Morphology and distribution of chromaffin cells in the adrenal gland of cordylidae (Reptilia,Sauria): A comparative study

The distribution of the adrenaline and noradrenaline chromaffin cells in the adrenal glands of 10 members of the family Cordylidae have been examined. In the genus Gerrhosaurus, all the catecholamine cells lie on the surface of the adrenal gland, forming a continuous envelope of one or two layers of cells that mainly contain noradrenaline (NA). In the genus Platysaurus, the chromaffin envelope is intermittent. There are relatively large tracts of interspersed interrenal tissue containing some adrenaline cells (A). Islets of chromaffin cells are scattered between these interrenal tracts. In the genus Pseudocordylus and the genus Cordylus, the superficial chromaffin cells tend to gather into a multilayered dorsal mass, containing mainly NA cells. Inside the interrenal parenchyma, there are always numerous chromaffin islets, containing mainly A cells.     

Chromaffin cells and interrenal tissue in the head kidney of the grouper,Epinephilus tauvina (Teleostei,Serranidae): a morphological (optical and ultrastructural) study

This work analyses the distribution, histology and ultrastructure of chromaffin cells (CCs) and interrenal tissue (It) in the head kidney of Epinephilus tauvina. Histological examination revealed that chromaffin cells are found in small groups under the endothelium of the posterior cardinal vein (PCV) and are mostly closely associated with the interrenal tissue. Ultrastructure examination confirmed the existence of two main chromaffin cell types, distinguished by different types of secretory granules. The first type was characterized by the presence of vesicles with round, strongly electron dense core granules, which were eccentrically located. Such cells were interpreted as being noradrenaline cells. Meanwhile, cells with vesicles that were completely electron lucent or that contained small less dense eccentric granules were identified as adrenaline cells. Nerve endings were invaginated into the chromaffin cells through synaptic junctions. Interrenal tissue consisted of nests, cords, or strands of cells in contact with the posterior cardinal vein (PCV) and interposed with haematopoietic tissue. Ultrastructure analysis revealed only one interrenal cell type, which contained abundant smooth endoplasmic reticulum (sER) and numerous mitochondria with tubulo‐vesicular cristae, characteristics of steroid‐producing cells. The interrenal tissue cells have different cytological aspects that can be linked to a steroidogenic cell cycle allowing a periodical renewal of organelles.     

Electron microscopic localization of 3β-hydroxysteroid dehydrogenase and nadh-ferricyanide reductase activities in amphibian interrenal cells

Summary 3-hydroxysteroid dehydrogenase and NADH-ferricyanide reductase activities were localized at the ultrastructural level in amphibian interrenal (adrenocortical) cells previously fixed in a mixture of formaldehyde and glutaraldehyde. Potassium ferricyanide was used as an electron acceptor.Copper ferrocyanide deposits resulting from 3-HSD activity were seen in close association with the external faces of the membranes of the smooth endoplasmic reticulum. Very rare grains of precipitate appeared in mitochondrial cristae. The addition of phenazine methosulfate to the incubation medium had no effect on these localizations.The interrenal cells showed also a strong NADH-ferricyanide reductase activity. The copper ferrocyanide grains were abundant in the mitochondrial cristae and in the hyaloplasm, where they were not preferentially associated with the smooth endoplasmic reticulum.The author is grateful to Miss Marie-Eve Moritz for skillful technical assistance     

Functional Morphology of Steroidogenic Tissue and Adrenomedullary Cells in the Adrenal Gland of Rana tigrina (Daud.) and Rana cyanophlyctis (Schn.)

The presence of δ⁵-3β-HSDH, 17β-HSDH, 11β-HSDH, G-6-PDH and DPNH-diaphorase activity has been demonstrated in the interrenal cells of two frogs, R. tigrina and R. cyanophlyctis. The substrate specificity of δ⁵-3β-HSDH and 17β-HSDH was tested by utilizing different specific hydroxysteroids. DL⁵-3β-HSDH, G-6-PDH and NADH-diaphorase activity was relatively more in the peripheral region of the interrenal tissue compared to the cells in the middle region, apparently indicating a zonation of the adrenocortical tissue. The presence of 11β-HSDH, G-6-PDH and NADH-diaphorase has been also observed in the renal tubules which indicates that the renal tubules might convert hydroxysteroids to ketosteroids during steroid excretion. The presence of two types of adrenal medullary cells showing positive iodate and chromate reactions was observed.     

Steroid dehydrogenases in the adrenal gland of four species of birds: a histochemical study

Synopsis The presence of⁵-3-hydroxysteroid dehydrogenase, II-hydroxysteroid dehydrogenase, 17-hydroxysteroid dehydrogenase and glucose-6-phosphate dehydrogenase has been demonstrated histochemically in the adrenal gland of the rain quailCoturnix coromendalica, barn owlTyto alba, brown crakeAmaurornis akool and painted partidgeFrancholinus pictus. All these enzymes occurred in the inter-renal cells. No activity was observed in the chromaffin cells. It is suggested that the inter-renal cells of these four species of birds are capable of synthesizing both corticosteroids and sex steroids.     

A light- and electron-microscopic investigation of gametogenesis in Typosyllis pulchra (Berkeley and Berkeley) (Polychaeta: Syllidae)

Summary The ultrastructure of the interrenal (adrenocortical) cells of trout (Salmo fario L.) was studied after dexamethasone treatment. A procedure for identifying and isolating interrenal tissue fragments from the surrounding head kidney tissue prior to their preparation for electron microscopy is described. The peripheral plasma cortisol concentrations were measured in order to evaluate the steroidogenic activity of this tissue.The interrenal cells of control animals contain numerous mitochondria with tubular cristae, and a well developed and highly organized smooth endoplasmic reticulum (SER). The scarcity, or absence, of lipid droplets contrasts markedly with the abundance of SER.Treatment with dexamethasone results in a decrease steroidogenic activity of the interrenal cells, as indicated by the fall in plasma cortisol concentrations. The interrenal cells are small, but still contain numerous mitochondria. The SER is poorly developed, but masses of densely intermeshed smooth cisternae subsist. Lipid droplets do not accumulate in these cells; this peculiarity is discussed in connection with the virtual absence of liposomes in teleost interrenal cells.     

Histophysiologische Untersuchungen der Entwicklung des Interrenalorgans beim Krallenfrosch (Xenopus laevis Daudin)

Zusammenfassung Mit histologischen und histochemischen Methoden lassen sich bei Xenopus-Larven 4 Zonen des interrenalen Gewebes (Nebennieren-Rinden-Gewebes) in caudo-cranialer Ausdehnung unterscheiden, die verschiedenen Phasen der Nebennierenentwicklung entsprechen. Diese Zonen wurden auf Grund der in ihnen am häufigsten auftretenden Zelldifferenzierungsformen als Blastemzone, Zone der Organbildung, Zone der Ausdifferenzierung und Zone der Funktionsphasen bezeichnet. Während der Entwicklung verändert sich ihre Ausdehnung, sie verschieben sich von cranial nach caudal. Daneben kommt es während der Ausdifferenzierung zu einer Verlagerung der Interrenalzellen von dorsal nach ventral. Die ausdifferenzierten Interrenalzellen durchlaufen einen Funktionszyklus. Charakteristische Differenzierungformen sind hierbei: wenig stimulierte, stark stimulierte, überstimulierte Interrenalzellen sowie Zellen in der Restitutionsform und Degenerationsform.Kerngrößenmessungen und Bestimmungen der Steroiddehydrogenase-Aktivität an Normaltieren zeigen eine Abhängigkeit der Nebennierenaktivität von dem Entwicklungsstadium. Maxima der Aktivität liegen im Stadium 59 und am Ende der Metamorphose vor.Untersuchungen des Organes nach Hypophysektomie und/oder ACTH-Behandlung machen den Einfluß der Hypophyse auf die Normalentwicklung der Nebenniere deutlich. Hypophysektomie verhindert zwar nicht die Abgliederung der Nebenniereninseln vom Blastem, die Zellen differenzieren sich jedoch nicht vollständig aus, sondern verharren in einer funktionellen Ruhepause. ACTH-Behandlung beschleunigt die Differenzierung und funktionelle Entwicklung der Interrenalzellen und führt zu einer Vergrößerung der einzelnen Interrenalkomplexe. Sie kompensiert auch die hemmende Wirkung der Hypophysektomie auf die Nebenniere. Außerdem vermag ACTH die durch Hypophysektomie bewirkte Wachstums-und Entwicklungshemmung teilweise auszugleichen.

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Histophysiological investigations of the development of the interrenal organ in the clawed toad (Xenopus laevis Daudin)

Summary Histological and histochemical investigations have given evidence for 4 zones of the interrenal tissue in caudo-cranial direction in Xenopus-larvae. These zones correspond to 4 phases of the interrenal development. Due to the representative differentiation form of the interrenal cells they were characterized as: 1) zone of the blastema, 2) zone of the newly formed interrenal organ, 3) zone of differentiation and 4) zone of the various functional stages. During the development these zones shift from cranial to caudal. Besides there is a displacement during the differentiation of the interrenal cells in dorso-ventral direction. Differentiated interrenal cells undergo a functional cycle: hardly stimulated, clearly stimulated and overstimulated adrenocortical cells as well as cells in the state of restitution and degeneration are to be observed.Calculation of the mean nuclear diameter and the activity of the steroiddehydrogenase in normal animals demonstrate a correlation between the activity of the adrenal cortex and the stage of development. Maxima of this activity are found in stage 59 and at the end of metamorphosis. Experiments with hypophysectomy and/or ACTH-treatment clearly demonstrate the influence of the pituitary on the adrenocortex during normal development. Hypophysectomy does not prevent the separation of interrenal islets from the blastema. These cells, however, do not completely differentiate but stay in an inactive phase of their function. ACTH-treatment accelerates the differentiation and functional development of the interrenal cells and further results in an enlargement of the single adrenocortical islets. This treatment compensates the inhibition of the adrenal cortex after hypophysectomy and partly compensates the retardation of development and growth in hypophysectomized animals.

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Mit dankenswerter Unterstützung durch die Deutsche Forschungsgemeinschaft.     

Co-localization of vasoactive intestinal peptide (VIP) and enkephalins in chromaffin cells of the adrenal gland of amphibia. Stimulation of corticosteroid production by VIP

Recent studies have shown that biologically active peptides and monoaminergic neurotransmitters coexist in certain neuronal cell populations. Using the immunofluorescence technique, we have examined the localization of enkephalins, vasoactive intestinal peptide (VIP) and tyrosine hydroxylase in the adrenal gland of the frog Rana ridibunda. Most chromaffin cells which stained for tyrosine hydroxylase contained VIP-like immunoreactivity, whereas methionine- (Met-) and leucine- (Leu-) enkephalin-like immunoreactivity was detected in about 40% of the cells revealed by the anti-tyrosine hydroxylase serum. No VIP- or enkephalin-like immunoreactive nerve fibres were observed. Since in the frog, the chromaffin cells are in close contact with the adrenocortical (interrenal) tissue, a possible action of VIP and opiates on corticosteroidogenesis has been investigated. At doses 10(-6) and 10(-5) M, 20-min infusions of synthetic porcine or chicken VIP elicited a significant increase in corticosterone and aldosterone production by perifused frog adrenals, in a dose-dependent manner. As compared to ACTH, VIP was several orders of magnitude less effective in stimulating corticosteroid production. Morphine, Met- and Leu-enkephalins (10(-5) M) had no effect on spontaneous secretion of corticosteroids. In addition, Met- and Leu-enkephalins (10(-5) M) did not alter the production of corticosterone induced by ACTH. THese results suggest that VIP contained in the chromaffin cells of the frog adrenal gland may exert a local action in stimulating corticosteroid production by the interrenal tissue.     

Distribution and Structure of the Adrenocortical Homolog in Polypterus palmas Ayres

The identity, distribution and structure of the adrenocortical homolog (AH) was made in Polypterus palmas Ayres using routine light and electron microscopy and histochemistry for the enzyme δ⁵-3β-hydroxysteroid dehydrogenase (3β-HSD). The AH is confined to yellow corpuscles which are positioned near the posterior cardinal and renal veins in the anterior two-thirds of each kidney. The 46–57 spherial to cigar-shaped corpuscles are placed end to end and are equally distributed in the kidneys. The tortuous cords of epithelial cells of each corpuscle are surrounded by sinusoids and are completely delimited from the haemopoietic and renal tissue of the kidneys. The steroidogenic nature of the cells is demonstrated by their 3β-HSD activity and their ultrastructure, namely lipid droplets, tubular smooth endoplasmic reticulum and mitochondria with tubulovesicular cristae. Giant mitochondria and gap junctions are notable features of AH cells in this fish. The yellow corpuscles of the kidneys in P. palmas represent the AH and this tissue has a distribution which is in accordance with the taxonomic position of Polypteriformes.     

Characterization of cDNAs encoding cholesterol side chain cleavage and 3beta-hydroxysteroid dehydrogenase in the freshwater stingray Potamotrygon motoro

The interrenal gland (adrenocortical homolog) of elasmobranchs produces a unique steroid, 1α-hydroxycorticosterone (1α-B). The synthesis of this and most other steroids requires both cholesterol side chain cleavage (CYP11A) and 3β-hydroxysteroid dehydrogenase (HSD3). To facilitate the study of elasmobranch steroidogenesis, we isolated complementary DNAs encoding CYP11A and HSD3 from the freshwater stingray Potamotrygon motoro. The P. motoro CYP11A (2182 bp total length) and HSD3 (2248 bp total length) cDNAs harbor open reading frames that encode proteins of 542 and 376 amino acids (respectively) that are similar (CYP11A: 39–61% identical; HSD3: 36–53% identical) to their homologs from other vertebrates. In molecular phylogenetic analysis, P. motoro CYP11A segregates with CYP11A proteins (and not with related CYP11B proteins) and P. motoro HSD3 segregates with steroidogenic HSD3 proteins from other fishes. CYP11A and HSD3 mRNA is found only in interrenal and gonadal tissues, indicating de novo steroidogenesis is restricted to these tissues. Because 1α-B is thought to act in the elasmobranch response to hydromineral disturbances, we examined the effect of adapting P. motoro to 10 ppt seawater on mRNAs encoding steroidogenic genes. The P. motoro response to this salinity challenge does not include interrenal hypertrophy or an increase in the levels of interrenal CYP11A, HSD3 or steroidogenic acute regulatory protein (StAR) mRNA. This study is the first to isolate full length cDNAs encoding elasmobranch CYP11A and HSD3 and the first to examine the regulation of steroidogenic genes in elasmobranch interrenal cells.     

Seasonal studies on the adrenal homologue of female Puntius sophore (Ham.).

In female P. sophore the adrenal homologue is located inside its head kidneys and comprises interrenal cells and chromaffin cells. The clumps of interrenal cells surround the postcardinal vein and its major branches; they also lie in the haemopoietic tissue. The chromaffin cells mostly lie in the endothelium of the post-cardinal vein, while a few of them also lie interspersed among the interrenal cells. The seasonal study of the adrenal homologue shows that the interrenal cells undergo changes in bulk and composition in different months, while the chromaffin cells do not show any significant variation.     

Action of vasoactive intestinal peptide (VIP) on amphibian adrenocortical function,in vitro

Vasoactive intestinal peptide (VIP) is located in chromaffin cells of the frog adrenal gland and is able to stimulate corticosteroid secretion in amphibians. In the present study we have investigated the possible involvement of prostaglandins, microfilaments and calcium in the mechanism of action of VIP on frog adrenocortical tissue. Rana ridibunda interrenal dice were perifused with amphibian culture medium for more than 10 hours. Corticosterone and aldosterone concentrations were measured in the effluent perifusate using sensitive and specific radioimmunoassay methods. In the presence of indomethacin (5 μM), a specific blocker of prostaglandin biosynthesis, the spontaneous secretion of corticosteroids was markedly reduced (80%) but the stimulatory effect of VIP was not altered. The administration of the microfilament disrupting agent cytochalasin B (50 μM) inhibited both spontaneous and VIP-induced corticosteroid secretion. In the absence of calcium, the spontaneous level of corticosteroid was reduced to about 60% but VIP was still able to stimulate corticosteroid secretion. From these data we conclude that the integrity of the cytoskeleton is required for the secretory response of adrenocortical cells to VIP, whereas neither prostaglandins nor calcium are involved in VIP-induced adrenocortical stimulation.     

Increase of steroid-producing cells in interrenal tissue and masculinization of gonads after long-term treatment of juvenile rainbow trout with cyanoketone

Summary Cyanoketone administered via the food (0.1, 0.2 and 2 mg/g) for 8 weeks from the first feeding (day 46 after fertilization) or via the aquarium water (3 and 30 mg/100 l) for 4 weeks from day 41 does not influence the activity of 3-hydroxysteroid dehydrogenase (3-HSD) in the interstitial cells of the gonads or interrenal cells of juvenile trout in vivo. However, the number of 3-HSD-positive interrenal cells was strongly increased by administration of the highest dose of cyanoketone via both routes. These high doses furthermore affect the sex ratio in favor of males. It is concluded that interrenal tissue is responsible for the masculinizing effect of cyanoketone via increased production of androgens and/or corticosteroids. Cyanoketone at concentrations of 0.01 to 100 g/ml causes a dose-response inhibition of 3-HSD activity in the interrenal cells, when the substance is administered to an incubation medium for demonstration of this enzyme in tissue sections. The controversial in-vivo and in-vitro effects of cyanoketone on 3-HSD activity are discussed.     

Radioautography of Presumptive Interrenal Cells in the Sea Lamprey After 3H-cholesterol Injection

Abstract Radioautography demonstrates that the injection of ³H-cholesterol into larval and adult sea lamprey, Petromyzon marinus L., results in labelling of their presumptive interrenal cells. This localization of cholesterol and/or its metabolites is: (1) further evidence for the homology of these cells with adrenocortical cells of higher vertebrates and (2) a suggestion of the possibility that these cells in lampreys are able to absorb cholesterol for utilization in corticosteroid synthesis.     

The Hemodynamically-Regulated Vascular Microenvironment Promotes Migration of the Steroidogenic Tissue during Its Interaction with Chromaffin Cells in the Zebrafish Embryo

Background

While the endothelium-organ interaction is critical for regulating cellular behaviors during development and disease, the role of blood flow in these processes is only partially understood. The dorsal aorta performs paracrine functions for the timely migration and differentiation of the sympatho-adrenal system. However, it is unclear how the adrenal cortex and medulla achieve and maintain specific integration and whether hemodynamic forces play a role.

Methodology and Principal Findings

In this study, the possible modulation of steroidogenic and chromaffin cell integration by blood flow was investigated in the teleostean counterpart of the adrenal gland, the interrenal gland, in the zebrafish (Danio rerio). Steroidogenic tissue migration and angiogenesis were suppressed by genetic or pharmacologic inhibition of blood flow, and enhanced by acceleration of blood flow upon norepinephrine treatment. Repressed steroidogenic tissue migration and angiogenesis due to flow deficiency were recoverable following restoration of flow. The regulation of interrenal morphogenesis by blood flow was found to be mediated through the vascular microenvironment and the Fibronectin-phosphorylated Focal Adhesion Kinase (Fn-pFak) signaling. Moreover, the knockdown of krüppel-like factor 2a (klf2a) or matrix metalloproteinase 2 (mmp2), two genes regulated by the hemodynamic force, phenocopied the defects in migration, angiogenesis, the vascular microenvironment, and pFak signaling of the steroidogenic tissue observed in flow-deficient embryos, indicating a direct requirement of mechanotransduction in these processes. Interestingly, epithelial-type steroidogenic cells assumed a mesenchymal-like character and downregulated β-Catenin at cell-cell junctions during interaction with chromaffin cells, which was reversed by inhibiting blood flow or Fn-pFak signaling. Blood flow obstruction also affected the migration of chromaffin cells, but not through mechanosensitive or Fn-pFak dependent mechanisms.

Conclusions and Significance

These results demonstrate that hemodynamically regulated Fn-pFak signaling promotes the migration of steroidogenic cells, ensuring their interaction with chromaffin cells along both sides of the midline during interrenal gland development.     

Histochemical detection of steroid synthesizing cells in the testes of goldfish,Carassius auratus,during the annual reproductive cycle

We investigated the sites of Δ⁵-3β-hydroxysteroid dehydrogenase (3 β-HSD) and glucose-6-phosphate dehydrogenase (G-6-PD) synthesis in the testes of goldfish, Carassius auratus, during the annual reproductive cycle. The histochemistry of fish gonads has been investigated previously in many species other than goldfish. The reproductive cycle of goldfish, is divided into five stages and the steroid synthesizing cells of the testes were studied during these stages, using histochemical techniques. We found that interstitial cells and seminiferous tubules are the main steroid synthesizing sites in testes of goldfish, and that enzyme activity was more intense in the interstitial cells than in the seminiferous tubules. During the pre-spawning months, 3 β-HSD and G-6-PD activities were weak compared to the spawning months.     

Histological, histochemical and ultrastructural studies on the interrenal and chromaffin cells of the fathead minnow, Pimephales promelas Rafmesque

Light and electron microscopic examination of fathead minnow head kidneys revealed that the interrenal and chromaffin cells were intermingled and always closely associated with the cardinal veins and their tributaries. Histochemical tests for lipids in the interrenal cells were positive, and two types of chromaffin cells were indicated by chromaffin reactions. Interrenal cells contained abundant smooth endoplasmic reticulum and mitochondria with tubulo-vesicular cristae, characteristic of steroid-producing cells. Only one interrenal cell type was found. Two types of chromaffin cells were present with differences in cytoplasmic density and in types of granules. In light cells, adrenaline granules were most common, and in dark cells noradrenaline granules predominated.     

Studies on the seasonal histocytological changes in the internal and chromaffin cells of male Puntius sophore (Ham) in relation to its reproductive cycle.

An attempt has been made to correlate the activities of interrenal and chromaffin cells with the reproductive cycle of Puntius sophore. Chromaffin cells do not have any significant bearing while interrenal cells undergo qualitative and quantitative changes which have been correlated with the various phases in the reproductive cycle of this fish. These cells undergo degranulation and vacuolisation during spawning phase, disorganisation during regression phase, and reorganisation and gradual growth during resting and prespawning phases. These cells thus appear to play an important role in the reproductive physiology of this fish.     

Histological and histochemical observations on the testis of Gobius paganellus

Summary Histological and histochemical observations on the testis of Gobius paganellus during all seasons of the year are described. In the yearly reproductive cycle, spawning in the Gulf of Naples occured from June through August, testicular recovery and relative inactivity from September through December, and active spermatogenesis from January to May.Germ cells develop as clones from single primary spermatogonia, each clone in a follicle enveloped by cells which are interpreted as Sertoli cell homologues.Glandular tissue is present in large amounts both in the form of interstitial islets and as a large mass along the length of the mesorchium. Cholesterolpositive lipids and ⁵-3-hydroxysteroid dehydrogenase, presumptive evidence for steroid production, are present exclusively in this tissue.Lipids are present in the glandular cells in acidic and neutral forms. The acid fats are the more abundant, but neutral lipids increase in amount in the period April-June. There is, furthermore, a cyclic variation in lipid droplet size, small droplets being present in the spring (during active spermatogenesis) and fewer, larger droplets during the fall (post-spawning period). Phospholipids are lacking.Lactic dehydrogenase activity was weak in the relatively inactive postspawning period, but was much more intense during active spermatogenesis.The glandular tissue in the testis of Gobius is interpreted as homologous with the interstitial (Leydig) tissue found in the testes of higher vertebrates.This investigation was supported by research grant RG-6455 from the Division of General Medical Sciences, U.S. Public Health Service.Postdoctoral Fellow from the Division of General Medical Sciences, U.S. Public Health Service. Supported in part by U.S. Public Health Service Training Grant 5 Tl GM-136, Department of Biological Structure, University of Washington, School of Medicine, Seattle, Washington, U.S.A.     

Metabolism of cortisone-4- 14 C by rat lung tissue

The metabolism of cortisone-4-¹⁴C has been studied in male rat lung tissue preparations. Data indicate the presence of 11β-hydroxysteroid dehydrogenase, Δ⁴-5α-reductase, 3α-hydroxysteroid dehydrogenase and 20α-hydroxysteroid dehydrogenase activity in this tissue. Metabolites identified were hydrocortisone, 17α, 20α, 21-trihydroxy-4-pregnene-3, 11-dione and 3α, 17α, 21-trihydroxy-5α-pregnan-11,20-dione.