Effect of acute heat stress on rat adrenal cortex — a morphological and ultrastructural study

The stereological structure of rat adrenal gland was analysed by light and electron microscopy after an acute (60 min) exposure to high ambient temperature (38°C). Under these conditions a significant increase in plasma corticotrophin (ACTH), serum corticosterone and aldosterone levels were observed. Histological and stereological investigation at light microscopy showed significant decrease in volume density of capsule and zona glomerulosa, increase in volume of fasciculata cells, and decrease of numerical density of zona fasciculata cells and mean diameter of blood vessels. At the ultrastructural level, volume density of nuclei and mitochondria of zona glomerulosa cells were significantly increased and that of lipid droplets decreased. Volume density of mitochondria of fasciculata cells was significantly increased, while number of lipid droplets per μm² of cell was reduced. In the cells of zona reticularis significant increase in the number of lipid droplets was found. The response of zona glomerulosa may be interpreted as immediate reaction to dehydration, while alterations detected in zona fasciculata, which were less extensive, were related to purely stressogenic effects of high ambiental temperature.     

Autoradiographic localization of vitamin A in the adrenal gland of rats.

Cellular distribution of vitamin A in the rat adrenal was evaluated by autoradiography. Vitamin A was concentrated in the lipid droplets of epithelial cells of the zona fasciculata and zona reticularis. A small amount of vitamin A was also present in the cytoplasm and nucleus of these cells. The zona glomerulosa contained very little vitamin A either in lipid droplets or in the remainder of the cell. The medulla had essentially no vitamin A. Such strong cellular specificity for uptake of vitamin A by the cells of the zona fasciculata and zona reticularis supports the hypothesis that vitamin A is involved in synthesis of glucocorticoid hormones.     

Morphology and ultrastructure of the adrenal gland in Bactrian camels (Camelus bactrianus)

In the present study, we examined the morphological features of the adrenal gland in Bactrian camel by means of digital anatomy, light and electron microscopy. Our findings testified that the gland was divided into three parts, capsule, cortex and medulla from outside to inside as other mammals, and the cortex itself was further distinguished into four zones: zona glomerulosa, zona intermedia, zona fasciculate and zona reticularis. Notably, the zona intermedia could be seen clearly in the glands from females and castrated males, whereas it was not morphologically clear in male. There was a great deal of lipid droplets in the zona fasciculate, while it was fewer in the zona glomerulosa and zona reticularis. The cytoplasm of adrenocortical cell contained rich mitochondria and endoplasmic reticulum. The adrenal medulla was well-developed with two separations of external and internal zones. The most obvious histological property of adrenal medulla cells were that they contained a huge number of electron-dense granules enveloped by the membrane, and so medulla cells could be divided into norepinephrine cells and epinephrine cells. Moreover, the cortical cuffs were frequently present in adrenal gland. Results of this study provides a theoretical basis necessary for ongoing investigations on Bactrian camels and their good adaptability in arid and semi-arid circumstances.     

Effect of biseptol on the adrenal cortex of the white rat in postoperative shock

The study aimed to find out the effect of sulfonamide combined with Trimetaprim-Biseptol 480 on the adrenal cortex in post-operative shock after removal of SPIGELian lobe (lobectomy of the lobus caudatus and unilaterally of one kidney with its suprarenal gland. The study was performed on a material of white rats which were post-operatively administered Biseptol 480 in doses 5 times bigger than those given to men. It was attempted to determine histochemically the intensity of the adrenal cortex' function by testing the number of lipid droplets, activity of the main enzyme of steroidogenesis (beta-hydroxy steroid dehydrogenase) and the level of alpha-ketols (as the final stage of steroidogenesis). Pathomorphologic examinations were also performe. On the basis of the present study's results, it was observed that - in the case of liver-lobectomy - the zona fasciculata and zona reticularis are functionally stimulated but the zona glomerulosa becomes insufficient. In the case of nephrectomy plus suprarenal gland's removal, all the adrenal cortex becomes insufficient. Administration of Biseptol in the 1st case contributed to hormonal inactivation of the zone glomerulosa cells, but in the 2nd case, it caused an increased activity of steroid dehydrogenase and an increase of the alpha-ketol level in the zona fasciculata.     

Trophic and steroidogenic effects of water deprivation on the adrenal gland of the adult female rat

The effects of a 3-day water deprivation were studied in adult female rats in order to know what are the different zones of the adrenal gland and the hormonal factors involved in the growth and the activity of the adrenal gland. Water deprivation significantly increased plasma renin activity (PRA), plasma Angiotensin II (AII), vasopressin (AVP), epinephrine, aldosterone and corticosterone concentrations but did not modify the plasma adrenocorticotropin hormone (ACTH) level. Water deprivation significantly increased the absolute weight of the adrenal capsule containing the zona glomerulosa without modification of the density of cells per area unit suggesting that the growth of the adrenal capsule was due to a cell hyperplasia of the zona glomerulosa. Water deprivation significantly increased the density of AII type 1 (AT₁) receptors in the adrenal capsule but did not modify the density of AII type 2 (AT₂) receptors in the adrenal capsule and core containing the zona fasciculata, the zona reticularis and the medulla. The treatment of dehydrated female rats with captopril, which inhibits the angiotensin converting enzyme (ACE) in order to block the production of AII, significantly decreased the absolute weight of the adrenal capsule, plasma aldosterone and the density of AT₁ receptors in the adrenal capsule. The concentration of corticosterone in the plasma, the density of AT₂ receptors and the density of cells per unit area in the zona glomerulosa of the adrenal capsule were not affected by captopril-treatment. In conclusion, these results suggest that AII seems to be the main factor involved in the stimulation of the growth and the secretion of aldosterone by the adrenal capsule containing the zona glomerulosa during water deprivation. The low level of plasma ACTH is not involved in the growth of the adrenal gland but is probably responsible for the secretion of corticosterone by the zona fasciculata.     

Steroidogenic characteristics of the adrenal cortex of the mare studied by electron microscopy

The three concentric zones of the horse adrenal cortex (zonae glomerulosa, fasciculata and reticularis) showed marked interpenetration and exhibited a different relative development according to their position in the gland. Whereas the three cortical zones each had a specific histological structure, the ultrastructure of their cells showed a certain qualitative homogeneity. The differences corresponded essentially to the relative abundance of the constituents which are generally considered typical of steroidogeneous cells: mitochondria with vesicular cristae, smooth endoplasmic reticulum and lipid droplets. Their importance increased progressively from the zona glomerulosa to the zona reticularis. In this zone, the presence of gap and septate-like cell junctions, and mitochondria with vesicular cristae in close proximity to a smooth endoplasmic reticulum with numerous dilated tubules, suggested that steroidogenesis may be the most active. Ultrastructural findings were indicative of only quantitative differences between the steroidogenic capacities of the three zones of the mare adrenal cortex.     

Effect of chronic ACTH treatment on the physical state of lipid droplets in rat adrenocortical cells

A histophysical method has been adapted to determine the thermotropic phase transitions of adrenocortical lipid droplets using a polarizing microscope equipped with a cold/hot stage. Cryosections of freshly-removed, unfixed adrenals, derived from control (untreated), and 14 days ACTH-treated rats were examined. The lipid droplets in the zona fasciculata and zona reticularis of the untreated rats were birefringent at room temperature (22°C). The birefringence of zona glomerulosa lipids selectively increased in the temperature range from −10 to −15°C. In cryosections prepared from ACTH-treated rats, thermotropic phase transitions of the lipid droplets appeared at a temperature range between −30 and −40°C in each cortical zone. The chemical analysis of the isolated lipids revealed that the relative amount of triglycerides in the zona fasciculata lipids increased, while that of free and esterified cholesterol decreased after chronic ACTH treatment. Present data suggest that the increased fluidity of lipid droplets promotes lipid mobilization in response to the enhanced demand of the chronically stimulated adrenocortical cells. Viscosity-dependent mobilization of free cholesterol from lipid droplets is not a rate-limiting process in adrenal steroidogenesis, but rather may represent an important control of the availability of precursor from lipid stores.     

Zonal distribution of cytochromes P-450 and related enzymes of bovine adrenal cortex--quantitative assay of concentrations and total contents

The zona glomerulosa, zona fasciculata, zona reticularis, and medulla were separated from bovine adrenal glands and cytochromes P-450 and related enzymes in each zone were investigated immunochemically by Western blotting using antisera from chickens or rabbits against cytochromes P-450scc, P-450(11)beta, P-450s21, and b5, NADH-cytochrome b5 reductase, NADPH-cytochrome P-450 reductase, NADPH-adrenodoxin reductase, and adrenodoxin. Concentrations of cytochrome P-450(11)beta, NADPH-cytochrome P-450 reductase, and cytochrome b5 per milligram of protein of homogenate were higher in the zona glomerulosa than in the other zones; the levels of the other components were higher in the zona fasciculata. The total enzyme content of all components was the highest in the zona fasciculata. The amount of adrenodoxin was about 10 times that of NADPH-adrenodoxin reductase in each zone.     

Chronic stress and high sucrose intake cause distinctive morphometric effects in the adrenal glands of post-weaned rats

We investigated the effects of chronic stress combined with high sucrose intake on the morphology of the adrenal glands in young rats. Male Wistar rats were fed a standard chow diet and allocated into control (C; tap water), chronic restraint stress (St), 30% sucrose diet (S30) and 30% sucrose diet + chronic restraint stress (S30 + St) groups. St consisted of 1 h daily sessions, 5 days/week for 4 weeks. Chronic stress reduced the thickness of the zona glomerulosa (ZG) and zona fasciculata (ZF) in both right and left glands; the thickness of the zona reticularis (ZR) was increased in the right gland. Cell density was greater in the ZF and medulla of both right and left glands, whereas cell density increased in the ZR of only the left gland. The percentage of small cells was lower in the ZG, whereas more large cells were found in the left gland. A similar result was obtained for the ZF, ZR and medulla in both right and left glands. Chronic stress increased the area covered by blood vessels in the ZR of the right gland, but decreased the area in the ZR of the left gland. The area covered by blood vessels was reduced in the medulla of both right and left glands in rats subjected to chronic stress. Infiltration of immune cells was increased by chronic stress in all layers of the cortex of the left gland, but was reduced in the medulla of the right gland. A high sucrose diet reduced the thickness of the medulla in the left gland. Cell proliferation increased in the ZG of the right gland and the weight of the right adrenal gland increased. Reduced cell proliferation in the ZG of the left gland was associated with a reduction in the area covered by blood vessels. In addition, the area covered by blood vessels decreased in the medulla of both glands. Our findings demonstrate that exposure to chronic stress during early life causes morphometric changes in adrenal glands.     

Immunohistochemical localization of the somatostatin receptor subtype 2A in the rat adrenal gland

The immunohistochemical localization of the somatostatin receptor subtype sst2A was investigated in the rat adrenal gland using SS-800 polyclonal antibody. The sst2A immunopositivity was found in all adrenocortical zones and in adrenal medulla, the reaction being slightly more intense in zona glomerulosa and medulla. The administration of the potent agonist of sst2 receptors - octreotide - resulted in the enhancement of the immunopositivity in zona glomerulosa and medulla, whereas chronic exposure of the rats to diethylstilbestrol led to enhancement of the immunopositivity in zona glomerulosa and in the external part of zona fasciculata.     

Immunohistochemical localization of adrenal ferredoxin and distribution of adrenal ferredoxin and cytochrome P-450 in the rat adrenal

Adrenal ferredoxin, the iron-sulfur protein associated with cytochromes P-450 in adrenocortical mitochondria, has been localized immunohistochemically at the light microscopic level in rat adrenals by employing rabbit antiserum to bovine adrenal ferredoxin in both an unlabeled antibody peroxidase-antiperoxidase method and an indirect fluorescent antibody method. When sections of rat adrenals were exposed to the adrenal ferredoxin antiserum in both procedures, positive staining for adrenal ferredoxin was observed in parenchymal cells of the three cortical zones but not in medullary chromaffin cells. Marked differences in the intensity of staining, however, where observed among the three cortical zones: the most intense staining being found in the zona fasciculata, less in the zona reticularis, and least in the zona glomerulosa. Furthermore, differences in staining intensity were also observed among cells within both the zona fasciculata and the zona reticularis. In agreement with these immunohistochemical observations, determinations of adrenal ferredoxin contents by electron paramagnetic resonance (EPR) spectrometry in homogenates prepared from capsular and decapsulated rat adrenals revealed that the concentration of adrenal ferredoxin in the zona glomerulosa was lower than that in the zona fasciculata-reticularis. Similar results were obtained when the contents of cytochrome P-450 were determined in capsular adn decapsulated rat adrenal homogenates. These observations indicate that adrenal ferrodoxin and cytochrome P-450 are not distributed uniformly throughout the rat adrenal cortex.     

Sex differences in adrenocortical structure and function

Summary Adrenal glands of adult female rats are heavier than the glands of corresponding male rats. Postpubertal orchiectomy increases the adrenal weight, an effect restored by testosterone replacement. Under the same conditions ovariectomy of 8 weeks duration does not change the adrenal weight while estradiol replacement enhances the relative adrenal weight.Karyometric studies have shown that nuclei in the female zona fasciculata cells are larger (app. 18%) than those of the male. Similar but only slight differences (2%) were observed in the zona reticularis. Orchiectomy results in enlargement of cell nuclei within all zones of the adrenal cortex; testosterone replacement has the opposite effect. Ovariectomy of 8 weeks duration slightly enhances the volume of nuclei of the zona glomerulosa cells, has no effect on the nuclei of the zona fasciculata and reduces the volume of nuclei in the zona reticularis. Estradiol replacement reduces the volume of nuclei of the zona glomerulosa cells but increases nuclear volume in the zona fasciculata and in the zona reticularis.Thus testosterone has an inhibitory effect on the adrenal cortex of the rat while the physiologic effect of estradiol on the morphology of this gland, particularly on the zona fasciculata cells is rather dubious.The author wishes to thank Mrs. B. Westerska and Miss K. Siejak for excellent technical assistance.This research was supported in part by a grant from the Zoological Committee, 2nd Department, Polish Academy of Sciences.This paper is dedicated to the memory of late Kazimierz Mietkiewski, M.D., Ph.D. whose encouragements and suggestions were most stimulating for my work.     

Adrenal cortical foci and nodules in Sprague-Dawley rats treated with N-nitrosomorpholine. Light and electron microscopic findings

Treatment of male Sprague-Dawley rats with N-nitrosomorpholine (NNM) in the drinking water at a dose of 120 mg/l for 7 weeks resulted in a subsequent enhanced development of focal and nodular lesions in the adrenal cortex. Sequential observation revealed that focal lesions in the zona reticularis/fasciculata or the zona glomerulosa developed both earlier and at a significantly higher incidence in animals treated with carcinogen than in untreated controls. Foci observed within or adjacent to the zona glomerulosa were all of pale cell appearance and contained large numbers of electron-dense cytoplasmic granules similar to those observed in normal granulosa cells. The foci and nodules which arose in the zona reticularis/fasciculata were, in contrast, characterized by a reduction or loss of the dense osmiophilic droplets normally seen in the cells of this region of the adrenal cortex, a pronounced increase in pleomorphic mitochondria of atypical appearance and the development of vacuoles.     

Ultracytochemical localization of particulate guanylate cyclase in rat adrenal gland exposed to stimulation by porcine brain natriuretic peptide

Summary We studied the cytochemical localization of particulate guanylate cyclase (GC) in rat adrenal gland after stimulation with porcine brain natriuretic peptide (pBNP) by electron microscopy. In the adrenal cortex, GC activity, as demonstrated by the presence of reaction product, was prevalently localized to the zona glomerulosa and zona fasciculata, while the zona reticularis showed little GC reaction product. In the adrenal medulla, GC reaction product was present only in adrenalin-containing cells. All GC positivity was associated with intracellular membranes. No GC reaction product was detected in specimens incubated in media devoid of pBNP. In parallel samples incubated in the presence of rat atrial natriuretic factor (rANF), the distribution of rANF-stimulated GC activity was similar to that of pBNP-stimulated GC activity.     

Histochemical localization of monoamine oxidase types A and B in the adrenal gland

Summary The distribution of monoamine oxidase types A and B within the adrenal galdn was studied in several mammals by histochemical methods. Controls showed that the methods were valid. The bovine adrenal medulla contained mostly the B type enzyme, distributed heterogeneously, with some A type associated with endothelium, nerves, and cells surrounding the nerves. The bovine adrenal cortex showed a marked zonation of the two types of monoamine oxidase. The zona glomerulosa contained the B type enzyme and the zona fasciculata and zona reticularis contained the type A enzyme. The adrenal medulla of the dog, cat, and rat demonstrated relatively little enzyme activity and it appeared to be both type A and B. The adrenal cortex of these animals appeared to contain mostly the B type enzyme, except the canine zona reticularis, which contained some A type monoamine oxidase as well.     

Morphometric and histological studies on the adrenal glands of the camel Camelus dromedarius

The adrenal gland of the camel consists of an outer cortex and an inner medulla. The general disposition of the cortex and medulla, however, differs occasionally from that of other mammals. Extensions of medulla could reach as far as the periphery of the cortex. Islet of medullary tissue may be found in sections of the cortex and cortical tissue consisting of all zones of the cortex may occur around arteries or nerves in the medulla. The medulla may be separated from the cortex by connective tissue especially in old camels. The arrangement of noradrenaline-secreting cells is different from that in other ruminants; they are found in groups scattered between the adrenaline-secreting cells. Bundles of smooth muscle occur in venules at the corticomedullary interface. Accessory adrenal glands are found embedded in the renal fat. They are similar in structure to the adrenal gland. The adrenal cortex forms 74% of the volume of the gland and the ratio of the cortex to medulla is 4:1. The zona glomerulosa, fasciculata and reticularis constitute about 13%, 53%, and 29% by volume of the cortex, respectively.     

Immunocytochemical localization of adrenodoxin in bovine adrenal cortex by protein A-gold technique

To investigate the intracellular localization of the enzymes that are involved in steroid hormone synthesis, an immunocytochemical study of the distribution of adrenodoxin in cells of the bovine adrenal cortex was carried out by the post-embedding immunostaining method and by immuno-cryoultramicrotomy in combination with the protein A-gold technique. Gold particles were seen on the matrix and the inner membrane of all the mitochondria examined, which have typical vesicular or tubulo-vesicular cristae, in parenchymal cells of the zona fasciculata and the zona reticularis. Gold particles were distributed homogeneously among the mitochondria. The density of gold particles on mitochondria in the parenchymal cells of the zona glomerulosa was lower than that of the zona fasciculata, which was similar to that of the zona reticularis. Gold particles were also seen on round, electron-dense intramitochondrial bodies in the parenchymal cells. The intramitochondrial bodies were abundant in the zona glomerulosa and the outer region of the zona fasciculata.     

Immunohistochemical localization of oxytocin and vasopressin in the adrenal glands of rat,cow, hamster and guinea pig

Summary The distribution of oxytocin and vasopressin in the adrenals of rat, cow, hamster and guinea pig has been studied by use of immunohistochemical techniques. In all the species studied the adrenal cortex contained both peptides; the staining in the zona glomerulosa being more intense than that in zona fasciculata or zona reticularis. The medulla, however, showed considerable species variation. In the cow, both peptides appear to be present in the adrenergic and noradrenergic cells, though staining was particularly prominent in cortical islands interspersed within the medullary tissue. In the rat, groups of medullary cells positive for both peptides were found, though it was not possible to associate these groups with particular chromaffin cell types. In the hamster oxytocin was present only in adrenaline-containing cells, whereas vasopressin was present in all medullary cells. The guinea pig medulla, which contains only adrenaline-secreting cells, was positive for both peptides. The possibilities that vasopressin and oxytocin have an autocrine or paracrine role in functioning of the adrenal gland is discussed.     

Carbonic anhydrase isoenzymes II and I are present in the zona glomerulosa cells of the human adrenal gland

Human carbonic anhydrase isoenzymes I and II (HCA I and II) were purified from human erythrocytes by inhibitor affinity chromatography and ion-exchange chromatography. These isoenzymes were then located in the human adrenal gland using specific polyclonal antisera raised in rabbits and specific detection by immunohistochemical techniques. Both HCA II and I were located in the zona glomerulosa cells, although the staining for HCA I was faint. The cells of the zona fasciculata and the zona reticularis failed to stain with either antiserum. Control stainings with preimmune or anti-HCA VI sera were negative. The presence of HCA II and I in the zona glomerulosa cells may be linked to regulation of the biosynthesis or secretion of mineralocorticoids.