new shell: & for controls that still fall within normal range shown by curves; P| for eyestalkless crabs after forced moult due to loss of moult-inhibiting hormone, MIH. ‘These show markedly low ratios and it is assumed that though loss of MIH does not affect nitrogen metabolism (11), the loss of the DIURETIC HORMONE in the eyestalk allows increased water imbi- bition and swelling that increases the crab’s size relative to its nitrogen content (from Koch, 1952). crease (‘Table 23). Whether the abnormally large volume increase observed during the first moult, following eyestalk removal in Eriocheir, might be followed later by a correlated increase in tissue synthesis remains to be investigated. Recent evidence on moulting hormones makes a tentative interpretation of these events possible, § 5.222 PROTEIN METABOLISM 203 if the apparent difference in nitrogen excretion in the intermoult and moulting periods may be taken as valid. The intermoult period is characterized by secretion of the moult-inhibiting hormone, MIH, and the lack of any Y-organ secretion. Removal of the eyestalk, or even of the sinus gland, allows the Y-organ to come slowly into action. Normal moulting occurs when the secretion of MIH stops naturally and the moult- promoting hormone, MPH, from the Y-organ becomes active (§ 4.211 and Table 24). From this it appears that during intermoult, the amount of protein catabolism, probably accompanied by steady synthesis, is restrained by MIH (as has been postulated) since lack of MIH results in increased catabolism. During moulting, catabolism and N-excretion are practically inhibited; since this cannot be attributed to the lack of MIH, it may perhaps be correlated with the presence of MPH from the Y-organ, which was absent during the intermoult period but is now active. ‘The lack of excretion at this stage is accompanied by evidence of transfer of proteins to the new integument, and is a process so clearly related to moulting that it might well be under the control of the MOULT-PROMOTING HORMONE. 5,222 Increase in protein synthesis INsEcTA. Evidence from both Periplaneta (Bodenstein, 1953) and Calliphora shows that removal of either the MEDIAN NEURO- SECRETORY CELLS OF THE BRAIN or of their stored products in the CORPORA CARDIACA results in reduced protein synthesis; in Pert- planeta this is shown rather indirectly by the disappearance of urates from the fat bodies, and their re-formation after re- implantation of the corpora cardiaca. In Calliphora an effect upon protein synthesis is deduced from the reduction or cessation of growth of the ovaries, accessory glands, oenocytes and corpora allata after ablation of the neurosecretory cells (E. Thomsen, 1952 and 1956). The effect is similar to that of keeping the flies on a protein-free diet of sugar and water. The effect of removing the corpus cardiacum is similar to, but not so profound as, removing the source of the neurosecretion. The removal of the corPorA ALLATA of Carausius is followed by an increase in amino acids in the tissues, which is interpreted as METABOLIC HORMONES 204 *yUITUNS94UI MOU O} JOFJSUBIY u19301d jo UOTJE[NUTYs pue UOTIIO -Xd-NJ jo mOBIETUE JenqaIA YIM poyeroosse Afussiedde ‘peasy ul ues10-X wo1y suOWIOY SuTOWOId-}JnNoW = F{gIV “UIsITOqe}e9 SUIUIBI}SII OS|R Ayyuoredde ‘yyeysoAo UI DUOULIOY SUTIQIYUI-I[NOW = FIV yuasoid yuosoid (AjaAT}e.190d0) juasqe (uor}9199S OU) juosqe (aduvys ou) SNUIIAD’) pue AY ION (Qquountiodxs surlinp poja199s 10U [[Ns) quasqe (perqrqur) yUISqe (AJaAtv19d0) yuasqe yuasoid SNUIIAD’) pue snsqvastia FT HdIN NOILV1NOUIO HIIW NI S€aNOWYOH SdVuoO NI NOILAYOXA NHOOULIN HLIM GALVIOOSSVY SHNOWYOP ‘$7 ATAV [, MOT porAsowol MO] youyur LTINOJ PeseotOUr pyI9AoWII unIpout 398 UT LINOWUYALNY SN TV.LSIAT NOILAYOXA- NT 4O ALV.LS SdVuyo § 5.223 PROTEIN METABOLISM 205 showing inhibition of protein synthesis (L’Hélias, 1953). This hormone, like that of the brain and corpora cardiaca, would there- fore presumably favour protein synthesis. It seems that secretion of these hormones in arthropods is stimulated mainly by the nervous system, with little trace of endocrinokinetic control (§ 4.21), unless the neurosecretion from the brain stimulates the secretion as well as the growth of the corpora allata; but on this the evidence is inconclusive (Thomsen, 1952). The neurosecretion does not pass into the blood. 5.223 Decrease in protein synthesis or increase in protein catabolism Crustacea. Evidence that in Carcinus the TIPS OF THE EYESTALKS stimulate catabolism, which is decreased by their removal, suggests the possible seat of a hormone that is antagonistic to those of the rest of the eyestalk (§ 5.221) ; but it is as yet unidentified (Needham, 2955): VERTEBRATA. The hormone chiefly connected with increasing protein catabolism and possibly restraining synthesis in the vertebrates is one of those from the ADRENAL CORTEX (Chester Jones, 1957a); but results as yet seem to be rather contradictory, and it is not clear whether this is really due to specific differences, or to the restriction of different reactions to different organs in the body. Mamma ta. The state of nutrition of the animals is important: in starvation, the administration of adrenocortical hormones of the HYDROCORTISONE type causes a much greater increase in protein catabolism than it does in well-fed animals. Both stress and ACTH (§ 4.231) injection into nephrectomized rats cause an increase in urea formation, which is restrained by glucose injec- tions. In part, this may be due to the concurrent action of the hormones concerned with carbohydrate metabolism (Noble, £955). It is clear that the endocrinokinetic control exercised by ACTH, from the adenohypophysis, plays an important part in these animals, in contrast to the situation in the Arthropoda. 206 METABOLIC HORMONES 5.3. BALANCE OF MONOVALENT ELECTROLYTES AND WATER The control of salts and water in the blood and tissues is of importance to all animals living in environments with which they are not in osmotic and ionic equilibrium; but evidence of this control being exerted by hormones is almost confined to the vertebrates. Hormonal control of these factors among invertebrates is as yet only known in relation to water in a few arthropods. There are, from the point of view of basic physiology, two contrasting situations in which an animal must be able to control its salt and water content if it is to survive. In the first, the blood and tissues tend to lose salts and become diluted by imbibition of water. This can arise when an aquatic animal moves into any hypotonic medium; but it is particularly associated with the migration from the sea to fresh water. Since the salt losses cannot, as a rule, be made good merely from the food, most freshwater animals not only reabsorb salts from their urine but also absorb them from the environment against considerable osmotic gradients. This involves the active transport of ions across certain cell membranes (cf. Kitching, 1957). There are various hypotheses concerning active transport, but they may all “‘be expected to contain the following principles: (a) The cations [such as Na*] are not transported as the free ions, but are bound in some complex, which operates as a carrier ; (b) This carrier operates in a cyclical manner across a mem- brane. Towards one side of the membrane it combines with the ion and towards the other side it releases the ion, the process being combined either with an exchange for another cation or it is accompanied by an anion [such as Cl7]. These associated exchanges may or may not be active in turn ; (c) The release of the ion must be associated with a simul- taneous release or transfer of the necessary energy if osmotic work is being done”’ (Conway, 1956). At the same time, water which enters the tissues by endosmosis from a hypotonic medium must be baled out by some form of § 5.3 BALANCE OF MONOVALENT ELECTROLYTES AND WATER 207 increased diuresis, either by a contractile vacuole or some other form of excretory organ. Movement of both salts and water may be controlled by hormones. In the second situation, the salts in the blood and tissues tend to become concentrated by water loss, with or without simultaneous salt endosmosis. This can occur when animals migrate to land where dehydration and thirst are the main dangers, even though water losses through the skin may be virtually abolished; for losses through the respiratory surfaces and by excretion are inevitable. ‘The main response to these water losses is hormonally controlled antidiuresis, involving reabsorption of water from the urine. In addition to this, the Amphibia and some insects can absorb water through the skin. Dehydration and salt increase also occur when animals move into any hypertonic aquatic medium; for instance, when teleosts migrate back to the sea, after having acquired a lowered salt content during their sojourn in fresh water. Here the necessary dilution and proportion of the salts in the blood is maintained, chiefly by active excretion of sodium and chloride ions either through the gills or the kidneys; there may also be antidiuresis, but the action of hormones that control this in fish is less clear than in tetrapods. Although in a stable environment little control may be required, most animals maintain a dynamic equilibrium between these two extremes; for the normal response to either hydration or dehydration (if carried to the limit) would lead to the opposite state being reached in the tissues. Control that allows of a variable response is demanded in particular of those animals which migrate periodically, like some estuarine crustaceans and fish, between fresh waters and the sea. They must be able to contend alternately with flooding of their tissues with water in one environment, and their dehydration by the high salt concentration of the other. In most of these cases, however, all too little is yet known of the part played by hormones. In those mammals, where the position is best understood, four hormones are probably concerned: one antagonistic pair to control the movement of salts and another pair for the water bal- ance. Complete elucidation is still difficult, not only because the movements of salts and water are so intimately linked by osmotic forces, but also because the associated hormones, active in a given METABOLIC HORMONES 208 s[BUIUe AY Sparel sopndoay Be PES Aoupry oe Hav é purls uTys (91991nIprjue) SisaAmipyuv puv snqdo1I9 AIBATTES SOG é SJSO9]9 J, sts{ydodAyoinayy Ajipiqvamutsad ut asvasquy Too S[BUIUIR A] (aurxo14y}) $}SO9[9 J, prorAy], S[PUIUUe JAY é vIudvig BOBIPIVS *d sopndoy SNSADIOSIU FT to /pue ureig (auOstIODOI1pAY) SNUIIAD’) (Aoupry) vung X9}109 [VUSIPY | snaps | pues snurs /uesio é sntuur}s SisaAnip puv snapqup’) -X-oruolsuey $}SO9]9 J, jo soposndioy | Ay2jiqvatusad ut aspaazaq—s | ZE"S AONVIVE WALVAA = TES — — s[ePuUTUe yy é “5 U01JUdJAL 4S (uOI}a19x9) s[eUIWe I (uI90}4x0) poo]q ut _|{_d — — pojoxy | stsAydodAyoinony puv en fo asvpaszaq §=ZIE'S — — s[RPUIWIR Ay es UOMAAIXI 4ST s[RPUIUUe]y (uoNdiosqeai 9A1}9R) so[doy (9U019}SsOpye) poo]q ut _{D -— --- DUDY X9}109 [BUDIPY puv .eN ut asvaanut = E's | GONVIVA LIVG [E'S ATM NVXa ANOIWYOH YO NVOUO ATH NVXd ANOWYOH YO NVOYO pe ALVUGALYAANI ALVUGLLYAA WaLVM ANV SULA TOULOUTA ONITIOWLNOO SANOWYOH OITOEVLAJ “SZ HTav T, $5,311 BALANCE OF MONOVALENT ELECTROLYTES 209 situation, are derived from the same source and are closely related chemically (Fig. 5-15). There are indications of a similar type of hormone control occurring in other vertebrates ; but the evidence is less conclusive, and workers are not in full agreement. There is, as yet, practically no evidence to show whether the same form of control occurs in any invertebrates. The hormonal control of salts will be considered first (§ 5.31), and that of water afterwards (§ 5.32), since the direction of move- ment of the water is so often determined by that of the salts. 5.31 BALANCE OF SODIUM IONS (Nat) AND OF ASSOCIATED MONOVALENT ELECTROLYTES (K+ anp Cl-) 5.311 Increase of Na* in the blood Active transport of ions across living membranes and especially the so-called “‘sodium pump” are now well-known phenomena in invertebrates as well as vertebrates. INVERTEBRATES. Both crustaceans and insects living in fresh water are known to take up sodium and chloride ions from the surrounding water by active transport, through the gills in Eriocheir, and by the anal papillae in the larval Chironomus; but, as yet, there is no evidence of any hormonal control of these activities. Although the isolated gills of Eriocheir make a beautiful preparation for demonstrating active transport of sodium, they have not apparently been examined for the effects of possible hormones; drugs have, however, been shown to have closely comparable actions here and on frog skin (Koch, 1954). VERTEBRATA. A number of cases now indicate some hormonal control of active transport of ions across cell membranes in vertebrates. This transport involves the use of metabolic energy, derived from aerobic respiration, to overcome the osmotic gradient. The action is stopped by lack of oxygen. Since among cold-blooded vertebrates much more is known of the situation in Amphibia than in Teleostei, they will be considered in that order. Concentration of the blood by inward passage of salts through the skin Ampuisia. The skin of the frog plays an important part in maintaining the internal salt concentration well above that of 1% 210 METABOLIC HORMONES the freshwater medium, and its action is facilitated by hormones of the ADRENAL CORTEX. 015 Rana temporaria Bufo bufo Rana esculenta 0:10 0-05 Sodium uptake, z equiv. /em?/hr Intact Intact Tsolated skin (a) (0) (c) Fic. 5-12. Increase in the apparent rates of active sodium-ion uptake through the skin, produced by salt-depletion of (a) the intact common frog and (b) the toad (after Jorgensen). (c) The apparently similar effect of Pituitrin (a neurohypophysial extract) on the isolated skin of the edible frog (from Fuhrman and Ussing, in Sawyer, 1956). In the intact animal there is evidence for independent absorption of both sodium and chloride ions against the osmotic gradient (Jorgensen, Levi and Zerahn, 1954), from external concentrations as low as 10-° M.NaCl. Like the gills of Erviocheir, even the isolated frog’s skin passes ions through itself from the outer surface to the inner (“‘controls”’, Fig. 5-12 a—-c) by active transport. This trans- port is increased in intact frogs and toads after the tissues have been depleted of salt by ‘‘washing”’ in distilled water for some time (Fig. 5-12a and 5). Since salt uptake is reduced in adrenal- ectomized animals, it is assumed that hormones of the adrenal cortex may stimulate the active transport of sodium ions inwards through the skin, as they do through the kidney tubules (see below). Despite the fact that the action of extracts of the adrenal cortex $ 5.311 BALANCE OF MONOVALENT ELECTROLYTES 211 in causing a marked increase in Cl~ uptake is similar to their action on Na*, it cannot be assumed that the transport of chloride ions follows passively upon that of sodium, since the rate of uptake of the two ions through the skin has been shown to be independent. In the intact animal the secretion of the adrenal cortex may be stimulated by adrenocorticotrophin ACTH, (§ 4.231), from the adenohypophysis. The effect of Pituitrin, an extract in this case of the neuro- hypophysis of a whale, on the sodium uptake of isolated skin 100 Fercentage reabsorbed O a fe) 15 Sodium filtered, mequiv/kg/ hr Fic. 5-13. Relationship between the percentage of the sodium reabsorbed from the kidney tubules (ordinates) and the total amount of sodium filtered through the glomeruli of the kidneys in m.equiv/kg/hr (abscissae) in the bullfrog, Rana catesbiana. ‘The percentage is high over a wide range of concentrations of sodium in the tubules (from Sawyer, 1956). (Fig. 5-12c) seems anomalous. The Pituitrin used was believed to be free of anterior lobe contamination, and therefore of ACTH, but oxytocin and antidiuretic hormone, ADH*, were certainly present in the extract and the latter caused so great a concurrent uptake of water that, although the absolute amount of sodium was increased as shown, the net effect was actually dilution (Sawyer, 1956). It would be curious, in view of the situation in other vertebrates, if neurohypophysial hormones should aid the uptake of salts in amphibians. * — vasopressin. Z12 METABOLIC HORMONES Concentration of the blood by reabsorption of salts through the kidney AMPHIBIA. In most circumstances, and especially in fresh water, the function of the kidney is to reabsorb sodium (Nat), and possibly chloride ions (Cl~) actively, and to excrete a certain amount of potassium (Kt). Figure 5-13 shows that, over a wide range of sodium ion concentrations in the glomerular filtrate, at least 95 per cent is reabsorbed in the tubules. Active transport of sodium here is comparable with that through the skin. In adrenal- ectomized frogs sodium is excreted more rapidly (presumably because less is reabsorbed) than in normal controls (Fig. 5-14). There is a slight concurrent increase in muscle potassium (Fowler, 1956). Moreover, saline treatment can maintain life in the adrenal- ectomized frog, as it can in mammals. This seems to support the case for supposing that an adrenocortical hormone (ACH) is concerned in stimulating salt reabsorption in the kidney tubules of frogs, as in mammals. It may well be that ACH secretion in Amphibia is under the endocrinokinetic control of ACTH (§ 4.231), since hypophysect- omy is followed by symptoms closely similar to those resulting from adrenalectomy ; other evidence is, however, defective (Chester Jones, 1957a). TELEOSTEI. The established function of the adenohypophysis in protecting some teleosts, such as Pundulus (Burden, 1956), against the osmotic stress of transfer from sea water to fresh is probably endocrinokinetic; it seems to favour the retention of sodium and, more particularly, of chloride ions; but it is not yet clear if this is due to the release of adrenocorticotrophin, ACTH (§ 4.231), or of thyrotrophin, TSH (§ 4.221). Neither of these are replaceable by the corresponding mammalian hormone, and it has even been postulated that some other “unknown factor”? may be present in the hypophysis of euryhaline teleosts. ACTH might be expected to stimulate secretion by the ANTERIOR INTERRENAL BODIES, which are the homologues in fish of the tetrapod adrenal cortex; but they do not seem to have been investigated in this connection (Rasquin and Rosenbloom, 1954). The effect of injecting extracts of adrenal cortex into trout has only been examined in the converse situation of their being exposed to excess salinity in sea water (see below). Seve! BALANCE OF MONOVALENT ELECTROLYTES 213 The suggestion that the hypophysis acts by secretion of TSH arises from the fact that the THYROID, which it would stimulate, is active during the upstream (anadromous) migration of salmon and 25 20 mg 5mg Noa ie) injected Sodium excreted , 05 Hours Fic. 5-14. Loss of sodium by excretion (ordinates) in hours (abscissae) from nine adrenalectomized frogs, Rana, (above) and from nine controls (below). After 24 hr in distilled water, (at hour 0) 5 mg Na (as frog Ringer solution) was injected into the dorsal lymph-sac of each, and excretion was recorded for the first 4 hr and at the end of 24 hr (points shown with standard deviation). Note the much greater excretion of sodium from the adrenalectomized frogs (from Fowler in Chester Jones, 1957). eels from the sea. Since these fish show a concurrent phase of growth and maturation, it seems more probable that the thyroid 214 METABOLIC HORMONES Cortical Neurohy pophysial hormones _ hormones _ i dieee in antidiuresis and blood concentration and blood dilution Glomerulus ACTH NEUROSECRETION Bowmans capsule | Oxytocin 3 i 7 _* ~ Aldo- [| Proximal sterone = > tubule ? Loop of Henle Hypotonic Kt sterone Distal tubule Hydro - LF 0->| Markedly cortisone [ #| <= | hypotonic Collecting duct To ureter and (a) bladder Isotonic Hypertonic urine urine c———> Active transport : > Jon exchange ----—> Diffusion — Concentrating mechanism (sanz, Hormone favours process “=ass==={ Hormone inhibits process Fic. 5-15. Diagrams to show the action of hormones on the kidney tubules of a mammal. a. The cortical hormones, active in blood concentration: ALDOSTERONE, increasing reabsorption of salts, and HYDROCORTISONE, increasing water diuresis; obligatory endos- mosis is then limited by the diffusion rate to about 80% of com- pletion. In the loop of Henle the urine still has ‘‘free’’ water. b. 'The neurohypophysial hormones, active in blood dilution: Paobl BALANCE OF MONOVALENT ELECTROLYTES 215 may be concerned with these processes than with osmotic regula- tion. Yet osmotic regulation also requires an increase in metabolic energy that might be stimulated by thyroxine (§ 5.111); it is noteworthy that the thyroid is more active in conditions of changing salinity than in stable conditions, even if these are abnormal (D. C. W. Smith, 1956). Experimental investigation of fish is clearly fraught with many difficulties; recently Jorgensen and Rosenkilde (1956) have issued a timely warning, in the form of observations on the very wide range of spontaneous variations that occur in the chloride content of relatively undisturbed and undamaged starving goldfish. RepTILia. Little is known of the electrolyte control in this class of vertebrates, but it is attractive to speculate that secretions of the ADRENAL CORTEX may act at the renal tubular level in reptiles to increase potassium excretion and sodium retention (Chester Jones, 1957a). Mamma tia. Hormones from the ADRENAL CORTEX (Fig. 5-15a). facilitate active transport of ions from the lumen of each convoluted kidney tubule into its cells, the surfaces of which are increased by a brush border. Reabsorption of sodium (Nat) and potassium ions (K+) from the glomerular filtrate, which is isosmotic with the plasma, occurs in the proximal tubules. Chloride ions (Cl—) also move back into the cells in the same region, either by independent active transport, or in company with the cations, because of the difference in electric potential set up by the cation transport across the cell surface. Cortical insufficiency, or removal of the adrenal cortex, is followed by loss of Nat from the plasma and tissues and its excretion in the urine (Fig. 5-16 and Table 26). The sodium OXYTOCIN, increasing salt excretion so that endosmosis can keep pace with the reduced rate of salt reabsorption; and ANTIDIURETIC HORMONE (vasopressin or ADH), increasing water reabsorption and reducing urine output (antidiuresis). The reactions in the tubules are similar to those in the Amphibia; but the mammal is peculiar in having a urine concentrating mechanism, probably in the collecting ducts (large arrows). More concentrated urine is indi- cated by closer striations. 216 METABOLIC HORMONES /& equiv. cm3 Sodium or potassium , Urine, Fic. 5-16. Volume of urine and the micro-equivalents of sodium ions excreted by normal (control) and adrenalectomized (expt) male rats, Rattus, over a 4 hr period after administration of water by stomach pump (roughly 11 ml was given to each rat, according to its size, 1 hr before the record started and again at hour 0 to inhibit ADH secretion). Food was removed 24 hr beforehand. Note the poor water excretion by the adrenalectomized animals (lack of DIURETIC HORMONE) but their very high output of sodium (lack of ALDOSTERONE, cf. Fig. 5-15). The simultaneous output of potassium, though not shown, was little altered (from Chester Jones, 1957). balance can be restored by injection of ACH, particularly aLpo- STERONE, though corticosterone is also effective (Chester Jones, 1957a). Although hydrocortisone can be present in 30 times the concentration of aldosterone in the blood, the latter is 300 times as potent in controlling the Na+ balance and possibly that of Cl—. It also has an action on the Nat+/Kt ratio of salts excreted by the sweat and salivary glands (Simpson and Tait, 1955). Table 26 shows that the effect of adrenalectomy on the plasma potassium, K+, in rats is opposite to that on sodium. This is in part due to the effect of ion exchange in the distal tubule (Fig. 5-15a). Sodium loss and potassium retention seem to be major factors in causing the death of adrenalectomized animals, though the former is the more important, since life can be maintained, despite high blood potassium, as long as a high level of sodium chloride is given in the diet. BALANCE OF MONOVALENT ELECTROLYTES 217 § 5.311 ‘aay tod sjyusyeambe-1yprTu y L9-0+ 00-92 6£-0+ 09-92 67-0 + 8Z°SZL FeO CSL UaALVM 7 78: 1+ | S8I-SOT | 7614 80-011 3 9C:66 66:0 + 9¢-001 M 09:0+ 89-61 6b 1+ ££-81 8b 1+ £6-S7 Leos 6E-f¢ YN “LM LOM *OM/OdIN aTOSON SL-0+ OL-E+ £0-9 9L-9F1 €l-O+F OCF $8-ZL C8-Tel 61-0+ CSobse C6°P OF-9F1 CC'O+ COVE 96°F 9¢-Srl wal YN y |/OAN VINSV1d O$-S-r OS-961 06-S + 88-681 19°O S8°921 +64 + £9-061 3 LHOIM -AGOd Ot cl Gi SNHWI0ddS IO WaaNON sAep Q IO} pozru0j99] -euaipe ‘snprdisur sojaqeiq, sAep Q 10F PEZIUI0}JSI[VUSIpe S[OIZUOD sy}UOUI ¢ 0} Z 10} snprdisur sajoqeiq, s[O.1}U07) dNOwD "(@ LS61 ‘souof Ia}soyD WIOIJ) JOIIO piepuvjs | ‘suvaur oie soinsy ouy, “(HCV JO 204nos ay} st yorum ‘stsAydodAyoinsu I19y} Jo [BAoUaI AG poonpur) snprdisutr sajoqerp NoyIIM pu YIM ‘ser a[VUoJ Jo sdno1s JOF UMOYS Iv sasuLYo 9U, J, AWOLOFYTVNAYGVY ONIMOTTIOA ‘ATOSAW ANY VINSY 1d JO NOLLVULNAONOO WAISSVLOd GNV WAIdGOS NI SHONVHZ ‘97 ATAV J, 218 METABOLIC HORMONES Aldosterone is secreted in response to adrenocorticotrophin, ACTH, in rats, but not in the human (§ 4.231). 5.312 Decrease of Nat in the blood There is no evidence for any hormones stimulating Na+ and Cl- excretion in invertebrates, and so far it is rather tentative in vertebrates, especially in the cold-blooded classes. AMPHIBIA. It has been shown that injection of a neurohypophy- sial extract containing OXYTOCIN increases Cl~ excretion in the axolotl (Sawyer, 1956). _ "TELEOSTEI. Most marine teleosts maintain their body fluids at a lower level of salt concentration than that of the sea, by means of specialized salt excreting cells on the gills. If any of the endocrine organs are concerned in stimulating this process, their action has not yet been proved. Extracts of mammalian adrenal cortex, ACH, have been injected into trout in sea water, and have been found not to increase their survival time in the excessively saline medium. Despite the fact that in other vertebrates ACH causes an increase in salt reabsorption rather than its excretion, the author suggests trying even larger doses (D. C. W. Smith, 1956). A single test of mammalian neurohypophysial extract likewise failed to increase the survival time of these trout, although it increases salt excretion in mammals. It is known that other teleostean hormones are very specific; also that the neurosecretory store in the neuro- hypophysis of Callionymus is depleted when this teleost is exposed to a hypertonic medium (Arvy, 1957). It might, therefore, be expected that extracts of fish neurohypophysis might be more successful than mammalian hormones in increasing salt excretion. On the other hand, salt excretion by the gills may be under quite other control than that by the kidneys. Aves. The nasal glands of the cormorant, Phalacrocorax, and probably of other oceanic birds* secrete salts, enabling them to eliminate excess Na+ and Cl~ taken up from any hypertonic media in which the birds may feed (Schmidt-Nielsen et al., 1958). It is not yet known if this active transport is under hormonal control, as might be expected. * 'The need for some such mechanism for flamingoes feeding in highly alkaline water had already been indicated (Jenkin, 1957). § 5.32 WATER BALANCE 219 MamMALIA. Removal of the neurohypophysis, and therefore of the supply of oxyTocIN from rats (last line of Table 26), mitigates considerably the effects of adrenalectomy, at least as far as plasma sodium is concerned. Plasma sodium is practically equivalent to that of the control, and potassium is lowered, as compared with specimens that had had the adrenals only removed; plasma potassium is, however, not fully restored to normal, and the balance of ions in the muscles is decidedly abnormal. A series of such experiments shows that the neurohypophysis is at least partially responsible for the loss of sodium in adrenalectomized animals, and that its action must be to inhibit the active tubular reabsorption of sodium ions (Fig. 5-155). Its action on chloride ions may be similar to that on sodium; but that on potassium is, as yet, far less clear (Chester Jones, 1957a). The interesting if tentative suggestion has been made that the effect of neurohypophysial removal may be due to the loss of the oxytocin secretion, rather than the antidiuretic fraction, ADH. Injections of oxytocin, at least in pharmacological doses, are more effective than ADH in increasing sodium excretion in dogs with a low rate of urine flow (Brooks and M. Pickford, 1957). If this last suggestion is further substantiated, the situation would be that sodium as well as chloride ions, at least in rats (Dicker and Heller, 1946), are controlled by the balance between aldosterone, favouring their reabsorption, and oxytocin, favouring their excretion. Potassium ions are excreted during periods of aldosterone activity. Secretion by the adrenal cortex is usually stimulated by ACTH, but this has less effect upon aldosterone than on other cortical hormones; the only control of neurohypophysial secretion is nervous. 5.32.5 WATER, BALANCE The direction of movement of water through a cell surface is determined by osmotic forces; but the rate of movement can be affected by hormones, which vary the permeability of the surface. Decrease in permeability, which is associated with diuresis, will be considered first (§ 5.321), as it tends to concentrate the blood salts and to accompany salt reabsorption (§ 5.311); increase in permeability associated with antidiuresis will be taken second 220 METABOLIC HORMONES (§ 5.322), since, like salt excretion (§ 5.312), it tends to dilute the blood-salts (Table 25). Such hormones are well-established in Arthropoda as well as in Vertebrata. In the latter, the hormones tending to concentrate the blood come from the adrenal cortex, and those tending to dilute it from the neurohypophysis, as they do for the control of Nat and Cl~ (Fig. 5-15). 5.321 Deerease in cell permeability and diuresis leading to con- centration of the blood CrusTAceA. Water uptake is a normal accompaniment of moult- ing, and probably plays a crucial part in helping to force off the old cuticle. In the crayfish, Cambarus, eyestalk removal induces both precocious moulting and abnormally great water uptake (from 23 % increase in body weight 0 3 4 6 8 io 12 4 16 Ig 20 Time, days Fic. 5-17. Percentage increase in water content (ordinates), as measured by changes in body weight in the crayfish, Cambarus immunis and C. propinquas, during 16 days (abscissae) after eye- stalk removal. Black circles show values for one specimen with two SINUS GLAND implants (curve A). This specimen shows less water uptake than the mean for eight eyestalkless controls with no implants (open circles, curve B). (From Scudamore, 1947). to 50 per cent of the body weight) during premoult; but this can be reduced again to about the normal level by implantation of two SINUS GLANDS (Scudamore, 1947; Fig. 5-17). Opinions differ as to whether eyestalk removal has a significant effect upon the water § 5.321 WATER BALANCE 221 uptake of the fiddler crab, Uca (Scudamore, 1947 and Guyselman, 1953); but in the shore crab, Carcinus, as in Cambarus, eyestalk removal before moulting results in a volume increase of 180 per cent instead of 80 per cent in the normal crab. Injected sinus gland extracts reduce the former value to 80 per cent and the latter to 50 or 60 per cent (Carlisle, 1956). At premoult, when the natural secretion of moult-inhibiting hormone begins to fade out, there is known to be an increase in internal osmotic pressure, owing to the mobilization in the blood of materials resorbed from the old skin. In forced moults, following eyestalk ablation, water absorption is greater than in natural moults, although this mobilization proceeds more slowly. The change in internal osmotic pressure in forced moulting cannot therefore be the only cause of increased water uptake. The loss of eyestalks and their hormones must increase the permeability of the tissues to water, as well as inducing the moult and the increase in osmotic pressure. Swelling at the time of natural moulting must then be due to a decrease in the secretion of this hormone, but not to its complete cessation. This would allow the skin, and particu- larly the branchial epithelium, to become more permeable than in the intermoult stage, but not as permeable as in forced moults. Increased permeability of the excretory tissue would also allow of increase in reabsorption of water from the urine, but there seem to be no figures relating to urine flow during moulting. Even under natural conditions in sea water, there must be considerable water endosmosis in Carcinus to maintain the high rate of urine flow, amounting to 14 per cent of the blood volume daily. The necessary internal osmotic pressure to achieve this, (unless active transport of water is to be postulated) must be partly due to active inward transport of such ions as Nat, Cat* and Cl- ; but this is small, though it may, perhaps, be aided by the presence in the haemolymph of ionized proteins, which do not pass into the urine. At the same time the endosmosis must be limited by a certain degree of impermeability of the tissues, maintained by the eyestalk hormone, otherwise eyestalk removal would not result in increased water uptake. It therefore seems reasonable to postulate that the action of one of the eyestalk hormones is the same as that of other diuretic hormones, namely, 222 METABOLIC HORMONES to decrease the permeability of the skin and of the excretory organs. Inadilute medium such as 50 per cent sea water, active transport of ions is increased, gill permeability is decreased, and urine flow is increased to about 24 per cent of the blood volume daily. At the same time there is a limited swelling of the tissues (Webb, 1940). The increased diuresis is presumably achieved by an increased secretion of the diuretic hormone from the sinus gland causing increased impermeability of the kidney tissues, as well as of the gills. Carlisle (1956) has claimed that the DIURETIC HORMONE of Carcinus differs from the moult-inhibiting hormone that is also obtained from the eyestalk, because only in the winter, at least at Plymouth, could he extract a moult-inhibiting substance, even from the sinus glands of the same species; extracts of SINUS GLAND from a number of other Crustacea were effective at all times of the year in reducing water content. The diuretic extracts could not be obtained from the cerebral ganglia, although these yield a strong moult-inhibiting extract. Passano (1953) claims that the differ- ences are quantitative and that the tissues have a lower threshold value of sensitivity to the water-balance effect than to the moult- inhibiting hormone. The observation that eyestalk removal reduced deaths of Carcinus from exposure for 24 hr to conditions of lowered salinity (Knowles and Carlisle, 1956) remains obscure. Survival would seem to require the presence, rather than the absence, of the diuretic hormone in the eyestalks. (Carlisle, 7m lit. 4.3.1957, agrees that the situation is far from clear, and thinks that perhaps in Crustacea, as in vertebrates, the balance of water, or of water and salts, is under the control of two hormones. ‘The moult-promoting hormone from the Y-organ may be one of them.) INsEcTA. A DIURETIC HORMONE has been claimed for the beetle larva, Anisotarsus cupripennis, and perhaps for other insects such as Blaptica (Nufiez, 1956). It appears to be secreted by the BRAIN, and is possibly stored in the CORPORA CARDIACA, which were not separated from the brain in the experiments. The larva lives in a damp environment and can take up water by endosmosis through the integumental cells lining the tracheoles ; but it normally remains constant in size and weight by excreting the excess water into the e532) WATER BALANCE 223 Malpighian tubules (Fig. 5-18, upper specimen, and Fig. 5-19a, dotted curve). If either the neck is ligatured or the head is cut off, preventing any flow of hormone to the body, or if the source of hormone is removed by ablating the upper part of the brain, the operated specimen swells steadily by retention of water (Fig. (a) Fic. 5-18. Water uptake in larvae of-the beetle, Anisotarsus cupri- pennis. a. Normal larva, and b. operated larva to show the swelling due to water uptake after removal of the upper part of the brain, including the CEREBRAL NEUROSECRETORY CELLS, and the corpora cardiaca (from Nufiez, 1956). 5-18, lower specimen, and Fig. 5-19a, full curve). Injection of brain extract restores the water balance, presumably by increasing excretion (Fig. 5-190, full curve); but extracts of suboesophageal ganglion have no such diuretic effect (Fig. 5-195, dotted curve). The seat of action of this DIURETIC HORMONE does not seem to have been established, but it acts on excretion rather than on water uptake. Probably it inhibits the reabsorption of water as it passes from the Malpighian tubules through the intestine. If so, a decrease in cell permeability in the intestine could be responsible, and would be comparable with that occurring in the skin of Carcinus, or in the distal kidney tubules of vertebrates, during diuresis. One curious feature is that the secretion of this metabolic 224 METABOLIC HORMONES 130 ied Operated with E saline e 1 ! e / / / Normal | control 90 With brain extract injected Fic. 5-19. Water uptake in larvae of Anisotarsus. a. The dotted line gives the mean weight (ordinates) in 3 days (abscissae) for five normal larvae, which remain constant in weight by diuresis. The full line gives the mean weight of five initially rather smaller, operated larvae (as Fig. 5-185) over the same period. Vertical lines show the range of weights between the extremes in each group. These do not overlap after the first day. b. Effect of injected extracts made at the times marked by arrows. The curves give weights of indi- vidual larvae, in which secretion of their own hormone was prevented by section of the circumoesophageal connectives. The full line shows an arrest in water uptake, 7.e. facilitation of diuresis, following injection of an extract of brain and corpora cardiaca in 1% NaCl. The dotted line shows continued water uptake after injection of a control extract of suboesophageal ganglia in 1% NaCl; salt solution alone gave a similar result (from Nufiez, 1956). § 5.321 WATER BALANCE 225 hormone is stimulated nervously from the ganglionic network round the gut; if this is damaged, or if the circumoesophageal connectives to the brain are cut, hormone secretion is not induced when the abdomen begins to swell. This nervous stimulation of hormone secretion recalls the activation of the pigment-controlling hormone of Carausius (§ 3.221), but is unusual for a metabolic hormone (§ 5.522), unless it be for the neurohypophysial hormones ($5:322): VERTEBRATA. HyYDROCORTISONE*, from the ADRENAL CORTEX (§ 2.31), has been found to have a diuretic effect on the kidneys, and also on the skin, of some vertebrates; but the evidence is most definite for amphibians and mammals (Table 25). AMPHIBIA. When amphibians enter their normal freshwater environment, excess water tends to enter through any per- meable tissues and to dilute the salt concentration in the blood. No observations on the action of hormones on the skin have so far been recorded in connection with limiting this endosmosis; it would clearly be of interest to know whether the relative imper- meability that is usual for the skin (controls, Fig. 5-20 a-c,) results merely from the lack of antidiuretic hormone, (§ 5.322), or whether a positive, pore-contracting action of an adrenocortical hormone may also be involved. The similarity in behaviour between isolated skin and that in whole animals would lend no support to the latter supposition, unless hydrocortisone persists for a long time in the tissues after isolation. The reactions are slightly better known in the kidney, where diuresis seems to be facilitated by HYDROCORTISONE. On exposure to a hypotonic medium, salts are actively reabsorbed in the proxi- mal tubules (§ 5.311). This would result, as in the mammalian kidney (Fig. 5-15a, p. 214) in an obligatory endosmosis of water. In so far as the inward diffusion of water failed to keep pace with the transport of the salts, “free water’ would remain in the tubules, converting the isosmotic glomerular filtrate into hypotonic urine. The greater the degree of impermeability to water possessed by the kidney tubules, the greater would be the hypotonicity of the urine. There is no clear evidence as to how this impermeability is * Several related 17-OH steroids have the same effect (Part IT). Q 226 METABOLIC HORMONES maintained in frogs, but it is probable that hydrocortisone may act here as in mammals. The removal of such hormones by adrenalectomy in Rana temporaria (but not in R. pipiens) is followed by oedema, or an excessive accumulation of water in the tissues; but the effect of injecting cortical extracts has not yet been shown (Chester Jones, 1957a). 30 20 Water uptake, pvcem*/ hr Controls Controls Controls Intact Intact Isolated cnn (a) (0) (c) Fic. 5-20. Increase in rate of water uptake following dehydration or injection of neurohypophysial extract (frog ADH); a. and Bb. in intact Rana pipiens, with the cloaca ligatured to prevent loss by excretion; c. in isolated frog’s skin (from Sawyer, 1956). TELEOSTEI. Although freshwater teleost fish are in much the same osmotic relation to the environment as Amphibia, the scales usually make the skin more waterproof, and water only enters the tissues through the gut and gills. Diuresis by the kidneys seems therefore to be the main means of water control, and there is some evidence that the cortical cells of the ANTERIOR INTERRENAL BODY (§ 2.311) may be concerned; but the situation is different in species adapted to different normal environments. In Fundulus, the cortical cells remain inactive when the fish are in sea water, § 5.321 WATER BALANCE 227 which is for them a hypertonic medium (cf. G. Pickford and Atz, 1957). The protective action claimed for the thyroid, in allowing fish to migrate into waters of low salinity, has not been fully elucidated (cf. § 5.311); but it may be similar to that of thyroxine, in causing water (and salts) to pass from the tissues of mammals to their blood, thereby increasing the possible rate of water diuresis by the kidneys (Fontaine, 1956). | The corpuscles of Stannius, derived from the pronephric ducts of teleosts, may also have a diuretic function, as they tend to hypertrophy when the tissues are loaded with water (Rasquin, 1956). MamMaAL_iaA. When water is plentiful in the tissues, the mam- malian kidney reacts like that of the frog and excretes a hypotonic urine. The volume of urine is considerably less than that of the glomerular filtrate, chiefly because reabsorption of salts by active transport in the proximal tubules results in a correlated reabsorp- tion of at least 4/5ths of the water by obligatory endosmosis, either in the same part or in the thin loop of Henle. Nevertheless, the diffusion rate for water limits the amount of endosmosis, so that, as in the frog, ‘“‘free water’ remains in the tubule. Most of the remaining sodium ions are actively reabsorbed, or exchanged for other cations, and especially for H+, in the distal tubule, which is made relatively impermeable by HYDROCORTISONE. ‘This causes further water to become “‘free’’ and to pass on to the collecting ducts. Thence it would be excreted, as it is in the frog, were it not for a ‘concentrating mechanism”, which is apparently independent of hormone control. This is of most significance in reinforcing antidiuresis, and it is referred to in more detail in that connexion (§ 5.322). “Osmotic diuresis” is also unaffected by hormones; it can occur if the blood is loaded with extra urea, which then passes into the glomerular filtrate and increases the tubular osmotic pressure. so that more water than usual is excreted, instead of being re- absorbed (Mudge, 1954). The action of hydrocortisone, in causing the relative imper- meability of the distal tubules in normal diuresis, has been examined indirectly in ‘‘water-loaded rats”, i.e. rats which have been given large doses of water by stomach tube. Such rats 228 METABOLIC HORMONES normally show a maximal urine flow, comparable to that of rats with ‘“‘diabetes insipidus’’, which follows inactivation, or removal, of the neurohypophysis and therefore of the store of the anti- diuretic hormone (§ 5.322). When adrenalectomized, the water- loaded rats, with or without their neurohypophyses, show less than 1/10th of the urine flow of either unoperated or merely neurohypophysectomized controls (Chester Jones, 19575; cf. Table 27). The decrease in diuresis in adrenalectomized rats (Fig. 5-16, p. 216) is greater than could be accounted for by the decrease of the blood pressure to a half, or even the correlated drop to 1/6th in G.F.R., the glomerular filtration rate. It may be assumed to be due to the lack of adrenocortical hormones. If rats are loaded with hypertonic saline, instead of water, the urine flow in the controls is halved, as compared with that in the rats with diabetes insipidus. This is mainly because the anti- diuretic hormone, ADH, is secreted in response to increased tissue salinity and higher osmotic pressure of the blood (§ 5.322). Adrenalectomy, by removing the source of the diuretic hormone, reduces urine flow still further, though not so greatly as in the case of water-loading. This is because the higher level of salts in the glomerular filtrate reduces the volume of obligatory endos- mosis. ‘he action appears to be independent of the presence or absence of the neurohypophysis (see Chester Jones, 1957a, for further details). The secretion of hydrocortisone, that causes relative tubule impermeability, is stimulated by adrenocorticotrophin, ACTH (§ 4.231). 5.322 Increase in cell permeability and antidiuresis leading to dilution of the blood-salts Dehydration can occur both on land and in any hypertonic aquatic environment. The animals’ response to this usually includes waterproofing some parts of the body surface to restrict water loss and increasing the permeability of other parts to allow of water uptake or reabsorption. INVERTEBRATES. No example of a hormone that increases tissue permeability to water has so far been found in the reports on 229 WATER BALANCE § 5.322 — _ _ — = = pezIuu0j}a[eUsIpe pue snpidisur sajaqriqg ———— pezrwoja[vusIpy snpidisur sojoqeiq, Ss[O1}UO-) e-1+ i 9006F © fb -1:60087 LL0°+ +000: + 6-SS | cLL-O L60:-0 | bLEe-0 9700-0 (|! C-I+ 910: + 7LO- + €S0: + $000: + C-7S £OT-O LOT-O OCE-0 L£00:0 O€ Lé+ | ETF | 600-= L€00-: + $700: + 0-96 18s-0 LéI-0 640-0 +£0-0 Ol LT 6s0-+~ |. 610-= S400: = +100: + 6:°S0T | 009-0 | CrIl-0 £S0-0 9¢0-0 6C Pi eo ae Se SS 9H WN NIW NIW NIW SNAWI0ddS aunssadd | /D OOL/IN /2 0O[/Oa-ONOIIN /® 0OL/1N dO aoold | Polen ee’ | M VN ANIA Yad WON “(QLS61 ‘souof 103s94D woz) jndino eulin pue “y"7'5) ‘ainsseid pooyq ur dorp ay} a31dsap Ausojoa[vusipe Aq pasvosour A]poyreu st yndjno wimrpos ayy, “BOAR JOBJINS WO (OT /,UID ¢ Jo adesop B 4k aqn} YoRUIO}s Aq 4a]D02 pay]4sip y110% papvo] 919M ITV ‘Pezruu0jaTeusrpe pue snpidisur sajoqeip yiIM sjer pue skep + TOF POZTWMOJzTeUsIpe Useq peYy yey} sjer ‘(stshydodAyoimou I19y}3 jo [BAOUaI J9je) snprdisur sajyoqeip YIM sie4 ‘(s[o1]U0_) s}er oyeUey pur ayeu yeuZIOU :1OF (10119 pivpurys | ) suvow se uMOYs 918 dInssoid pooyq pue (Y"Yq"D) 9381 Uoneayy Ie[N1awoys sy} ‘unisseyod pue umipos ‘sulin JO yndyno 94, J, AWOLOATYNAYGVY ONIMOTIOS SINAISSV.LOd GNV WOAIGOS ‘YaLYM JO NOILAYOXA NI SHONVHD °/Z aTav T, 230 METABOLIC HORMONES invertebrates, though it might perhaps be expected in land Crustacea. That claimed for the cockroach, Blabera, is an extract which has only been shown to act on rats (Stutinsky, 1953); that claimed for extracts ofthe post salivary glands of some Cephalopoda has likewise only been tested on unrelated animals (Erspamer and Boretti, 1950). VERTEBRATA. Increase in the water content of the tissues of vertebrates can be obtained either from the food in the gut, where no hormone action has been found, or from the environment, as when Amphibia move from dry to damp conditions, or back to water. An antidiuretic hormone from the neurohypophysis not only increases skin permeability, to facilitate this water uptake, but also favours water conservation by increasing its reabsorption from the urine in the kidneys and bladder. Its action at all three sites is to increase cell permeability and facilitate water endosmosis. Such antidiuretic hormones occur in most vertebrates and are closely akin to the so-called ‘‘vasopressin”’ of mammals. Since the main function, even of the latter, is not to control the tonus of blood vessels as its name implies, but to increase reabsorption of water from the urine in the kidneys and thereby reduce the urine flow, “‘antidiuretin’’* would seem to be a more descriptive name for this type of hormone, so often referred to as ADH. ADH is secreted from the hypothalamus and stored in the neurohypophysis (§ 2.111), whence it is released into the circulation (Table 25, p. 208). Increase in permeability of the skin AMPHIBIA. Since the skin of amphibians is moist and its per- meability to water can be varied, it plays as important a part in the control of the water balance in these animals as do the kidneys and the bladder. ‘There is no longer thought to be a “‘water balance principle’, distinct from the antidiuretic hormone and acting only on the skin of amphibians; but the amphibian ANTIDIURETIC HORMONE is chemically more akin to oxytocin than to mammalian ADH (= vasopressin). Asa rule, frogs do not drink an appreciable amount of water, unless they are immersed in a relatively saline * It should not be confused with ‘‘Antidiuretin’’, a commercial product with a similar action. & 5.322 WATER BALANCE 231 medium; but water losses following exposure to air can quickly be made good, on the animals’ return to water, by imbibition through the skin. This can be shown by ligaturing the cloaca of such de- hydrated frogs to stop loss of urine; then increase in weight over two or three hours from the time of their return to water will show the amount of water uptake. Comparison with control frogs, which had already been in water for some time, shows that the rate of water uptake after dehydration may be three times greater Wt. regained as % original wt. Hours in water Fic. 5-21 Rates at which normal and hypophysectomized frogs, Rana pipiens, return to their initial weight after being dehydrated and then placed in water at time 0. Hypophysectomy and the consequent lack of frog ADH reduces, but does not inhibit, water uptake through the skin (from Levinsky and Sawyer, 1953). than in the controls. In the latter, the loss of urine, if the cloaca were not ligatured, would balance this uptake and the weight would remain constant (Fig. 5-20a). Removal of the hypophysis decreases, but does not inhibit, the rate of water uptake by dehydrated frogs (Fig. 5-21), presumably because the osmotic gradient is steeper than in the normally hydrated controls. In- jection of a neurohypophysial extract (FROG ADH) more than restores the capacity for water uptake (Fig. 5-205). Even isolated skin treated with the same hormone shows an increase in water uptake large enough to account for the changes seen in whole frogs (Fig. 5-20c). 232, METABOLIC HORMONES Xenopus, the wholly aquatic clawed toad, shows little increase in water uptake in response to ADH injection; but the terres- trial toad, Bufo americana, (though not Bufo bufo; Sawyer, 1956) shows a greater increase than the frog. Moreover, the toad is able ) fole) 200 300 400 Milliosmols NaCl / litre, in bath Fic. 5-22. Percentage change in weight of the frog, Rana, after immersion for 3 hr each time, in baths of different osmotic concentration. Increase and decrease in weight, due to change in water content, are shown as ordinates above and below the hori- zontal line; values for osmotic concentration as abscissae range from distilled water, O, to values above that of plasma (ca. 325 milliosmols). Control frogs maintain a constant value over much of this range; but frogs that had received 5 units/100 g body weight of NEUROHYPOPHYSIAL EXTRACT (frog ADH) allow a free flow of water through their skin, so that their weight changes steadily until they come each time into osmotic equilibrium with the external medium (from Sawyer, 1951). to take up water from damp moss and does not need to be sub- merged like the frog. There therefore seems to be an adaptive correlation between the sensitivity of the skin to the hormone and the degree of adaptation to land life. The neurosecretory material in the neurohypophysis is found to be depleted in dehydrated frogs, indicating that it has been secreted in response to the need to absorb water. 5.322 WATER BALANCE 233 Experimentally the skin of a frog or toad behaves as if it had ultra-microscopic pores that are enlarged by the neurohypophysial hormone to allow an osmotic flow of water, instead of the slower diffusion (Ussing, 1954). The rate of flow is then determined by the osmotic gradient (Fig. 5-22). Even with 1/10th Ringer as the “outside medium”’ for isolated skin, the net influx of water is several times greater than it could be by diffusion. In fresh water the rate would be still greater. Ussing supported his “pore theory” by putting thiourea on both sides of the isolated skin; but the thiourea on one side had its carbon, and on the other its sulphur, isotopically labelled, so that the flux could be followed in both directions. When neurohypo- physial hormone was present, the increased flux of thiourea was much greater than the increase in flux of the water. It may be assumed that the hormone dilates pores that are normally too small for molecules of thiourea, though just large enough for water. Moreover, ‘‘the influx of thiourea is more rapid than the outflux, even when the concentration is the same on both sides’’, and this reflects the fact that thiourea is swept through the pores in the osmotic flow of water (Sawyer, 1957). Increase in permeability of the bladder AMPHIBIA. Reabsorption of water from the bladder in Rana is similar to that from the skin and kidney tubules and responds in like manner to frog ADH injections and to dehydration, a state which presumably stimulates the secretion of the frog’s own neurohypophysial hormone (Fig. 5-24). Mamma.ia. There is no evidence of water being reabsorbed from the bladder in mammals. Increase in permeability of the kidney tubules Ampuisia. When amphibians are exposed to dehydrating conditions, conservation of water in the kidneys is stimulated by the independent action of ANTIDIURETIN at two points, the glo- merulus and the tubule. The glomerular filtration rate (G.F.R.) is determined by the arterial blood pressure, the resulting filtrate being isosmotic with the blood plasma and only differing from it in the lack of proteins 234 _ METABOLIC HORMONES or other colloids. ‘The top curve (Cer) in Fig. 5-23 shows the effect of the pressor action of frog ADH in reducing the volume of the filtrate by constricting the blood flow to the glomerulus. It can also reduce the number of glomeruli which are active. After injection of antidiuretin the kidney tubules, therefore, receive a reduced volume of isosmotic filtrate, containing salts, sugar and urea. Rona catesbicna 500g 1/100 frog ADH | (0-007 Oxytocic unit / kg) 100 Co R cmY kg / hr Hours Fic. 5-23. Responses of the bullfrog, Rana catesbiana, during 2 hr after an injection of the ANTIDIURETIC HORMONE from the NEUROHYPOPHYSIS. Frog ADH reduces G.F.R., the filtration rate (shown by creatinine clearance, “cp, on ordinate scale on left) but not sufficiently to account for the large reduction in volume (V on same scale) of excreted urine. This is chiefly due to the reduc- EP aE ’ C : tion in the ratio (shown as = on percentage scale on right) of CR clearance of ‘‘free water” to that of creatinine, due to reabsorption of water in the kidney tubules. Control injections caused no signifi- cant changes (from Sawyer, 1957). So.o22: WATER BALANCE 235 In the tubules all the sugar is reabsorbed, but the urea, to which the tubules are impermeable, is excreted. Monovalent salts, which account for much of the osmotic pressure of the filtrate, are less actively reabsorbed in the tubules during antidiuresis than during diuresis (§ 5.312 and Fig. 5-15), p. 214). The action of ADH on the tubules is to increase their permea- bility, presumably by increase in pore size, as in the skin. The obligatory endosmotic flow of water then follows closely on the active inward movement of salts, so that the final urine tends to be isosmotic with the plasma. The reduction in clearance of ‘free water” is shown in the lowest curve (Fig. 5-23) to be much greater than could be accounted for by the reduction in glomerular filtration rate alone, as shown by the creatinine clearance. It is therefore assumed that the reduction in volume of urine (V, Fig. 5-23) is mainly due to reabsorption of water because of the increase in tubule permeability, and not only to the reduced G.F.R. No Amphibia have been found to excrete urine more con- centrated than that which is isosmotic with the plasma. There is, therefore, no need to postulate for them any seat of active transport of water from the kidneys to the tissues. There is considerable specific variation among Amphibia in their sensitivity to their own ADH. Removal of the neurohypo- physis, and therefore of the store of this hormone, has no effect in Bufo bufo and Rana temporaria, as compared with intact controls. Bufo arenarum, when kept out of water, behaves like R. catesbiana (Fig. 5-23). Bufo marinus of South America may be even more sensitive than these to frog ADH; in hydrated and hypophysec- tomized specimens only about 35 per cent of the filtered water was reabsorbed by the tubules; but after injecting extracts containing frog ADH this rose to an average of 65 per cent (Sawyer, 1956). The rate of secretion of antidiuretin from the neurohypophysis in Amphibia may well be mediated through osmo-receptors in contact with some of the main arteries, as it is in mammals (Chester Jones, 1957a). TELEOSTEI. In sea water the body fluids of teleosts are maintained at a level considerably hypotonic to the medium by drinking sea water and excreting the excess salts through the gills. Kidney excretion is reduced to a minimum, largely by a loss of glomeruli, 236 METABOLIC HORMONES though the action of an antidiuretic hormone might also be expected. Although extracts of the fish neurohypophysis have a potent antidiuretic effect on frogs, there is no direct evidence of the hormone having the same action on the fish themselves. The best indirect evidence for the relation of hormones to an anti- body wt % Injection Bladder content, O 3 5 Time; > hr Fic. 5-24. Effects of injection of NEUROHYPOPHYSIAL EXTRACT (Frog ADH), at the time marked by the arrow, on the water content of the frog, Rana pipiens. ©: values calculated from changes in body weight; @: values for direct measurement of bladder contents. The cloaca was ligatured and injected with phenol red to show if any leakage occurred; the frogs then sat in water, which was imbibed through the skin and excreted into the bladder, for 3 hr; then the controls (dotted line) continued to excrete for the next 2 hr while the specimens, which received the injection at hour 3 (full line), showed a marked decrease in bladder water. This was taken to mean that water had been reabsorbed into the tissues from either the bladder or the cloaca (from Sawyer, 1956). diuretic function in Callionymus and Ammodytes (Arvy, 1957; Arvy, Fontaine and Gabe, 1954) is the observation that on transfer from normal sea water to hypertonic sea water, the increased dehydrating action of the environment is accompanied by depletion of the neurosecretory store in the hypothalamus and NEURO- HYPOPHYsIS. ‘he secretion is re-formed when the fish are restored to normal sea water and increased if they are placed for a short 89.522 WATER BALANCE 237 time in hypotonic sea water, where the tissues would tend to become over-hydrated and there would be no call for the secretion of an antidiuretic hormone. Presumably in normal sea water the supply of the hormone is in balance with the demand and no histological change is observable. ReprTILia. Little seems to be known of water control in reptiles. Chester Jones (1957a) writes: ‘‘Many reptiles are adapted to arid conditions and even those whose habitat is in or near water have no movement of water through the skin . . . nor is the reptilian renal tubule specifically adapted for water reabsorption beyond the plasma osmotic concentration. The alligator is very responsive to the antidiuretic effect of . . . pitressin [mammalian ADH] rather than the pitocin fraction, as in frogs. Antidiuresis is effected, at least partly, by lowering the glomerular filtration rate, pitressin [contracting] the smooth muscle of the afferent glomerular arteri- ole.” Metabolic waste products and especially the uric acid are eliminated not only by glomerular filtration, but probably to a much greater extent by tubular secretion. Aves. Even less seems to be known of birds, but they are prob- ably similar to reptiles. Mamma lia. The hormone control of water balance in mammals has been most fully investigated in rats, but seems to be basically similar in other forms, including the dog. In one way the situation is simpler than in the frog because the skin is practically waterproof, and the supply of water is, therefore, derived entirely from the gut contents by intestinal absorption, where no hormonal influence has been detected. The loss of water from the sweat glands does not appear as a rule to be under hormone control either (§ 4.14); nor does the loss through the lungs. The main control of the water balance occurs in the kidneys, but it is only partially dependent on hormones, because mammals have, in addition to the systems found in the frog, the “concentrating mechanism’’ already mentioned (§ 5.321). If the ANTIDIURETIC HORMONE, ADH, were to have the same effect in mammals as in the frog it would increase the permeability of the distal kidney tubules and allow of water reabsorption there until the urine became isosmotic with the plasma (Fig. 5-156, p. 214). It is clearly of importance for wholly terrestrial mammals 238 METABOLIC HORMONES to be able to conserve more water than this, and in fact it is commonly found that the urine is markedly hypertonic. Recent evidence shows that in addition to the action of an antidiuretic hormone (ADH injected as vasopressin in graded doses in Fig. 5-25) there is a process of active reabsorption of water. Although the exact means by which it is brought about is still under dis- cussion (e.g. Wirz, 1957 and Sawyer, 1957), it is clear that it is Urine cm3/10O min Time, JOmin intervals Fic. 5-25. Urine output (ordinates) of an unanaesthetized rat, Rattus, after being given a standard water load by stomach pump (5 % body weight, or ca. 11-3g) and then a dose of ANTIDIURETIC HOR- MONE (vasopressin). Successive doses were given intravenously, in the order b, c, d, a, e, at the times marked by arrows and were graded: a = 100, b = 200, c = 400, d = 600, e = 800 v units. The antidiuretic effect, shown by the drop in urine output, approaches a maximum after dose d, and the further increase in hormone given at e has practically no greater effect. The method can be adapted for assaying antidiuretic substances over the lower range of concentrations (from Ginsburg and Heller, 1953). independent of hormone control. This has been demonstrated in water-loaded dogs, in which urine flow is maximal and antidiuretin, ADH, is therefore not being secreted (Berliner and Davidson, 1957). Acute unilateral reduction of G.F.R., induced by occluding the blood supply to one kidney only, so reduces the volume of the filtrate reaching the “‘concentrating mechanism” of that side, that the urine collected separately from the two ureters becomes $5,322 WATER BALANCE 239 hypertonic on the treated side, while remaining iso- or even hypotonic on the other side. This concentration may or may not be limited to the collecting ducts rather than occurring in the renal tubules (Fig. 5-15). It is certainly limited to a maximal rate of water uptake, no matter what volume of fluid is delivered to it from the tubules; it is also limited to a maximal osmotic concentration of the urine, relative to, but con- siderably above, that of the plasma. “The degree of urinary concen- tration in the mammal is determined . . . largely by the volume of water delivered to this concentrating mechanism ”’ (Sawyer, 1957). The natural secretion of the antidiuretic hormone can be brought about by stimulation of osmo-receptors in the anterior hypo- thalamus of the brain, a region supplied by branches from the internal carotid arteries. It has been shown in dogs that infusing these branches of the carotids for about half an hour with suffi- cient saline to increase the osmotic pressure of the blood by some 2 per cent can reduce the urine flow by 90 per cent. This treatment is much more effective than giving similar saline infusions else- where in the circulation (Jewell and Verney, 1957). The presence of the saline presumably has the same effect as tissue dehydration ; yet it can be shown that although ADH injection into an isolated heart-lung preparation causes antidiuresis it does not reduce the G.F.R., whereas natural dehydration does both. The factor controlling G.F.R. in mammals is not known, since recent work all points to the vascular constrictor, ‘“vasopressin’’, being one and the same hormone as ADH. Haemorrhage also stimulates secretion of ADH, possibly by affecting vagal nerve endings sensitive to hydrostatic pressure in the heart or the great veins (Heller, 1956). Sawyer (1957) wrote: ‘‘Diuresis and antidiuresis can . . . be ex- plained in the mammal simply in terms of an increase in perme- ability of the distal segment to water under the influence of the antidiuretic hormone. This increase in permeability, since it involves reabsorption of water only up to the isosmotic state, could be interpreted in terms of a change in pore size, as the evidence indicates to be the case in the [frog] skin, and as we have inferred to be the case in the frog bladder” and kidney. But this statement was made prior to the publication of Chester Jones's 240 METABOLIC HORMONES evidence, referred to above, showing the importance of cortical hormones in causing water diuresis. 5.4 BALANCE OF CALCIUM AND PHOSPHATES Unlike water and the monovalent electrolytes, calctum and phosphates are of more importance in growth than in the direct relation of the animal to its environment. The changes in their concentration in the blood are, therefore, slower and less spectacu- lar, and fewer observations seem to have been made upon them. ARTHROPODA. Despite the differences in the metabolic signifi- cance of the control of calcium and phosphates, as compared with the salts previously considered, the hormones concerned in their control seem, nevertheless, to be derived from the same organs, namely the Y-ORGAN and EYESTALK in the Crustacea (p. 222) but not in the Insecta. In both classes, the changes in calctum and phosphates are associated with moulting, since in these forms with hard exoskeletons it is necessary for one or both salts to be with- drawn from the old skin before it is shed. During this phase they appear in the blood and rise to a maximum at the time of the moult, and then decrease suddenly as they are re-incorporated into the new shell, together maybe with fresh calcium rapidly absorbed from outside sources. The changes are more marked in Crustacea, which have heavily calcified shells, than in Insecta, where such structures are unknown and would clearly be too heavy to permit flight in air. VERTEBRATA. Since vertebrate growth is a continuous process and not as a rule accompanied by moulting, the changes in calcium and phosphates are more gradual; but the accumulation of these salts is an essential preparation for their deposition in bone in the later stages of growth. For convenience in recording what evidence has so far been put forward on the action of hormones on these salts, their concentra- tion in the blood will be used as the indicator. 5.41 BALANCE OF CALCIUM 5.411 Increase of calcium in the blood Crustacea. In Carcinus, a single injection of an extract of the Y-ORGAN, that contains the MOULT-PROMOTING HORMONE, MPH, BALANCE OF CALCIUM AND PHOSPHATES 241 § 5.41 SNISNDADY) epodesaq SNISNDAD’) snaynuvg SNIDISFy snuizav‘y DIT NVXa puv[s d1IOvIOYIOIg é pues snurg pues uvs1IO-KX-OTUOT[SUPLy eyel[e e1od10 100, 101, 102, 108ff. sources of hormones in, 18, 22, 23ff., 24, 26, 27 Cuticle formation, 117 Cuticle-secreting glands, 117, 129 Cyclostomes, 36 (see also Agnatha) Cystic duct, 63 Cytochrome c system, 182 Dark-adapting hormones (see Reti- nal) Death, from adrenalectomy, 14, 15, 216 from hypophysectomy, 14, 64ff. from low blood-Ca, 245 from muscular tetany, following parathroidectomy, 244, 246 from post-operative shock, 14 from stress, 145 Decapoda (Crustacea), 24, 25, 26, 29, 88, 94ff., 96, 98, 108ff., 169 Definitions of hormones, 5ff. Dehydrating conditions, 233, 236 Dehydration, of tissues, 207, 228ff., 231, 231;233 and urine flow in Mammalia, 239 289 Desiccation, 37 Deuterocerebrum, 24, 25, 30 Diabetes, insipidus, 217, 228, 229 mellitus, 1, 143, 195 Diabetogenic hormones, 189ff., 190 (and see Adrenal cortex and Glucagon) endocrinokinetic control of, 142, 144ff., 255 in Crustacea, 189ff., 192, 257 in Insecta, 173, 191 in Vertebrata, 144, 191ff., 253ff., 290 nervous control of, 191, 195, 254 sources of, 46, 50, 51, 52, 189ff., 190, 197,253 Diapause, 182ff., 259 effect of ecdysone on, 136, 137, 184 linked in host and parasite, 185ff. Diapause hormone, D, 31, 136, 182if.,.254, 257 Digestive, enzymes, 44, 115, 116, 1241f., 125, 127 glands, 117ff., 118 system (see Gut) Diiodotyrosine, face 48, 49, 141 Dipnoi (lung fish), 50 Diptera (flies), 135ff., 168ff., 169 (and se Calliphora) Diuresis, phosphate, 251 water, 143, 201, 207, 208, 214, ZNO. 2245 225. 229,235, 240, 257 Diuretic, extracts of cortex, 226 hormone, 201, 216, 220ff., 222, 223, 225ff., .254,..255. (see also Hydrocortisone and Si- nus gland hormone) Diurnal changes in activity of Pert- planeta, 69, 153 Diurnal rhythm, in adrenal cortex, 144 in calcium metabolism, 242 in chromatophores, 85, 86, 103 290 Diurnal rhythm, in epidermal cells, 76 in hormone secretion, 243, 259 in retinal cells, 80 Duocrinin, 46, 118, 120, 126, 127, 156 Duodenum, 46, 63, 120, 121, 124, 425,126; 427 156 | Duodenal glands, 118 Duodenal mucosa, 63, 124, 125, 126 Ecdysone (see Moult-promoting hormone of Insecta) Ectoderm (see Endocrine glands from) Ectohormones, 3 Eggs, growth of, 138 Eggshell and Ca, 246 Elasmobranchii (dogfish and skates), and chromatophores, 83, 105f7., 108, 109, 110 and control of glands, 122, 124, 139 and metabolism, 174, 197 and muscles, 68 sources of hormones in, 36, 47, BOE. ore Electrolyte balance, 8, 10, 143, 206 (see also Ions and Salts) Ca and phosphates, 240ff., 247, 242, 245, 247, 248, 254, 255 monovalent, 208, 209ff., 211, 213, 214, 216, 217, 218ff., 240 Electron, micrographs, between 22 and 23 microscope, 20 Electrophoresis, paper, 89, 109 Embryos, 2, 3, 4, 35, 48, 64, 151, 182, 184 Emergency reactions, 63, 67, 190 Endocrine glands or organs se- creting kinetic and metabolic hormones, 8, 61, 184 cells of, face 22, 32, face 48, 53, face 140, face 141 SUBJECT INDEX direct control of, 253ff., 254, 257 ectodermal, 9, 18ff., 22, 30, 38ff., 40, 48, 56, 132, 133, 134f. endocrinokinetic control of, 14, 15, 31; °915; 1318133 ee 140, face 141, 145, 146, 148, 258 endodermal, 18, 46, 46ff., face 48, 56, 132, 133, 139ff. location of, 119 mesodermal, 9, face 48, 50ff., 51, 53, 133, 143ff. (and see 22, 46, 51 for names, under which further refer- ences to particular glands are given) Endocrine glands secreting mor- phogenetic hormones, 8 antennary, 12, 134ff. gonadial, 70, 128, 133, 148ff. maxillary, 12, 134ff. (and see Peritracheal, Prothor- acic, ‘"hymus, Vas deferens and Ventral glands, and Y-organ) Endocrine secretory cells, face 22, 22, face 48 (and see Neurosecre- tory cells, and cells of Adenohy- pophysis, Adrenal cortex, Ad- renal medulla, Corpus allatum, Corpus cardiacum, Gut mu- cosa, Islets of Langerhans, and of Parathyroid, Salivary, and Thyroid glands) Endocrinokinetic hormones, 7, 9ff., 11, 12. 14,115,113 ae 133 actions in Arthropoda, 134ff., 158ff. actions in Vertebrata, 139ff., face 140, 143ff., 145, 146, 148 and growth, 134, 142, 256 and kinetic hormones, 128, 149, 187;:162 SUBJECT INDEX Endocrinokinetic hormones, and metabolic hormones, 167, 254,255, 2588. and metabolism in Arthropoda, 1737191195 , 205, 243, 258 (see also Hanstré6m’s sensory pore organ and Prothoraco- trophin) and metabolism in Vertebrata, 7 29 195, 205, 20141., 243, 252, 256, 259 (see also ACTH, STH and 'TSH) and morphogenetic hormones, 12, 134, 148ff., 151ff. (see also ICSH and LSH) control of secretion of, 145ff., 149ff., 157, 158ff sources of, 10, 31, 40, 44, 160 Endoderm (see Endocrine glands from) Endosmosis, 206, 207, 221, 225, 230 - obligatory, 227, 228, 235 Endostyle, 47, 173, 174 *“Energetic hormones’’, 7 Enterocrinin, 46, 118, 120, 126, 1275156 Enterogastrone, 46, 58, 64, 117, 116, 120, 126; 127, 153, 156 Environment (habitat), adaptation to, 7, 9, 85, 149, 151, 206ff., 240, 259 (see also Background) and diapause, 182ff. aquatic, 225, 228 arid, 237 control of, 86 damp, 222, 230, 232 dehydrating, 236 terrestrial, 228, 232 Enzymes, and secretagogues, 5, 117 and thyroxine, 140, 177 hormone-destroying, 62 intracellular, 196 291 secretion of digestive, 49, 115, 116, 128, 121, 1246., 125, 127 Ephemeroptera (Mayflies), 30, 133, 135 Epinephrine, 148 (and see Adrena- line) Epiphysis in Phoxinus, 94 Epistellar body, 21ff., face 22, 22, 23, 205/90; 09, 156 Epithelium as hormone source, 22 coelomic, 38, 50ff., 517, 52 ectodermal, 18, 22, 38 gut, 46 stomodaeal, 18, 39, 40, 48 thyroid, 48, face 140, face 141 Erythrophores (see Chromato- phores, red pigment in) Eupagurus, chromactivating hor- mones, EDH and ELH, 84, 87,93, 98 Excitement, response to, face 62, 67, 191 Excretion, 208 of calcium or phosphates, 241, 247, 249, 251ff. of salts, 213, 214, 216, 218ff., 220-0258 of sodium ions, 217, 218, 219, 229 of urea, 227 of urine, 214, 216, 227, 229, 235 of water, 207, 223 (see also Diuresis and Water) Exocrine glands, 9, 66, 115ff., 118, 129 Brunner’s duodenal, 126, 127 buccal, of Gastropoda, 116 cuticle-forming, 117, 129 digestive, 5, 117ff., 118 gastric (stomach), 119ff., 126ff. in Arthropoda, 116, 117 nasal, 218 under hormone control, 44, 45, 115, 1183129) 152k 292 Exocrine glands (and see Duoden- um, Intestine, Mammary, Ovi- ducal, Pancreas, Skin and Stomach glands) Exocrine secretory cells, 49, 118, £19 129 ASS Exoskeleton, 240 Experimental investigations examples of, 12ff., 14, 168ff., 183 pharmacological methods in, 13, Gie 86;°88; 103; 1407.5 13 0, 219, 238 physiological methods in, 13, face 62, 85, 103, 119 use of controls in, 12, 168, 171, 172,210, 216, 217,226,229, 232, 236, 242, 245, 250 Extracts, commercial (see Antuitrin, Pito- cin, Pitressin and Pituitrin) compared in Crustacea and In- secta, 29 fractionation of, 89 of hormone sources (see Adreno- cortical, Brain, Corpus car- diacum, Eyestalk, Neuro- hypophysis, Parathyroid glands, Pericardial organs, Sinus gland, Suboesophag- eal ganglion, Thyroid gland and Y-organ) relation to hormones, 28, 35, 86, 94, 119 Upjohn’s cortical, 185 uses of, 13ff., 14, 51 Eyes, adaptation to light, 79, 154 appositional, 77 compound, 74, 77ff., 78, 81 illumination of, face 83, 100ff., 101, 105, 106, 149 retinal cells of, 71 sessile, 100 stalked (see Eyestalk) superpositional, 77 SUBJECT INDEX Eyestalk, as source of kinetic hormones, 2, 58, 71, 108 (and see Sinus gland) as source of metabolic hormones, 178, 190, 205, 240, 248, 248 extracts of, 78, 79, 88, 180, 191 removal of, 178, 179, 180, 181, 187ff.; 189.) 200: 204 (see also Eyestalkless speci- mens) structure of, 24, 25ff., 26, 28 Eyestalk hormones, 2 (and _ see Sinus gland and HSPO) calcitum-decreasing, 241, 247 diuretic, 201, 202, 221ff. kinetic, 69, 71, 81, 153 metabolic, 178, 180, 188, 190, 199ff., 204, 248 moult-inhibiting (see Moult-in- hibiting) source of, 25ff. Eyestalk tip, 22, 190, 205, 252 Eyestalkless test specimens, and chromatophores, face 82, face 83, 88, 89, 90ff., 90, 92, 96, 971K, 9S and metabolism, 168, 178, 180, 192, 202, 220, 248, 249, 251 Eyestumps, cautery of, 168 Fallopian tube, 65 Fat, and flow of hormones, 63, 126 metabolism of, 168, 169, 186ff., 188, 200 Fat-preserving hormone, 188 Fat translocation, 186, 255 Fatty acids, 63 ‘‘Feed-back”’ control, in Vertebrata, 11, 134, 141, 146, 149, 150, 252 Fish, and chromatophores, 73, be- tween 82 and 83, 106, 108 SUBJECT INDEX Fish, — - and control of endocrine glands, 133, 139, 144, 256 and metabolism, 174, 175, 176, 193° 197, ‘212,226; 2438,, 250 sources of hormones in, 33, 35, 36, 37, 38, 40, 41, 43ff., 49, 50 (see also Elasmobranchii, Holo- stei and 'Teleostei and “‘fish”’ in Animal Index) Flagella, 10, 167 Follicle-stimulating hormone, FSH, 8, 10 actions of, 133, 150ff., 161 source of, 44 “Free water” in kidney tubules, 214,225,227, 235 Fresh water, transfer to and from, 206, 207, 212, 226, 244 Frontal ganglion, 30 Frontal organ of Apterygota, 31 Fructose, 198 Galactose, 193, 196, 197, 198 Gall bladder, 58, 63, 120, 127 Gametes, 8, 152 Gamones, 3 Ganglion, abdominal, 29, 30, 91, 92 frontal, 30 hypocerebral, 30, 33, 39 optic, 81 (see also Optic lobe and Ganglionic-X-organ) parasympathetic, 45 pedal, 23 stellate, 21, 22, 23 suboesophageal, q.v. sympathetic, 33, 37, 68 thoracic, 24, 29, 30 ventral, 24, 30 ventricular, 30 Ganglion cells, 27, 37 293 Ganglionic-X-organ, as source of hormones, 80, 82, 169, 178, 179, 188, 191, 208, 241, 247ff., 251 structure of, 22, between 22 and 23, 25ff., 24, 26, 27, 28 Gastrin,; 5,6; 156 kinetic actions of, 57, 58, 64, 118, 119.) 1208 .; 120, +121;.122, 124, 126ff., 152, 153 source of, 45, 46 Gastro-intestinal hormones, 45, 46 Gastroliths, calcareous, in Crusta- cea, 242 Gastropoda (univalve molluscs), 116, 128, 129, 163 Genes, 3, 4 Genital (reproductive) ducts, 12, 64ff., 115, 149, 150, 151 Gestation, 152 (see also Pregnancy) Gills, 226, 235 **Glandotrope Wirkung’’, 131fn.(see also Endocrinokinetic action) Glands (see Endocrine and Exo- crine Glands) Glomerular filtrate, 215, 225, 227 228 2330 238 25% reduced volume of, 234 Glomerular filtration rate, G.F.R., 228. 229. 233.2304, 2393¢257, 239 unilateral reduction in, 238 Glomerulus, 214, 233ff., 235 Glucagon, control by: STH, 133, 142ff., 193ff., 194, 199, 255, 256 diabetogenic action of, 190, 193ff., 197, 252, 253, 256 secretion stimulated by low blood-sugars, 195, 254, 256 source of, 46, face 48, 50 “Glucagonotrophin”’, 142fn. (and see Growth hormone) Glucocorticoid hormones, 144 (and see Hydrocortisone) 294 Gluconeogenesis, 195 Glucose, 6, 11, 191,193, 194, 195ff., 198, 256 Glucose-6-phosphate, 196 Glycogen, 188, 193, 194, 195, 196, 200 Glycoproteins, 43, 173, 174 Gonads, 4, 8, 10, 18, 128, 129, 132, 133148. 1597 Gonadial hormones, 70, 128, 133, 148ff., 151 - Gonadotrophic hormones, 148ff., 157 (see also ICSH and LSH) Gonadotrophins, 10, 148, 161 Graded responses, 154, 238 Granules, pigment (see Pigment) secretory (see Neurosecretory) Growth, and Ca and P, 240 and metabolism, 253 gradients, 3 hormone, 8, 11, 194, 195 (see also Somatotrophin) hormone in plants, 3 Guanophores (see Chromatophores, white pigment in) Gut, absorptive cells of, 167 action of chemicals in, 123, 124, 1252255 argentaffine cells of, 45 control of digestion in, 117, 118 ADT glands of (see Endocrine and Exocrine glands) mechanical distention of, 5, 119, Waa DE mid-, crypts of, 117 mucosa, isolated hormone- secreting cells in, 3, 5, 6, 9, 4467... 46,417, 1275 155, 156, 257 ____muscle (see Muscles) 194, 195, 145: SUBJECT INDEX water absorption by, 226, 230, 237 Haemolymph, carrying hormones, 136, 137 3 Haemorrhage, 11, 145, 239 non-fatal, 145, 145 Hair follicle muscles, 67 Hanstréms sensory pore organ, as source of hormone, 133, 134ff., 243, 254, 258 structure of, 22, 24, 26, 28, 31 Heart-accelerator hormone, 38, 59ff., 154, 156 (see also Ad- renaline) Heart beat, 58, 59, 60, 62, 67 inhibition of, 59 Heart muscle, 57ff., 63 Heat, output, 177 regulation, 186 Henle, loop of, 214, 227 Hepatopancreas, phosphate con- tent of, 249 Herring bodies, 32, 34, 35 Hexokinase, 196 Hibernation, 186 Hirudinea (leeches), 83 Histamine, 5, 59; 120, 122, 1245 145ff., 145, 146, 148, 757 Histology, 12, 19 Holocrine secretion, 116 Holostei (e.g. Amaia), 36, 52 Hormones, actions ,,of, 77,8, Sof. 115i, 167ff. (see also specific effec- tors and reactions) antagonistic reactions of, 10, 84, 86, 91ff., 103. Isao. 205, 207, 219, 245, 252,256 (and see below, pairs of hormones) breakdown in tissues, 59, 90, 91, 140, 155, 184 SUBJECT INDEX Hormones, chain reactions of, 9, 11, 14, 181 (see also Endocrinokinetic hormones) characteristics of, 6, 152ff., 252ff. classification of, 2, 7 concentration of, in blood, 59, 90, 99, 109, 141, 145f., 160 (and see below, dosage) definitions of, 5ff. ' direct control of, 120, 155, 253ff. discovery of, 1 dosage, effects of, 59, 63, 67, face 68, 92, 154, 238 endocrinokinetic, q.v. “energetic”, 7 identification of, 12ff. in balance with demand, 237 inhibitory, 6, 126ff., 154, 178ff., 184 (and see MIH) in host and parasite, 185 in tissues, 91, 140, 184 - “Gntracellular”’, 3 kinetic, g.v. metabolic, q.v. morphogenetic, q.v. nervous control of, 9, 38, 156, U58it.; 191, 205, '2545 257f. pairs of, maintaining balanced reactions, 10, 84, 92, 107, 153, 189; 2078., 2142 19ir., 239ff., 245, 252, 256 plant, 2,3 rate of destruction of, 155 release of, 23, 258 (and see Storage-and-release organs) sources of, 7, 13, 14, 18ff. stimulation of secretion of, 46, Holt. 1 slits Psott., 196,497, 253th 294, 299, Lot. synergistic actions of, 150, 252, 256 tabulation of, 57 (and see List of Tables, p. vii) types of, Off. 295 vascular, 3, 4, 6, 8, 12, 19 et seq. Hydrochloric acid, and flow of hormones, 123, 123, 124, 125, 1275253 Hydrochloric acid secretion, 118, 1495-120, 127, 153 inhibition by enterogastrone, 126 stimulation by gastrin, 119, 122, 127, 106 Hydrocortisone, and metabolism, 169, 190, 194ff., 205, 208, 214, 216;,-225i: stimulated by ACTH, 133, 228, Zao Hydrocortisone-like hormones, 51, 143f., 205;°225' faz Hydrogen ions, Ht (see Ions) Hydrostatic pressure, affecting vag- us, 239 5-Hydroxytryptamine, 116 Hyperglycaemia, 142, 143, 191, 195, 197 Hypertonic, media, 37, 207, 218, 221.256 urine, 214, 239 Hypocerebral ganglion, 30, 33, 39 Hypoglycaemia, 142, 193 Hypophysectomy, or removal of hypophysis, and ACTH, 1/4, 147ff., 212 and ADM: 67, 2315235 and Ca metabolism, 243, 245, 246 and melanophores, 103, 104 and oxytocin, 64, 67 and TSH, 140, 141 Hypophysial, arteries, 41, 42 hormones, 142, 152, 245 (and see Adenohypophysis and Neurohypophysial hormo- nes) portal system, 41, 42 Hypophysis, 40, 48, 64, 103, 104 (and see Adenohypophysis and Neurohypophysis) and balance of salts, 212ff. 296 Hypophysis, and calcium control, 243ff. and phosphate control, 256ff. functions altered by transplanta- tion, 141, 161ff. Hypothalamus, and pituitary for- mation, 40, 48 as source of neurosecretions, 22, 33ff., 34, 42, 64, 236 control of adenohypophysis by, 141, 147, 149 injury to, 64ff., 244 neurosecretory cells of, 33, 35 osmoreceptors in, 235, 239, 257 Hypotonic, media, 206, 225, 237 tissues, 235 urine, 214, 225, 227, 239 Infundibulum, 35, 40ff., 48 Inhibition of reactions, by brain, 183 by brain extract, 59 by hormones, 6, 58, 64, 126, 153, 178ff., 180, 184, 200, 203, 204, 223 by nerves, 154, 191, 257 Inhibition of pigment granule con- centration, 74 Insecta, 2, 11, 257, 258 (see also Apterygota, Diptera, Ephe- meroptera, Lepidoptera, Odo- nata, Phasmida, Plecoptera and Syrphidae) and control of kinetic hormones, 156, 160 and control of metabolic hor- mones, 254, 255, 257, 258 and diapause, 136, 137, 169, 182ff. and glands, 129, 135ff. and metabolism, 186ff., 190, 191, 195, 203ff., 222ff., 223, 224, 240, 243, 250, 251 and muscles, 56, 58, 59ff., 62ff., 69 SUBJECT INDEX and pigmentary effectors, 71, 74ff., 75, 76, 82, 85, 87, 102 and respiration, 168ff., 169, 171, 17Z sources of hormones in, 18ff., 22, between 22 and 23, face 23, 29ff., 30; .32,.38, 398., 62 Insulin (antidiabetogenic hormone) 16282155253 action of, 190, 194, 195ff., 196, 197, 198 control of secretion of, 6, 198, 253, 254 : secretion and STH, 199, 256 source of, 46, face 48, 49 Intercerebrum (pars intercerebra- lis) of Arthropoda, 8, 62, 133, 135ff., 195 (see also Neuro- secretory cells of) Intermedin, 44, 87, 94, 104, 107, 160 (see also MSH) Internal ‘‘milieu”’ in the tissues, 259 Interrenal body, 38, 50, 51, 51 (and see Corpuscles of Stannius) Interrenal tissue, anterior, 51, 5/, 144, 212, 226 Interstitial-cell-stimulating —hor- mone, ILCSH, 6, 40.) 22,22 133, 149ff., 157 Intestinal absorption of water, 223, 237 Intestine (including jejunum and ileum), 120 endocrine secretions of, 46, 126, 156 exocrine secretions of, 117, 118, 126 Invertebrates, 2, 7, 8, 12, 153, 206 and glands, 115, 118, 129, 132-4 (33 and metabolism, 169, 190, 208, 228, 241 and muscles, 58, 61, 64, 67 and pigmentary effectors, 71, 74, 87 SUBJECT INDEX Invertebrates, possible hormonal control of salt transport in, 209, 252 sources of hormones in, 18, 21, 23. 355/39 Iodides, 140 Iodine, 12, 47, 140ff., 173 radioactive, 189 Ions (anions and cations), active transport of, 206, 209, 215, FAS: 2216 22 7e252 exchange of, 214, 216 Catt, 221, 240ff., 241,242, 245, 247, 248, 254, 256 Cl> 37, 206i... 208, 2008.,.212. 214, 218if.. 221; 252 El 214,027 Ke 205, 209i, 214, 215, 216, ZN 229 Naz. .206; 208, 2098. 212, 213, ot 216; 717, 218t.,221, 229 252 - Phosphate, 240, 241, 247, 249, Zale 204,250, 256 Iridosomes, between 82 and 83, 83 Iris of eye, 67 Islets of Langerhans, cells of, face 48 (6), 49ff. control of, 132, 133, 142ff., 155, Zate255 source of glucagon and insulin, 46, 47, face 48 (b), 49ff., 193, 194, 195 Isopoda (see also Ligia and Para- tya), 28, 29, 98, 99, 101, 108 Juvenile hormone, from corpora allata, 12, 39, 251 fn. Kidney tubules, impermeability of, 225228 permeability increased by hor- mones, 214, 233ff., 239 proximal, 225, 227 U 297 Kidneys, 214, 225ff. concentrating mechanism in Mammalia, 214, 227, 237ff. creatinine clearance in, 234, 235 excretion by, 226, 235 reabsorption of water in, 230, 233ff., 254520741. tubular reabsorption of P in, 250, 252 tubular secretion in, 237 Kinetic hormones, actions of, 8, 56ff., 98, 71, 87,.96,:98; 1156... 118, 129-259 and effectors (see Chromacti- vating and Endocrinokin- etic hormones and Exo- crine glands and Muscles) characteristics of, 7, 9, 152ff. control of, 155ff., 156, 157 effects of dosage with, 59, 63, 67, 154 endocrinokinetic control of, 128, 149, 157, 162 lack of antagonists for many, 154 likened to metabolic hormones, 252,297, sources of, 18ff., 22, 46 Kymograph records, 60, 62, face 62, face 68 Lacertilia (lizards), 36, 85, 175, 193 Lactation, 152 Lamellibranchia (bivalve molluscs), ES Larvae in diapause, 184, 185 Lepidoptera (moths andsilkworms), 136, 182ff. (see also Bombyx, Antheraea, and Hyalophora) Light, adaptation to, 75, 77, 78, 81, between 82 and 83, 154 direct effect of, 77, 84, 85, 96, 98 intensity, 79 reactions to, 75ff., 83, 84ff., 91ff., 101, 106 298 Ligia, chromactivating hormones, LDH and LLH, 87, 98, 100ff., 101 Lipophores, 83 Liver, bile flow from, 124 Locomotion, reduction of, 69, 70 Lumbricidae (earthworms), 4 Langs, 237 Luteal-stimulating hormone, LSH (see Luteotrophin) Luteinizing hormone, LH, 8, 44, 128; 1508; 152, 16118. Luteotrophin, LSH, 22, 44, 128, £535) ASO.) (LSS \(see?also Prolactin) Macrura (= Decapoda, other than Brachyura) 98fn. Malacostraca, 26, 28, 82, 83, 93 Malpighian tubules, 63, 223 Mammalia, 1, 253, 256 (see also Cetacea, Carnivora and Roden- tia) and balance of electrolytes and water, 207, 208, 214, 215ff., 216.27 7"219, D2 7it., 229; 230,233, 2378. 3238 and calcium and_ phosphates, 241, 246ff., 247, 249, 250, 251 and control of endocrine glands, 133, 140ff., 142, 144ff., 145, 146, 148, 1494. and control of kinetic hormones, 155; 196; 1575160; 162 and exocrine glands, 66, 68, TAT TSy 20120, A122, 1226f.; 123.125; 128if.,; 129; 130ff. and fat, 187 and intermediary metabolism, 190, 194, 1944f., 196, 197ff., 198, 205 SUBJECT INDEX and muscles, face 62, 63ff., 65, 67, face 68, 70 and respiration, 169, 173, 174, 175, 176;°177;- 135i sources of hormones in, face 22, 34, 35, 38, 41ff., 42; 44, 46, face 48, 50, 51, 53, face 140, face 141 Mammary glands, 65, 66, 68, 119, 121, 129, 130, 1511. 953 Median eminence of vertebrate brain, 35, 36, 41, 42, 142 Medium, hypertonic, 37, 207, 236 hypotonic, 206, 225, 237 saline, 218, 230ff., 232 “outside’’, 233 Melanin, 75, 76, 83 Melanophore-concentrating hor- mone, MCH or W, 8 in Amphibia, 87, 103, 104, 160 in fish, 87, 97, 106ff., 106 source of (in Pars tuberalis), 22, 43, 103ff., 157, 245 Melanophore index (see Chromato- phore index) Melanophore-stimulating (dispers- ing) hormone, B, MSH, or Intermedin, 8, 102ff. in Amphibia, 87, 103ff., 104, 160 in fish, 87, 94, 97, 105ff., 106 in Reptilia, 107 source of, 22, 44, 105, 157 Melanophores (see Chromatoph- ores, black pigment in) Meso- and Meta-adenohypophysis of fish, 40, 107, 212 Mesoderm, as hormone source, 18, 50:47 endocrine cells of, face 48 Mesodermal endocrine glands, 9, 508., 51, 53,1430: Mesonephric duct, 52 Metabolic effects, on activity, 69 affected by endocrinokinetic hor- mones, 132 SUBJECT INDEX Metabolic hormones, actions of, 167ff., 169, 190, 208, 241 (and see Calcium, Car- bohydrates, Electrolytes, Fat, Phosphates, Protein, Respiration and Water) and internal ‘“‘milieu’’, 259 characteristics of, 7, 8, 10, 252¢f. control by feed-back, 11, 134, 252 control of secretion of, 253ff., 25 250, 259 endocrinokinetic control of, iS2tP., 133,137,139) 1428. , 145, 258ff. morphogenetic actions of, 173, 257 259 pairs of antagonistic, 10, 189, 207i, 214, 219. 220° 245, 22,256 sources of, 18ff., 38, 40, 44, 46, 47,50, 51, 1341. Metamorphosis, 8, 11ff., 138, 140, Za lin, 259 in Amphibia, 12, 140, 176 in Arthropoda, 11, 138 Methylthiouracil, MTU, 139 Migration of birds, 149 Milk, 65, 68, 130 “‘let-down’’, 66, 68, 152, 154 pigeon’s, 130 Muilk-secreting glands, 129, 130ff. (and see Mammary glands) Mineralocorticoids, 185 Mitosis and secretion, 39, 52, 116, 117 Mitochondria, between 22 and 23 Moisture, and pigmentary effectors, 75ff., 76, 85 and water balance, 222, 230, 232 Mollusca, 11, 19, 115ff. (see also Cephalopoda, Gastropoda and Lamellibranchia) Monoiodotyrosine, 141 Monosaccharides, 197 299 Morphogenetic hormones, actions of, 128, 134, 139, 148, 149, 150, 151ff. (see also Part IT.) and feed-back, 134 characteristics of, 4,7, 8, 10, 11ff. definition of, 7, 11, 167 distribution of, 11 endocrinokinetic control of, 10, 132, 13d, 157, 1433 ieet, 162ff. kinetic actions of, 70, 128, 149, 157, 1628. of gonads, 133, 149ff., 157, 259 sources of, 18, 19, 38, 40, 50 Moult-accelerating hormone, from HSPO, 135, 156, 258 Moult-inhibiting hormone, MIH, of Crustacea, and protein metabolism, 200ff., 202, 203, 204 from eyestalk or sinus gland, 12, £35, 098621871%.,4 90 221i. 247, 248, 252 nervous control of secretion of, 254257 Moult-promoting hormones, 12, 18 and diapause, 184ff. endocrinokinetic control of se- cretion of, 133, 254, 255,258 of Crustacea, from Y-organ, MPEG, 135,,-2902 203.2204. 222 JAGR. 24 oan oe. 254 of Insecta, from prothoracic gland, etc. (= ecdysone), 129, 134, 135ff., 137,, 184, 190, 195, 241 251 255 sources of, 18, 38 Moulting, changes in Ca and P during, 240ff., 242, 242, 248, 249, 251 hormonal control of, 8, 11, 117, 1345 1358.4 181, 284, 2299; 200, 203, 220, 251fn. 300 Moulting, metabolic changes during, 181, 187, 195," 1998. 202; 203, 204 water uptake during, 220ff. Moults, time of, 40, 74, 200ff. forced, 202,221, 257 Mucosa, 3, 44, 45, 63, 120, 124 Mucus glands of epidermis, 129, Pd Muscle tone, increase of, 69, 70, 156,157 Muscles, 8, 9, 13, 56f., 58; 152f., 157 calcium content of, 247 of blood vessels, 67, face 68, 237 of chromatophores, 39, 71, 72, 726k. of gall bladder, 63, 127, 153 of genital ducts, face 62, 64ff., 65, 153 of glands, 66 of out, 57; 62ff.,“face 62, 127, 153.154 of hair follicles, 67 of heart, 57ff., 60, 154 of iris, 67 6f mantle, 21, 23, 69 sodium concentration in, 217, 219 somatic, 63, 69ff. specificity of, 153, 154 tetany of, 244, 247 Myoepithelial cells, 58, 65, 66, 68, 152,°453 154,159 Nasal glands, 218 Nematocysts, 10 Nerve, action, 9, 20 axons, 19, 41 cells, 19ff., face 22, 32, 33, 45 impulses, 20, 155, 258 SUBJECT INDEX Nerve section, and secretion, 183, 191, F9OZ2257 Nerves, adrenergic, 56, 106, 131 cholinergic, 56, 106, 131 parasympathetic, 45, 62ff., 127, 154 stellate, 23 sympathetic, 37ff., 41, 42, 45, 621.467; 68, 122, 1232125. 131, 154 vagus, 62, 64, 123, 124, 125, 127 Nervous control, ; of blood supply, 41 of chromatophores, 72, 83, 94, 97, 102, 106, 107 of hormone secretion, 9, 23, 38, 66, 68, 101, 106, 155, 156, 157, 158... 191,205; 259: 259295; 2 50 of salivary glands, 117 Nervous system, 7, 155, 205, 259 cells derived from, 19ff., face 22 central, of Crustacea, 24, 24ff. central, of Insecta, 29ff., 30 central, of Vertebrata, 33ff., 48 of Cephalopoda, 23, 69 parasympathetic (see Nerves) relieved by hormones, 155 stomatogastric, of Insecta, 30, 39 - sympathetic (see Nerves) Neural crest, 45 Neural lobe of pituitary (see Pars nervosa) Neurofibrillae, 19 Nuerohaemal organs, 4, 20, 56 (and see Storage-and-release or- gans) Neurohormones, 3, 20, 39, 56, 61, 135;-158 Neurohumoral secretion, 3 Neurohypophysial extracts, with frog ADH, 131, 210, 218, 226, 232, 236 mammalian, face 62, 218 SUBJECT INDEX Neurohypophysial hormones, 33, 2AT 214, 2195 230,233, 234, 235, 252 (and see ADH and Oxytocin) storage-and-release organs for, 22, 33, 34, 35, 42, 64 Neurohypophysis, and kinetic hormones, 64, 67 and metabolic hormones, 208, 220, 234, 241, 245, 248ff. axons in, between 22 and 23 (g), 34, 42 control of secretion from, 157, SOT e 21, 239,294" 257 extract of, face 62 (and see Neurohypophysial extracts) neurosecretions stored in, 34, 35, 64,167, 218; 232.1236 removal of, 217, 219, 228, 229; press 2 55 structure of, 22, 33ff., 34, 36, 40, 41, 42 (= ME + NL), 48 -Neurointermediate lobe of pituitary (see Pars intermedia) Neurons, 2, 19ff., face 22, 23 Neurosecretion, 4, 10, 19ff., 63 aetions of, 50,°57.(61,.75, 138it., 190; Lo/ lO 203i, 254 (and see many Kinetic and especially Chromactivating hormones and Sinus gland. etc.) nervous control of, 10, 23, 156, Poe Aoi. 2Ue2o4. 257 sources of, 19ff., 23, 24, 26, 30, 34, 258 (and see Neuro- secretory cells) storage-and-release of, 20, 22, 28,32, 34, 35, 62, 203, 236 (and see Storage-and-release organs) Neurosecretory cells, 2, 4, 19ff., face 22 compared with neurons, 19ff., 258 301 controlled by nerves, 156, 157, 158ff., 254, 257, 258 in Cephalopoda, 21, 23 in Crustacea, 24ff., 24, 26, 27, 188 in Insecta, between 22 and 23, face. 23, 29ff, 30; .32°62 in. Vertebrata, 33i.,o4, 37,44, 42, 45, 68 of intercerebrum (l.n.c. and mn.c.),8, face 23, 30, 314., 13345 135i. 251-79, 195. 20a, 223,255 suboesophageal (s.n.c.), 29ff., 30 tissue culture of, 35 Neurosecretory, granules, 9, 19ff., face 22, between 22 and 23 (Figs 2-1 g and 2-2), face 23, 27 material (Frog ADH), 232 store, 236 substances, 9 systems, 22, 23ff., 24, 29ff., 33ff., 34, 62 Nissl bodies, 19, face 22, between 22 and 23 Nitrogen, excretion of, 201, 204 (and see Protein catabolism) metabolism of, 199ff., 200, 202 Noradrenaline, 21, 37, 38, 56, 59, 60, 68 Nutrition, 205 Octopoda (octopus), 21, 58, 69, 73 Odonata, 135 Oestrogen, 58, 64, 70, 129, 130, 134.133 51501525137 Oestrone, 150 (see also Oestrogen) Oestrus cycle, 70, 131,°150, 157, 152 Ommatidia, 74, 77, 78, 80, 81, 100 Ophidia (snakes), 36 302 Optic lobe, 24, 24, 25, 26, 26, 30 ganglion in, 26 terminal medulla of, 24, 25, 26, 28 Organiser, 2, 3, 4 Organisine, 3, 4, 8, 12 Ortho-dihydroxytryptamine, 59 Ortho-diphenol, 61 Osmoreceptors in hypothalamus, 239,239, 257 Osmotic concentration of urine, 239 Osmotic diuresis, 227 ‘Osmotic gradient, 206, 231, 232, EX) Osmotic pressure, 74, 221, 227, 235,237 Ovarian growth, 170 Ovary4, 150, 153; 157; 170; 182; 186 maturation, 245 Oviducal glands, 128ff., 129 Ovulation, 246 Oxygen consumption, 167, 168ff., 169 (and see Respiration) accompanying changes in fat storage, 186, 188 ana TSH,.139 in Crustacea, 168, 178ff., 178, 179, 180, 181 in Insecta, 138, 168ff., 171, 172, 182ff., 191, 250 in Protochordata, 173 in Vertebrata, 173ff.,; 175, 176, 185ff. Oxyntic cells, 119 Oxytocin, and milk “‘let-down’’, 65ff., 66, 68, 130ff., 154, 159, 258 and muscles, 58, face 62, 64ff., 65,'67;°152°153,:15:7 and salt excretion, 208, 211, 214, 21S; 249 Jad and water balance in Amphibia, 230 source of, 35ff. SUBJECT INDEX Palaemonetes, chromactivating hor- mones, and red pigment, PDH and PLH, 8, 87, 91ff., 92, 98, 156 and white pigment, PWCH and PWDH, 87, 95, 96 Pancreas, as endocrine gland, 44, 46, 132, 142, 193, 194 (and see Islets of Langerhans) as exocrine gland, 1, 45, 117, 118, 120, 122ff.; 1231248 125 transplantation of, 123, 123, 125, 125 Pancreatectomy, 193, 197 Pancreozymin, 46, 118, 120, 124, $2554.25 2027-156 Para-activators, 2, 73 Parabiotic graft, 69 Parasympathetic nervous system (see Nerves) Parathormone, actions of, 8, 243, 246, 247, 250, 251 means of control, 254, 256 secretion of, stimulated by dark- ness, 246 source of, 46, 48, 49 Parathyroid glands, 49, 24] and calcium, 241, 243, 244, 246 and phosphates, 250, 251ff. cells of, 49 control by level of Ca and P in blood, 142, 254, 256 extirpation of, 246ff., 247 extract of, 246, 251 hypertrophy of, 252, 256 not under endocrinokinetic con- trol ;132, 142 secretion stimulated by darkness, 246 structure of, 46, 47, 48, 49 Parathyroidectomy, 246, 247 Paraventricular nucleus of hypo- thalamus, 22, 33, 34, 35, 37 SUBJECT INDEX Pars distalis of pituitary, 22, 34, 36, 40, 41, 42, 43, 44, 157, 255 (see also Adenohypophysis) Pars ganglionaris X organi, 25 (see also Ganglionic-X-organ) Pars intermedia of pituitary, as source of hormone, 94, 103, 104, 106, 106, 107, 157 structure of, 22, 34, 36, 40, 41, 42, 43 Pars nervosa (neural lobe of pitui- tary), 34, 35ff., 36, 37, 40fn., 42, 43, 48 (see also Neurohy- pophysis) Pars tuberalis of pituitary, as source of hormones, 87, 103, 104, 104, 105, 106, 157, 241, 245 regeneration of, 245 structure of, 22, 34, 36, 40, 41, 42, 43 Parturition, 64, 65, 65, 152 Pepsin secretion, 119, 120 Peptones, 123, 125, 125, 126, 156 Periodic-acid Schiff reaction, 44 Pericardial organs, 22, 25, 29, 57, 35, 59, 1396 extracts of, 59, 60 Perirenal organ, 50 Peristalsis, 58, face 62, 62, 63, 155 Peritracheal glands, 135ff. Permeability of cell membranes, decrease in, 208, 219, 220ff. increase in, 208, 228ff. (and see Pores, ultramicroscopic) of chromatophores, 74 of kidney tubules, 214, 233ff., 234, 235ff., 238 of skin, 226, 230, 231, 232 Pharynx, glands of, 44, 46, 46, 47, 48, 49, 173 Phasmida (see Carausius) Phenylalanine, 125 Phosphates, and moulting, 249ff. 303 Phosphates, and parathyroids, 142, 251ff., 256 balance of, 241, 249ff., 247 in blood, 254, 255 metabolism of, 142, 240, 249 Phospholipids, 43, 187 Phosphoric hexose esters, 197, 250 Phosphorus turnover, 117 Phosphorylation, 177, 196ff. Physiological colour change (see Colour change) Pigeon’s milk, 130 Pigment granules, 73ff., 75, 77, 79ff., 82ff., 108 black, 74, between 82 and 83, 83, 97ff., 98 blue, 83 concentration of, 73ff., 79, 83, 88ff. dispersal of, 73ff., 79, 83, 91ff. in epidermal cells, 74ff., 75, 158 in mesodermal cells, 102 in retinal cells, 77ff., 78, 81 movements of, 73, 74, 80, 82ff. red, 74, face 82, 83, 86ff., 98 reflecting, 83 under pressure, 74 white, 77, 83, 94ff., 96 yellow, 83 yellow-green, 74, 75 (see also Chromatophores) Pigment-concentrating hormone of Penaeus, 90 Pigmentary effectors, 56, 71, 73ff., OF, 107i. Vo 2tk. Pineal organ (see Epiphysis) Pitocin, 237 Pitressin, 237 Pituitary body, 34, 36, 48, 245 anterior lobe of, 40ff., 40fn. (and see Adenohypophysis) extracts of, 104, 199 posterior lobe of, 33ff., 104 (and see Neurohypophysis) Pituitary gland, 42 304 Pituitrin, and calcium, 245, 246, 249 antidiuretic fraction, 210, 211 oxytocic fraction, 68 Plant hormones, 2, 3 Plasmosol phase, 74 Platyhelminthes (flat worms), 12 Plecoptera (stone flies), 30 Pleuronectidae (flat fish), 83, 110 Pores, ultramicroscopic, and ADH, 233-230,239 Porifera (sponges), 4, 167 ' Post-commissure organs, 24, 25, 29, 90 Potassium, Kt (see Ions) Pouch, from fundus of stomach, 119. 428; 122 from pylorus, 119, 120 Precursors of hormones, 91, 140 Pregnancy (and gestation), 64, 150, 1515-152 Preoptic nucleus, 33 Pro-, meso-, and meta-adenohy- pophysis, 40 Progesterone, actions of, 64, 128ff., 129,152,153 endocrinokinetic control of, 133, 1508. 137, 162 source of, 50 Prolactin, LUSH, 128;1295130; 150, 152.153, 157, 260; 161 Pronephric ducts, 52, 227 Protective, coloration, 70, 84 responses, 259 Protein, anabolism, 199, 201 catabolism, 205, 249, 250, 251 contraction of molecules, 80, 82 digestion of, 125 in moulting crabs, 202, 204 metabolism, 10, 11, 135, 189, 190, 1998200; 202,252 of chromatophores, 74 restraint of catabolism of, 190, T998f., 204, 204, 257 SUBJECT INDEX synthesis, 190, 203, 205, 249, 2524293, 2o4a2o0 Prothoracic gland hormone (se Moult-promoting) Prothoracic glands, and metabolism, 183ff., 790, 195, DAL, 258 as source of ecdysone, 8, 18, 30, 38 endocrine control of, 133, 135ff., 2995258 Prothoracotrophin, 12, 136, 137, 156, 184, 258 (see Intercere- brum, for source) Protocerebrum, of Arthropoda, 22, 24, 25, 30, 31 Protochordata, 47, 173, 174 Proventriculus of birds, 122 Pupae, in diapause, 137, 184, 185 Reflecting cells, 77 Regeneration, 8, 11, 135 Release mechanisms for secretions, 258 Renal calculi, and Ca metabolism, 243 Reproduction, 148ff., 151ff. Reptilia, 33, 36 (see also Chelonia, Lacertilia and Ophidia) and kinetic hormones, 74, 83, 85, 94, 107, 109, 110, 124 and metabolic hormones, 169, 193, 208, 215, 237 Respiration, 8, 10 aerobic, 209 and cytochrome c, 182 cyanide stable, 182 decreased rate of, 168, 169, 171, 178ff. increased rate of, 8, 168ff., 169, 172 Respiration-inhibiting 178, 179, 180, 254 hormone, SUBJECT INDEX Respiratory accelerator hormones, 168if., 169; 254, 255 Respiratory inhibitor hormones, 178ff., 254 Respiratory quotient, 180 Responses, graded, 154 Retina, 24 Retinal cells, contractile fibres in, 71, 79, 80, 81 distal, 71, 74, 77ff., 78, 80£f., 81, 108 . proximal, 77, 79 Retinal-dark, and -light-adapting hormones, 80ff., 81. Retinal-pigment-concentrating and -dispersing hormones, RPCH and RPDH, 78, 79ff., 108, 156 Retinula cells, 74, 81 Retrocerebral system of insects, 33 Rhabdome, 81 Rhythm, diurnal, in Ca metabolism, 242 diurnal, in pigment cells, 80ff., $5; 86, 93; 99, 103, 259 of nocturnal activity, 69 of muscle contractions, face 62, 63 of secretion, 116, 144 tidal, 85, 259 Ring-gland, of Diptera, 135, 195 Ringer’s solution, 59, face 62, 63, 213,233 Rodentia (see Cavia and Rattus) Saccus vasculosus, 41 Saline, hypertonic, 228, 239 injections, 14, 37, 148, 212, 224 medium, 218, 232, 233 (see also Ringer’s solution) Salinity, changing, 215, 227 excessive, 212, 218 Salivary glands, of Cephalopoda, 22, 36it., 735.1 16,208,230 of Mammalia, 117, 216 305 Salt, NaCl, concentration in blood, 143, 206, 208, 209ff., 225ff. excretion of, 13, 213, 214, 218, 218fn., 220, 258 injection of, 14, 92, 175, 191, 224 in medium, 224, 232 reabsorption of, 206, 211, 212, 214, 218, 235 uptake, 210, 211 (see also Electro- lyte balance and Ions) Seawater, 92, 212, 218, 226, 236 hypertonic, 37, 207, 236 hypotonic, 237 migration from, 206, 212 Secretagogues, 5, 117, 119, 124 secretin, action of, 4, 8,116, 119. 120120512 2ir-2 127 control of secretion of, 57, 123, 123,196; 253 source of, 45, 46, 56, 64, 123ff. Secretion, by nerves (see Neurosecretion) inhibition of, 117, 126ff. of acid, 118, 119ff., 120, 122, 127, 15352253 of bicarbonate, 120, 122ff., 123, 253 of enzymes, 115, 116, 117, 118, 1248f., 12555127 of hormones, q.v. phases of, 20, face 22, between 22, and: 23; 325 52 Secretory cells (see Endocrine and Exocrine secretory cells) giant, 32 Sensitivity, threshold of, 154 specific variation in, 235 Sex-differentiation, in pigmentation of Uca, 99 in secondary characters, 150, 151 Sinus gland, as storage organ, 22, 24, 25ff., 26, 72 IO MS J equivalent organ in Ligia, 102 149, 306 Sinus gland extracts, 59, 89, 90, 91, 92.94 5° 97; 9 78H 191, 221, 222, 248 Sinus gland hormones (and _ see Eyestalk hormones) chromactivating, 87, 88ff., 89, 90, 92, 94ff., 96, 97ff., 98, 108 control of secretion of, 156, 254 decreasing fat consumption, 169, 187ff. diabetogenic, 8, 189ff., 190, 192, 257 diuretic, 208, 220, 220ff. heart-accelerators, 58, 59 moult-inhibiting, q.v. respiration-inhibiting, 169, 178ff. restraining protein catabolism, 190, 199ff., 203, 204, 254 retinal-light-adapting, 71, 80ff. storage of, 25ff. Sinus gland implants, 220, 220 Sinus gland, means of controlling, tao; 2o4 removal of, 80, 94, 178, 179, 181, 187, 188, 189, 192, 199, 200, 203, 248 sectioning nerve to, 192, 257 structure of, 24, 25ff., 26 supplying kinetic hormones, 58, 71, 80, 81, 87, 88ff., 89, 96, 97ff., 93, 108 supplying metabolic hormones, 169, 178ff., 189, 190, 208, 220, 241, 247, 248 Skeleton, decalcification of, 243, 252 Skin, absorption of ions by, 209ff. glands, 117, 129, 131 imbibition of water by, 206, 231, 232 impermeability in Reptilia and Mammalia, 207, 237 permeability to water in Am- phibia, 230ff., 231, 232 SUBJECT INDEX reactions of isolated, 226, 231 Skin sensitivity to hormones, 231 ultramicroscopic pores in, 233, 239 Sodium chloride (see Salt) Sodium ions, Nat, 37, 206ff., 208, 214,217, 252 increase in blood, 209ff., 210, 241. 212i eee decrease in blood, 213, 216, 218ff., 229 Sodium pump, 209 Somatotrophin, STH, control of uncertain, 157 relation to insulin secretion, 256 stimulation of glucagon secretion, 133, 142ff., 194, 194ff., 199, 239, 299 source of, 22, 44 Sources of hormones, 2, 18ff., 22, 46, 51 (and see Endocrine glands and Endocrine secre- tory cells) ectodermal, 18ff., 22, 38ff., 44 endodermal, 46, 46ff., face 48, 48, 119ff., 127, 139ff. extracts of (see Extracts) from nervous system, 18, i9ff., 22, face 22, 38 from neurosecretory cells q.v. in Cephalopoda, 21ff., 22, 23, 38 in Crustacea, 22, 23ff., 24, 26 in Insecta, 22, between 22 and 23, face 23, 29it., 30pa2.39. 62 in Vertebrata, 18, 19, face 22, 22, 33ff., 34, 36, 40ff., 42, 44, 44ff., 46, face 48, 48, 50ff., 51,53 location of, 12ff., 14, 86, 109 mesodermal, face 48, 50ff., 51, 53 Sperm in fallopian tube, 65 Starvation, 187, 188, 200, 205 Stellate ganglion, 21, 22, 23 Sterolic hormones, 50, 52 SUBJECT INDEX Stimulation of hormone secretion, aa Sou. V1 Ssit., 1596, 157, 255i. 294255 chemical, 57,.123f., 125f., 155, 156, 253ff., 254 hormonal; 57; 115, 1318., 132, P27 MSS 01 G2, 2545255, 258 (and see Endocrinokin- etic hormones) mechanical, 56, 119ff., 155, 156 mervous, 38,.57, 156,157, 158, 160ff., 254, 257ff. Stomach, 5, 44, 45, 119, 120, 156 glands (gastric), 118, 119ff., 126 pouches, 119ff., 121, 122 Stomatogastric system of insects, 30, 39 Stomatopoda, 25, 29 Stomodaeum, 30, 33, 40, 48 Storage-and-release organs for neurosecretion, 4, 20, 22, 258 in Crustacea, 23ff., 24, 26, 28, 80, 188, 248 in Insecta, 30, 31ff., 32, 61, 62, 173,203) 222 in Vertebrata, 33, 34, 35, 42, 44 Stress, 167, 189, 195, 205 (and see Emergency) and ACH, 52, 144ff., 145 from asphyxia, 189ff., 192 from haemorrhage, 11, 145 Suboesophageal ganglia, and diapause, 169, 182ff., 254, 257 and motor activity, 58, 69, 153 and pigment dispersal, 71, 75ff., 76, 156 as source of hormones, 22, be- tween 22 and 23, 24, 29ff., 30 endocrinokinetic action of hor- mone from, 133, 135 extract of, 223, 224 Succus entericus, 126 Sugar, in blood (see Blood-sugars) in tissues, 189, 195, 197ff., 253 307 Sugar, in urine, 234 (and see Diabetes mellitus) Sugar molecules, responsive to insulin, 197, 198, 253 Supraoesophageal “‘brain”’ of Arth- ropoda, 28ff. Supra-optic nucleus of hypo- thalamus, 22, face 22, 33, 34, 35 Suprarenal body, 33, 68 (and see Adrenal medulla) cells of, 51 Suprarenal, gland, 38fn. tissue, 22, 37 Sweat glands, 129, 131, 216, 237 Sympathetic nervous system (see Nerves) Syndrome, emergency, 67 Synergistic action of hormones, 150, 252,256 Syrphidae (syrphid flies), 185 ‘Teleostie, and kinetic hormones, 83, 94, 102, 106, 106ff., 109, 110, 139 and metabolic hormones, 169, 174, 176, 193, 197, 208, 212, 2163-226, (2oa hey ao hes | sources of hormones in, 46, 47, a1 Sit. use of mammalian hormones on, 212, 218, 243ff. Temperature, effect on chromato- phores, 85 Terminal medulla of optic lobe, 24, 25, 26, 28 Terrestrial mammals and water, 237 Testis, 1, 4, 6, 8, 149, 151, 157 Testosterone, 6, 8, 70, 133, 149, 15d toe, Tetany, muscular, and Ca lack, 244ff., 246, 247 308 Tetrapoda, face 22, 33, 36, 40, 46, 47, 49, 50, 51, 51, 67 (see also Amphibia, Reptilia, Aves and Mammalia) Thiouracil, 141 Thiourea, 233 ‘Vhacst: 373207, 239 Thoracic ganglia, 24, 30 Threshold, levels of sensitivity, 154 values for hormone action, 249 values for hormone stimulation, 256 Thymus gland, 47, 48 Thyroglobulin, 140 Thyroid gland, and blood-P, 241, 243, 244 as source of thyroxine, 46, 47ff., face 48(a), 48 cells of, 48ff., face 140, face 141 endocrinokinetic control of, 133, 139ff., face 140, face 141, 255, 256 (and see TSH) extracts of, 174, 175, 176 in Agnatha, 173ff. in Amphibia, 140, 174. in Aves, 177 in fish, 139, 174,189, 208, 213ff., 227, 244 in hibernation, 186 in Mammalia, 140, 175, 176, 177, 187, 188ff., 194, 208, 241, 243 in Protochordata, 173 morphogenetic effects of secre- tion, 6, 12, 140, 257,259 Thyroidectomy, difficulty of, 174, 177, effect on blood-P, 250 Thyrotrophin, TSH, and iodine metabolism, 140ff. and parathyroid glands, 243, 245, 246, 256ff. control of secretion of, 141, 149, 157, 161 SUBJECT INDEX endocrinokinetic, action of, 12, 49, 133, 139ff., face 140, face 141, 174, 177, 187, 188, 213, 299, 259 growth-promoting action of, 10, 142 . source of, 22, 43, 44 ‘Thyroxine, action of, 169 in Amphibia, 175, 259 in Chordata, 70, 169, 173, 187, 188ff., 208, 243 in: fisla.4 245, 227 in Mammalia, 176, 177, 195, 250 Thyroxine, and cretinism, 1 and cycle of iodine metabolism, 140ff. and feed-back, 141, 157 and metamorphosis, 8, 12, 259 and parathyroids, 243, 256 precursors of, 49, 140, 141 secretion stimulated by TSH, 133; 1398 255 source of, 46, 47, face 48 (a) ‘Tissues, and sugar supply, 197£.; 253 embryonic, 182, 184 enzymes of, 140 hormone breakdown in, 59, 91, 140, 155, 184 iodine cycle in, 140 permeability changes in, 220, 225, °226h0. protein synthesis in, 190, 249 (and see Protein) protection from diapause, 184 ff. salt content of, 206, 207, 215 water content of, 206, 226ff., 230, 237,239 Translocation, of calcium phosphates, 250 of fats, 255 Tritocerebral, commissures, 24, 28, 29, 80, 90, 91, 92, 94 lobe, 25 189, 195, and SUBJECT Tritocerebrum of Arthropoda, 24, 255, 2830 Trophic hormones, 9ff., 132 (see also. Endocrinokinetic hor- mones) Tropic hormones (see ‘Trophic hormones) ‘Tryptophane, 125 iycarmine, 22, 38, 71, 73 Uca, chromactivating hormones of, 87 for black pigment, UDH and ULH, face 83, 98, 99 for red pigment, URCH and URDH, 89, 93, 98, 99 for white pigment, UWCH and UWDH, 95, 96 Ultimobranchial bodies, 46, 47, 48, 49, 241, 243ff., 251 compared with parathyroids, 244, 256ff. Ultra-violet light and Ca, 246 Urea, 227, 234 Ureters, separate collection of urine from, 238 Uric acid, 237 Urine, 214, 231, 234, 235, 237, 239 hypertonic, 238, 239 hypotonic, 206, 225, 227, 239 output, 216, 229, 238 Urochordata, 173 Urodela (Necturus and _ sala- manders), 35, 36, 41, 128, 193 Uterine glands, 128ff., 129 Uterus, face 62, 64, 65, 151 Vagal nerve endings, 239 Vagus nerve, 45, 62, 64, 121, 123, A124 21255127 Vas deferens gland, 8, 132 Vascular constrictor, 239 (and see Vasopressin) INDEX 309 Vascular hormones, 3, 4, 6, 11, 19, 20 et seq. Vascular plexus, 36, 41, 42 Vascular plexus in cortex, 53 Vascular system, 67 Vascular tissue, 119 Vasoconstriction, in Cephalopoda, 116 in Vertebrata, 66, 67, face 68 Vasodilation, 66, face 68 Vasopressin, ADH, 58, 67, 157, 159, 211fn., 230, 234, 239 (see also Antidiuretic hormone) Venous plexus of pituitary, 36, 41, 42 Ventral gland, 30, 30, 133, 135 Vertebrata, 2, 7, 8, 11, 12 (and see Agnatha, Amphibia, Aves, Elasmobranchii, Fish, Mam- malia, Reptilia and 'Teleostei) and balance of Ca and P, 240, 241, 243ff., 247, 249, 250ff. and balance of electrolytes and water, 208, 209ff., 214, 216, 220, 225ff., 230i. 235 and chromatophores, 83, 86, 87, 94, 102ff., 104, 106, 109, 110 and control of endocrine glands, 132, 133. 139, 145; 156, 157, 159; 160H. 254255 and control of metabolic hor- INGMES, 205, 204, cIa, cod, 259 and exocrine glands, 66, 115, 116, 117,116), 120; 122, 123, 125, 128i.,429, 1308. and intermediary metabolism, 185ff., 187, 188, 189, 190, 191ff., 195ff., 196, 205 and muscles, 56, 58, 59, 61ff., 63f.; 70 and respiration, 167, 168, 169, 1738; 1757. 176,485 sources of hormones Sources) in (see 310 Vertebrata cold blooded, 5, 44, 63 and kinetic hormones, 82, 122, 124, 144, 149, 155 and metabolism, 173, 193, 253 Vertebrata, lower, 45, 47, 74, 110, 127 Visceral arches, 48, 49 Visceral muscles, 57ff. Visceral nervous system, 30 Vitamin D, 246 Viviparous development, 151 W (see Melanophore concentrating hormone) Water, antidiuresis, 228ff., 238 balance, 8, 10, 143, 168, 206, 208, 214, 219ff., 220, 232, 239 ‘‘Water-balance principle”’, 230 Water, diuresis, 143, 201, 207, 208, Zits 209R.. 223,224, 2258f., 240, 257 excretion, 222, 223, 229 flux through skin, affected by hormone, 233 loss, 228, 231 reabsorption, 230, 234, 235, 236, 237 SUBJECT INDEX retention, 223, 246 uptake, 220ff., 223, 224, 226, 228, 2308. Z3L 259 Water-loaded, rats, 227, 228, 229 tissues of fish, 227 Weismann’s ring (gland), in Dip- tera, 135, 195 Xanthophores, between 82 and 83, 83 X-organ, confused use of name, 28 (see Ganglionic-X-organ and Hanstr6m’s sensory-pore or- gan) Y-organ in Crustacea, 8, 18, 38 and metabolic hormones, 190, 199, 203, 204, 222, 240, 241, 242ff., 249, 251 endocrinokinetic control of, 133, 134ff., 254, 258 extract of, 242 Zona fasciculata, 53 vvvvws vw <3 yy Z xz ¢ < < < < < < < < < < < 953 4 < 4 < < < < < < < < < < < < < 4 < é 4 < < < 4 4 < < € € é 1 < wy ee Ck <54 <3e <7 <8 o< «7k ? , < « « é ‘ < < < é < ‘ < < & é < z ne Z < é “SK y < < < < < < r < < ‘ < ‘ 4 < y < < 4 < < % % < < < < é é y < < 4 é 4 < < < < Z “ ang <4 ce & Se >?) 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