The nerve fibres in the basement membrane and related structures in Saccoglossus horsti (Enteropneusta)

Summary The structure of the basement membrane of Saccoglossus horsti has been examined with the electron microscope. The membrane consists of two lamellae each of two layers. An outer amorphous layer 150 nm across and an inner fibrillar layer 1–3 m across. The fibrils of the fibrillar layer are two sizes, the majority are 5–9 nm in diameter and at least 2 m long. The thicker 30 nm fibrils occur in small patches and have striations with a 30 nm period.Within the lamellae of the basement membrane are blood spaces. The only regularly found structures in these spaces are blood particles some 12–16 nm in diameter.Nerve fibres of varying diameters traverse all the layers of basement membrane. These fibres run longitudinally and obliquely through the basement membrane, and emerge amongst the muscle cells inserted into the coelomic side of the membrane. No motor end plates have been seen. Preliminary observations suggest that many of the nerve fibres have no sheath other than the cell membrane of the fibre itself.The muscle cells are attached to the basement membrane by structures that resemble hemidesmosomes. The blood vessels of Saccoglossus have a basement membrane on the lumenal side of the endothelial cell cytoplasm.I wish to thank Professor J. Z. Young, F. R. S. for continuous encouragement and advice. To Dr. R. Newell I am indebted for the collection and identification of the specimens. I am pleased to acknowledge my debt to Dr. R. Bellairs for the use of electron microscope facilities, and to Mrs. J. Hamilton and Mr. R. Moss for skillful technical assistance.     

朱鹮卵壳的微观结构和成分研究

本文首次对世界珍禽——朱鹮的卵壳在电子显微镜下的微观结构进行了研究;对卵壳中的26种无机元素进行了定量分析。根据两巢卵壳所含有害元素的对比,以及两巢区土壤中有害元素含量对照情况,指出保护朱鹮自然种群、研究其环境因子,刻不容缓。     

The structure of a photoreceptor organelle in the eye of Pterotrachea mutica

Summary The photoreceptor cell of Pterotrachea consists of an elongated cell some 100 m long with recogniseable inner and outer segments. The photoreceptor membranes point towards the light. There are about 300 discs per photoreceptor, a small number of discs arising from a single ciliary base. There are bout 75–100 such bases on each receptor cell. The receptor cells themselves (the inner segments) have four recognisable regions. The vacuolated region, the region of mitochondria, the nuclear region, and the axonal region.The photoreceptor cells are organised in five roughly parallel rows, and separated from one another by pale supporting cells.My thanks are due to Professor J. Z. Young, F. R. S. for his enthusiastic support and help during this work. Dr. R. Bellairs kindly provided electron microscope facilities. Mr. R. Moss, Mrs. J. Hamilton and Mr. A. Aldridge provided excellent technical and photographic assistance.     

Osmium tetroxide-zinc iodide reactive sites in the photoreceptor cells of the retina of the rat

Summary The osmium tetroxide-zinc iodide fixative of Champy-Maillet has been used to study the rat's retina at the electron microscope level. Electron opaque deposits were observed all along the photoreceptor cells and concentrated in the outer segments of rods and cones and in the nerve endings. In the outer segments that deposits are located in the inter and intra disk spaces as well as between the disk and outer membranes. In the outer plexiform layer reactive sites include synaptic vesicles and mitochondria; other minor reactive sites are described in the inner segment and inner plexiform layer.Electron opaque deposits were not seen if potassium iodide substitutes zinc iodide in the fixative. However, if osmium tetroxide-potassium iodide fixed retinae are immersed in osmium tetroxide-zinc iodide the characteristic electron-dense material is evidenced at those same sites. The effect of other several fixatives were studied with a similar double fixation procedure. Our finding points to the histochemical demonstration of an unidentified component (s) of the retina which shows a striking specificity of localization and which is made evident when zinc iodide is used in the Champy-Maillet mixture.This work has been supported by grants of the Consejo Nacional de Investigaciones Cientificas y Técnicas, Argentina and U.S. Air Force AF-AFOSR 67-0963 A.We are greatly indebted to Miss Haydée Agoff and to Mr. Alberto Saenz for their skillful technical assistance.     

The fine structure of the olfactory tract in the teleost Carassius carassius L.

Summary Electronmicrographic montages of the olfactory tract at two levels in each of two fish (Carassius carassius L.) were constructed and fibre diameters measured using a Zeiss TGZ 3 particle size analyzer. Medial and lateral tract divisions, rhinocele and dorsal tela were identified. Ciliated ependymal cells line the rhinocele. Meninges form the outer covering of both tract divisions and the tela roofing the central canal.The lateral tract consists of 10–14 fasciculi in which myelinated nerve fibres are prominent. These fibres range in diameter between 0.2 and 1.8 (mean 0.7 ) consistent with conduction velocities averaging 0.6 m/sec recorded in the carp lateral olfactory tract.The medial division of the olfactory tract contains two larger fasciculi within which are numerous fine unmyelinated nerve fibres (mean diameter 0.17 ) arranged in bundles partly enveloped by glial cell processes. Myelinated nerve fibres are unevenly distributed within both fasciculi and have mean diameters of 0.6 .An interesting observation is the consistent presence of synapses within the largest bundle of the medial tract at all levels.Supported by Grant 5 Ro5 TW00154-03 from the National Institutes of Health, United States Public Health Service.The authors are indebted to the Fisheries and Wildlife Department who generously provided the fish from Snob's Creek Fish Hatchery, and gratefully acknowledge the technical assistance of Mr. T. Armitage, Mr. J. Simmons and Miss D. Harrison.     

Calcium-stimulated respiration and active calcium transport in the isolated chick chorioallantoic membrane

Summary Calcium markedly stimulates the respiration of the isolated chick chorioallantoic membrane. This stimulation of oxygen uptake appears to be closely associated with the membrane's active transcellular calcium transport mechanism. In the presence of 1mm Ca⁺⁺ the rate of uptake increases from 9.3±0.15 to 13.0±0.2 liters O₂/cm²/hr, an increase of about 40%. The calcium-stimulated respiration is specific for the ectodermal layer of cells, the known location of the calcium transport mechanism, and only occurs when the calcium transport mechanism is operative. Sr⁺⁺ and Mn⁺⁺ are transported by the tissue at a lower rate than Ca⁺⁺ and cause a smaller stimulation of oxygen consumption. Mg⁺⁺ and La³⁺ have no effect on tissue respiration. In the presence of Ca⁺⁺, the organic mercurialp-chloromercuribenzene sulfonate (PCMBS) inhibits calcium transport and specifically decreases the oxygen uptake of the ectoderm to a rate identical to that obtained in a calcium-free medium. Stripping the inner shell membrane away from the chorioallantoic membrane mimics these effects. The specificity and locus of action of these two inhibitors suggest that a vital component of the active transcellular calcium transport mechanism resides on or near the outer surface of the plasma membrane of the ectodermal cells and that sulfhydryl groups are important to the normal function of this component.     

The fine structure of the olfactory mucosa and nerve in the teleost Carassius carassius L.

Summary Morphological studies on teleost olfactory mucosa confirm the findings of previous authors regarding the general arrangement of conventional cell types, viz. receptor, sustentacular, mucous and basal, but teleosts show certain distinct differences. The receptor cells have the general mammalian bipolar shape but their peripheral dendrite does not project beyond the epithelial surface. In addition to numerous typical cilia, an exceptional ciliary formation was observed in which the filaments, instead of forming individual cilia, are grouped together in clusters and are enveloped in a single limiting membrane.At the junction between the finger-like process and the mucosal fold myelinated nerve fibres are observed within the subepithelial stroma.Within the postero-medial zone of the mucosa is a conspicuous well-differentiated new cell type. A thick rim of electron-dense cytoplasm, bounded by an outer trilaminar membrane, encloses prominent foliate (leaf-like) organelles, a basal nucleus, numerous mitochondria and vacuolar spaces. These foliaceous cells communicate with the external environment through a small stoma, their close association with epithelial components suggesting a possible secretory or absorptive function. Their intricate morphology, however, suggests that they may be receptors, but their role and neural connections still require definition.Supported by Grant 5 RO 5 TWOO 154-02 from the National Institutes of Health, United States Public Health Service.The authors are indebted to Dr. A. S. Wilson for his helpful criticism and gratefully acknowledge the photographic technical assistance of Mr. J. Simmons and Mr. S. Frank.     

The fine structure of cephalopod blood vessels II. The vessels of the nervous system

Summary Blood vessels of the perioesophageal nerve ganglia (brain) of Octopus vulgaris and the stellate ganglia of Sepia officinalis are described. The vessels have an incomplete endothelium, a complete basement membrane and a complete investment of pericytes. The pericytes are joined by specialised membrane junctions but these are not tight junctions. The main type of neuron/vessel arrangement is one where there is a collagen-filled space between the pericytes and the surrounding glial cells. Axons or neurons are sometimes applied directly to the vessel pericytes and in the neuropil, pericytes contact glial cells that ensheath bundles of axons. Blood spaces between neurons are also present.We would like thank Professor J. Z. Young and Dr. E. G. Gray for encouragement and advice, Mrs. Jane Astafiev for drawing Fig. 11, Mr. S. Waterman for photographic assistance and Miss Cheryl Martin for secretarial and other assistance.     

The yield of synaptosomes from the cerebral cortex of guinea pigs estimated by a polystyrene bead “tagging” procedure

Summary The number of synaptosomes (pinched-off nerve endings) produced/g guinea pig cortex on homogenizing this tissue under defined conditions is estimated to be in the region of 4×10¹¹ using two different polystyrene bead tagging procedures. This is the same order of magnitude as the number of nerve endings/g cortex calculated from histological estimates given in the literature of the number of neurones in the cortex and the extent of their cortical connexions.This investigation was supported by U.S. Public Health Service Grant no. NB 03928 from the National Institute of Neurological Diseases and Blindness. Dr. Sheridan was a Postdoctoral fellow of the U.S. Public Health Service during the period of his participation in this investigation (1963/64). The electron microscope facilities were provided by the Wellcome Trust. Mr. T.F.J. Hobson, of the ARC Statistics group, Cambridge, kindly advised on statistical aspects of the work. We are most grateful to him, to Miss L. Swales and Mr. G.H.C. Dowe for their skilled technical assistance and to Dr. E.G. Gray for helpful discussions.     

The behavior of retinal tissue in vitro,light and electron microscopic observations

Summary Retinae from two day old rats were used in this study and the cultures were handled according to standard methods used in this laboratory. In the first few days of cultivation an abundant outgrowth of nerve fibers into the cell-free medium was observed. These fibers later degenerated and by the beginning of the second week they had completely disappeared. In the living cultures, differentiating ganglion cells, bipolar and horizontal neurons could be seen in the main explant in association with various types of glial cells. Rod cells became arranged as epithelial sheets or as clusters of cells which often formed rosettes. The nuclei of these sensory cells possessed a characteristic chromatin pattern by which they always could be differentiated from other cells in the cultures. Cytoplasmic extensions that developed from the free surface of the sensory rod cells were observed within a week following explantation. A limiting membrane separated these extensions from the nucleated part of the rod cells. Morphologic details of the different neuronal cell types could be demonstrated in cultures by Bodian's silver impregnation technique.With the electron microscope, retinal development in culture was observed and compared to the development of the retina of the intact eye. Cilia developed from processes extending from the rod cell free surface. These processes were the rod cell inner segments in which many mitochondria were seen. At the bases of these segments terminal bars developed forming the outer limiting membrane. In the area of the terminal bars microvillous extensions projected between the rod cell inner segments. After twelve days in vitro a bulb-like enlargement containing a lamellar membrane system developed at the end of the cilium. This bulb-like enlargement was a beginning of the rod cell outer segment. The lamellar system did not acquire the symmetry or precise organization during cultivation that was observed in the retina of the intact eye. The distinguishing characteristics of individual neuronal cell types seen in cultivated retinae were the same as those described for their counterparts in the retina in situ, but regular plexiform layers failed to develop. Likewise, there were no indications of typical synapses in the neuropils of the cultures. There were many processes containing vesicles similar to those in presynaptic endings and mitochondria but membrane thickenings were not apparent.The results indicate that the retina cultivated in vitro does not behave as an organized entity. The component cells dissociated more and more with time, and developmental differentiation was observed only at the cellular level.Supported by USPHS Grants 5R01NB03114-06 and 5T01GM00459 from the National Institutes of Health, Bethesda, Maryland.Sincere appreciation is expressed to Mrs. Eleanor Morris for management of the cultures, and to Mr. E. E. Pitsinger, Jr. for his photographic assistance.     

An electron microscopical study of the ventral nerve cord of the leech

Summary Three types of fibrillar structure can be seen with the electron microscope in nerve cells of the vental nerve cord of the leech: the neurofibrillar bundles, the tubules and the tonofibrils. In neuroglial cells only the tonofibrils are present. The three types are structurally distinct, and, contrary to past suggestions, there is no evidence that neurofibrillar bundles may consist of tightly packed or badly fixed tubules.In vertebrates the electron microscope reveals bundles of discrete neurofilaments that form the basis for the argyrophilic neurofibrillae seen by light microscopy. Each neurofilamentous unit appears as a dot in cross section. In contrast, in the leech, the electron microscope shows compact fibrillar bundles that clearly correspond to the neurofibrils described by light microscopists. These bundles are made up of closely packed units rather than discrete filaments and where the units occur singly they are seen to have an angular or stellate outline in cross section. To make this distinction clear these have been termed neurofibrillar bundles rather than neurofilaments.Attachment plaques occur in both neurons and neuroglia. These plaques have tonofibrils attached, and the glial tonofibrils are far more numerous than the neuronal tonofibrils. The glial fibrils are identical with the tonofibrils in the glial cells.The attachment plaques are invariably related to an extracellular space that contains material identical with the basement membrane. This material is continuous, by a complex system of channels and diverticulae, with the outer basement membrane in the neuron packets, but forms isolated patches in the other parts of the nervous system.We are grateful to Prof. J. Z. Young, F. R. S., for his encouragement to Mrs. Astafiev for the drawings, to Miss B. Shirra and Mr. K. Watkins for technical assistance and to Mr. S. Waterman for photography.     

Changes of lysosomal enzymes during hereditary degeneration and histogenesis of retina in mice

Summary In the normal histogenesis of mouse retina localized distribution of acid phosphatase positive granules has been seen around the photoreceptor cell nuclei along the outer limiting membrane. These granules disappear during the development of the rod elements. Temporarily increased activity is also seen along the nuclei of the inner layer adjacent to and in the course of the development of the outer and the inner plexiform layers. Within the inner nuclear layer, the cells at the outer and inner rows develop localized acid phosphatase positive granules which persist in the adult retina. Ganglion cells and the layer of nerve fibres show little change. In the pigment epithelium the enzyme gradually increases. In mice, homozygous for the retinal degeneration gene, degenerating photoreceptor cell nuclei, characterized by perinuclear acid phosphatase staining, can be detected before morphological signs of degeneration. Increased frequency of such nuclei and intensity of staining are recorded with the progress of degeneration. Enzyme activity in the photoreceptor cells, within the inner nuclear layer and in the degenerating photoreceptor cell nuclei is demonstrable using naphthol substrates but not -glycerophosphate. Positive reaction with -glycerophosphate is obtained in these sites in the presence of dimethyl sulphoxide. Existence of differential permeability among the retinal lysosomes is tentatively suggested.     

The origin of annulate lamellae in the oocyte of the ascidian,Boltenia villosa stimpson

Summary The formation of the extranuclear annulate lamellae has been revealed to be connected with a process of nuclear emission which is very active during the previtellogenetic stages of the Boltenia oocyte development. This process involves both of the nuclear membranes. At many spots on the surface of the nuclear envelope, the outer membrane pulls away from the inner membrane, thus forming what has been designated as blisters of various sizes and shapes. Masses of nuclear content, apparently not from the nucleolus, are pushed into the blisters. These blisters may become detached from the nuclear envelope and lie free in the cytoplasm. But in many cases, the detachment seems delayed, and in each blister many emission masses are squeezed tightly together and flat one on top of the other. These masses, in sections, may present the appearance of a stack of elongated outlines. The membrane, limiting any two adjacent masses in close contact, develop annuli. It is thus that an annulate lamella is formed. Whether an annulate lamella is formed between a pair of neighboring masses depends on their proximity. So the production of the annulate lamellae is incidental to, but not a necessary part of the process of nuclear emission. After the original outer nuclear membrane forming the blister has disintegrated, the annulate lamellae are left exposed in the cytoplasm.It is clear that, 1. both membranes of an annulate lamella are of inner nuclear membrane origin, 2. they hold between them some of the content of the enlarged perinuclear space resulting from the raising of the outer nuclear membrane when the blister is formed, and 3. the material held between any two lamellae is from the nucleus.The intranuclear annulate lamellae simply arise from the narrow pouches formed by the inner nuclear membrane towards the interior of the nucleus, and on these narrow pouches annuli are developed. So the intranuclear annulate lamellae is also composed of two membranes of an inner nuclear membrane origin holding between them a quantity of the content of the perinuclear space.Supported by Grant GM-11858 of National Institute of Health. The author is indebted to Dr. Richard Cloney of the Department of Zoology, University of Washington, for the use of the electron microscope.     

The mineralization and hardness of the radular teeth of the limpet Patella vulgata L.

Summary A study of the Patella vulgata radula has been made using: the scanning electron microscope in its normal and compositional contrast modes of operation, the electron microprobe analyser, ion etching with argon ions and microhardness testing.Only iron, silicon and small amounts of sulphur were detected in the radula. The teeth can be subdivided into a cusp, a junctional area where the cusp is joined to the base, and the base which is embedded in the radular membrane. From a study of longitudinal vertical and transverse sections of the mature teeth it was found that the cusp could be subdivided into a posterior iron-rich area (44–51% Fe, 1–6% Si) and an anterior silicon-rich area (22–30% Fe, 27–32% Si). The junctional zone consisted of a poorly mineralised layer at its border with the cusp and an iron-rich layer where it joined the base. The upper part of the base (5% Fe, 16% Si) could be clearly differentiated from the silicon-rich anterior and lower parts of the base (3–4% Fe, 28–35% Si). No minerals were detected in the membrane. The changes in the mineral content of the teeth cusps along the length of the radula were studied. Iron appeared in the cusps at the 25th row and the concentration increased to 28% at the 50th row. The iron was here evenly distributed throughout the cusp. Silicon appeared in the anterior part of the cusp at the 50th row and as it increased in concentration so the iron was displaced, and at the same time the concentration of iron increased in the posterior part of the cusp. Mineralization appeared to be complete by the 150th row.The teeth cusps appear to consist of 800 Å fibres grouped into 1 thick bundles and the tooth appears to be covered by a thin enamel-like layer. It is suggested that the fibres contain the silicon-rich phase and the matrix the iron-rich phase.The significance of the arrangement of the fibres and the distribution of the minerals are discussed with relation to the function of the teeth.We wish to thank Mr. A. Rees and Mr. A. Davies for their technical assistance; Prof. Lewis and Dr. James for the use of the Electron Microprobe; and the S.R.C. for their financial support.     

Sulla organizzazione dello strato granulare esterno e della membrana limitante esterna nella retina di coniglio

Summary The organisation of the outer nuclear layer and the structure of the outer limiting membrane of rabbit retina have been studied. In specimens stained by the Golgi method it was observed that in the outer nuclear layer each Müller cell envelops with its thin lamellar expansions ten to fifteen rod and cone cell bodies.The only cytoplasmic organelles in rod and cone cell bodies are a few free ribosomes and smooth surfaced vesicles. Neurotubules are prominent in the outer and inner fibres of the rods and cones.The processes of the Müller cells are distinctive because of the presence of many glycogen granules and glial filaments. Also present but only found near the outer limiting membrane are mitochondria, occasional centrioles and cilia that lack inner fibres. Long microvilli originate from the Müller cell processes on the scleral side of the outer limiting membrane.The photoreceptor cells on the vitreal side of the outer limiting membrane are completely isolated from each other by glial processes. On the scleral side of the membrane, the inner segments of the photoreceptor cells are not completely isolated by glial processes and so are frequently found in mutual contact. In the outer nuclear layer the granule of each photoreceptor is surrounded by more than one glial process while the fibres are often deeply embedded in a single glial process and provided with a mesofibre.At the level of the outer limiting membrane the visual cells and the glial expansions enveloping them are joined together by a junctional complex formed by a zonula adhaerens interposed between two very short zonulae occludentes. The same junctional complex joins to each other the contiguous expansions of the Müller cells and the mesofibres of the visual elements.     

Oxygen and CO2 Exchange and Acid-Base Regulation in the Avian Embryo

The avian embryo exchanges the oxygen and carbon dioxide withthe ambient air by diffusion. The respiratory organ is the chorioallantois,endowed with a rich circulation. Between ambient air and chorioallantoiccapillary blood are interposed the porous shell fibrous shellmembranes, and the chorioendothelium which compose the diffusionbarrier. The air cell is formed between the two shell membranesin the blunt end of the egg. The diffusion barrier is dividedinto an outer barrier (shell plus outer membrane) and an innerbarrier (inner membrane plus chorioendothelium and capillaryblood). The resistance to gas diffusion (the reciprocal of thediffusive conductance) in the outer barrier is almost fixedthroughout incubation while that in the inner barrier decreasesas the embryo develops. Because of the fixed outer barrier conductance,the embryo is obliged to take up oxygen under hypoxic conditionsagainst increasing metabolism with development and encountersa relative respiratory acidosis. In connection with the diffusivehypoventilation caused by the fixed outer barrier conductancethe respiratory factors of the allantoic circulation changeprogressively with development to moderate the restraint ofgas exchange through the shell. Blood oxygen capacity and hemoglobinincrease with development in association with an increase inerythrocyte count and hematocrit value. In addition, a progressiveleftward shift of the oxygen dissociation curve occurs. Theincreases in the allantoic blood flow and chorioallantoic capillaryvolume contribute to the increasing conductance of the innerbarrier. Furthermore regulation of acid base balance is inferredin the developing embryo.     

Ultrastructure of spermatozoa of Peregrinus maidis (Homoptera,Delphacidae)

Summary The spermatozoa of Peregrinus maidis Ashm. are thread-like, approximately 650 long and 1 wide including the head (approximately 28 ).The main part of the spermatozoa consists of two mitochondria derivatives, a central body between them, the axial filament complex, and a newly found element consisting of two wing-shaped bodies. Each mitochondrion derivative shows a peripheral and an inner part. The peripheral part is formed by cristae arranged perpendicularly to the long axis of the spermatozoon. The cristae are approximately 70 Å wide. The dense layers between them measure approximately 280 Å. The inner part of the mitochondrion derivative shows a crystalline array, formed by sub-units of approximately 100 Å diameter. The wing-shaped bodies consist of tubular elements.The head has an elongated nucleus with an electron transparent space inside. At the anterior end of the nucleus lies a tapered acrosome. This appears fibrous and parts of the acrosome fibers seem to run along the nucleus. Acknowledgements. The authors wish to thank Dr. G. H. Bergold for suggestions and support, Drs. J. André, D. W. Fawcett, P. Maillet and G. F. Meyer for very helpful discussion. They are also grateful to Mr. O. Suárez for assistance in the preparation of the organs of P. maidis and to Mrs. M. de Pingarrón for technical assistance.     

Placentation in the lizard Gerrhonotus coeruleus with a comparison to the extraembryonic membranes of the oviparous Gerrhonotus multicarinatus (Sauria,Anguidae)

Paraffin sections of an ontogenetic series of embryos of the viviparous lizard Gerrhonotus coeruleus and the oviparous congener G. multicarinatus reveal that although general features of the development of the chorioallantoic and yolk sac membranes are similar, differences are evident in the distribution of the chorioallantoic membrane in late stage embryos. An acellular shell membrane surrounds the egg throughout gestation in both species although the thickness of this structure is much reduced in G. coeruleus over that of G. multicarinatus. The initial vascular membrane to contact the shell membrane in both species is a trilaminar omphalopleure (choriovitelline membrane) composed of ectoderm, mesoderm of the area vasculosa, and endoderm. This transitory membrane is replaced by the vascularized chorioallantois as the allantois expands to contact the inner surface of the chorion. Prior to the establishment of the chorioallantois at the embryonic pole, a membrane begins to form within the yolk ventral to the sinus terminalis. This membrane, which becomes vascularized, extends across the entire width of the abembryonic region and isolates a mass of yolk ventral to the yolk mass proper. The outer membrane of the yolk pole is a nonvascular bilaminar omphalopleure (chorionic ectoderm and yolk endoderm). In G. multicarinatus the bilaminar omphalopleure is supported internally by the vascularized allantoic membrane, whereas in G. coeruleus the allantois does not extend beyond the margin of the isolated yolk mass and the bilaminar omphalopleure is supported by the vascularized intravitelline membrane. Both the chorioallantoic placenta (uterine epithelium, chorionic ectoderm and mesoderm, and allantoic mesoderm and endoderm) and the yolk sac placenta at the abembryonic pole (uterine epithelium, chorionic ectoderm, and yolk sac endoderm) persist to the end of gestation in G. coeruleus.     

Two substrate sites in the renal Na+-d-glucose cotransporter studied by model analysis of phlorizin binding and-d-glucose transport measurements

Summary Time courses of phlorizin binding to the outside of membrane vesicles from porcine renal outer cortex and outer medulla were measured and the obtained families of binding curves were fitted to different binding models. To fit the experimental data a model with two binding sites was required. Optimal fits were obtained if a ratio of low and high affinity phlorizin binding sites of 1:1 was assumed. Na⁺ increased the affinity of both binding sites. By an inside-negative membrane potential the affinity of the high affinity binding site (measured in the presence of 3 mM Na⁺) and of the low affinity binding site (measured in the presence of 3 or 90 mM Na⁺) was increased. Optimal fits were obtained when the rate constants of dissociation were not changed by the membrane potential. In the presence of 90 mM Na⁺ on both membrane sides and with a clamped membrane potential,K _D values of 0.4 and 7.9 M were calculated for the low and high affinity phlorizin binding sites which were observed in outer cortex and in outer medulla. Apparent low and high affinity transport sites were detected by measuring the substrate dependence ofd-glucose uptake in membrane vesicles from outer cortex and outer medulla which is stimulated by an initial gradient of 90 mM Na⁺(out>in). Low and high affinity transport could be fitted with identicalK _m values in outer cortex and outer medulla. An inside-negative membrane potential decreased the apparentK _m ofhigh affinity transport whereas the apparentK _m of low affinity transport was not changed. The data show that in outer cortex and outer medulla of pighigh and low affinity Na⁺-d-glucose cotransporters are present which containlow and high affinity phlorizin binding sites, respectively. It has to be elucidated from future experiments whether equal amounts of low and high affinity transporters are expressed in both kidney regions or whether the low and high affinity transporter are parts of the same glucose transport moleculc.     

Localization of melanin synthesis within the pigment cell: Determination by a combination of electron microscopic autoradiography and topographic planimetry

Summary Localization of melanin synthesis within the pigment cells of the Cloudman S-91 mouse melanoma was determined by means of a combination of high resolution autoradiography and topographic planimetry. Initial melanin biosynthesis occurred predominantly in the endoplasmic reticulum and associated ribonucleoprotein particles of the melanocytes. By measuring a number of cell organelles and employing the index of relative specific localization it could be shown that the nucleus and mitochondria are of little or no importance in the process of melanogenesis.This investigation has been supported by the following research grants: CA 06548 CB, NIH, PHS and an Institutional Grant from the American Cancer Society (to H. M. H.); CA-05887, NIH, PHS (to A. S. Z.); M-00388 and NB-00782, NIH, PHS (to J. F. H.). One of us (H. M. H.) holds a Research Cancer Development Award (5-K 3-GM-2634) of the National Institute of General Medical Sciences, Public Health Service.We are grateful to Mr. Ronald Abler for his help with the topographic measurements; to Dr. Erhard Haus for help and advice; to Mr. J. Thornby and Mr. A. P. Basu for assistance with the statistical aspects of this study; and to Mrs. Lenore Mottaz, Miss Bernice Uittenbogaard, and Mrs. Judith Strong for careful technical assistance.