Immunocytohistochemical characterization of pituitary cells of the bluefin tuna, Thunnus thynnus L

In this paper we report the first complete mapping of the pituitary in a tuna species. The various different adenohypophysis cell types of the bluefin tuna, Thunnus thynnus L. have been identified and located using different antisera against mammalian and piscine hormones and various histochemical techniques: PAS, Alcian Blue pH 2.5 and lectins -ConA and WGA(Neutral and Acidic Glycoproteins); Bromophenol Blue (Proteins) and Tioglycollate-Ferric-Ferricianide-FeIII (-S-S- groups). Prolactin (PRL) and adrenocorticotrophic (ACTH) cells were located in the rostral pars distalis (RPD) of the pituitary, while the proximal pars distalis (PPD) displayed gonadotrophic (GTH), thyrotrophic (TSH), somatotrophic (GH) and also a few PRL cells. Moreover, somatolactin (SL) and melanotrophic (MSH) cells were identified inside the pars intermedia (PI). Interestingly, some SL-immunoreactive fibers were also detected in the neurohypophysis. Some GTH cells were also located on the exterior surface of the PI. Glycoproteins containing mannose (Man) and/or glucose (Glc); N-acetyl-glucosamine (GlcNAc) and/or sialic acid sugar residues, as well as -S-S- groups, were observed in GTH, TSH and SL cells. The Bromophenol Blue technique stained amphiphilic SL, acidophilic GH cells and weakly ACTH cells. GH and ACTH cells were unreactive to PAS, Alcian Blue, Tioglycollate-Ferric-Ferricianide-FeIII and lectin (Con A and WGA) techniques. Finally, PAS reaction was positive in amphiphilic SL cells, which were PbH unreactive, while MSH and ACTH cells were stained with PbH technique.     

Immunocytochemical characterization of adenohypophyseal cells in the greater weever fish (Trachinus draco)

The adenohypophysis of the greater weever fish (Trachinus draco) was studied using histochemical and immunocytochemical methods. The adenohypophysis comprised the rostral pars distalis (RPD), the proximal pars distalis (PPD), and the pars intermedia (PI). Neurohypophysis showed a patent hypophyseal stalk which was divided into several branches intermingled with the adenohypophysis. Salmon prolactin (PRL)-immunoreactive (ir) cells, arranged in follicles, resided in the RPD and the most rostral part of the ventral PPD. Human adrenocorticotropin (ACTH)-ir cells were located in the RPD between PRL-ir cells and the neurohypophyseal processes. Salmon and seabream somatotropin (GH)-ir cells were located in both the dorsal and the ventral PPD. Some GH-ir cells were seen in surrounding and in contact with neurohypophyseal branches, whereas other isolated or clustered GH-ir cells were embedded in adenohypophyseal cells of the PPD. In addition, isolated or clustered GH-ir cells were also detected in the tissue of the PPD covering the most rostral part of PI. Only one class of salmon and carp gonadotropin (GTH)-ir cells was detected. Isolated or clustered GTH-ir cells resided in both the dorsal and the ventral PPD and were seen surrounding the PI and in the tissue of the PPD covering the most rostral part of PI. In addition, a few scattered GTH-ir cells were observed in the ventral RPD. Scattered groups of thyrotropin (TSH)-ir cells were present in the anteroventral PPD. Salmon and seabream somatolactin (SL)-ir and bovine melanotropin (MSH)-ir cells were intermingled surrounding the neurohypophyseal tissue. SL-ir cells were negative to periodic acid-Schiff technique. MSH-ir cells showed a very weak immunoreactivity to anti-human ACTH((1-24)) serum. In addition to the PI location, few isolated or clustered SL- and MSH-ir cells were observed in the dorsal PPD.     

Pituitary cytology of the teleost fish Tilapia mossambica (Peters)

In the pituitary of Tilapia mossambica, eight cell types were identified on the basis of their staining reactions. the RPD consists of erythrosinophils and PbH positive cells. PbH cells border the NH and give amphipilic reaction to tri- and tetrachrome dyes and Halmi's stain. Erythrosinophils are also stained with acid fuchsin and orange G. The PPD is made up of acidophils and cyanophils. The acidophils are stained with orange G, acid fuchsin and erythrosin. These cells form a palisade zone between the neurohypophysis and the cyanophils. The cyanophils are AB, AF, ATh, PAS and aniline blue positive. They include both thyrotrophs and gonadotrophs and cannot be differentiated. The gonadotrophs may be those which are degranulated and vacuolated in the breeding season. The PI is formed of PAS- and PbH positive cells which lack any definite pattern of arrangement. Apart from the chromophils, scattered chromophobes were seen throughout the adenohypophysis. Occasionally, cells resembling the cyanophils of PPD were noticed in the RPD and PI also.     

The Gonadotropic Cell in Parr,Precocious Parr Male and Smolt of the Atlantic Salmon,Salmo salar. An Immunocytological,Light- and Electron Microscopical Study

Lindahl, K. 1980. The gonadotropic cell in parr, precocious parr male and smolt of the Atlantic salmon, Salmo salar. An immunocytological, light- and electron microscopical study. (Department of Zoology, University of Stockholm, Sweden.) — Acta zool. (Stockh.) 61(2): 117–125. Only one GTH cell type was identified in the pars distalis of the parr, smolt and precocious parr male. On a fine structural basis four phases with transitional forms of the GTH cell were recognized. In parr and smolt the GTH cells have numerous secretory granules and their cytology substantiate that they are in a storing phase with a possibly low release rate. On the contrary, in the precocious parr male all phases of the GTH cell are present. The great number of active cells and frequent degranulation and vacuolization indicate a high activity and release level.     

Cyclical changes in the pituitary gonadotrophs (GTH cells) in relation to the testicular cycle in Puntius sarana (Hamilton)

The pituitary gland of Puntius sarana is of the leptobasic type and has three subdivisions--a proximal pars distalis (PPD), a rostral pars distalis (RPD) and a pars intermedia (PI) arranged vertically one below the other. The gonadotrophs (GTH cells) of the PPD were identified by their PAS, ATh, AF, aniline blue and alcian blue positivity and by their immunoreactivity to ovine anti-LH beta-subunit, anti-salmon GTH and anti-silver carp GTH. The testes of Puntius sarana have been classified as the "unrestricted type". The morphological, histological and histometric findings indicate that the testing passes through four distinct stages--preparatory, fully mature, spawning and spent. Spermatogenesis begins in January and continues up to July. The spawning season lasts from late July until October. Cyclical changes in the gonadotrophs correlate well with changes in the testes. The histometric data show that the GTH cells attain maximum values (size and number) during the breeding season. The peak values for testicular weight, lobule diameter, the gonadosomatic index (GSI) and the sperm concentration in the testes, and the number, size and amount of secretory material in the cytoplasm of the gonadotrophs, coincide with the time of maximum environmental temperature and the longest days.     

Histochemical procedures for the simultaneous visualization of neutral sugars and either sialic acid and its side chain O-acyl variants or O-sulphate ester. I. Methods based upon the selective periodate oxidation of sialic acids

Summary Five new methods, based upon the selective oxidation of sialic acid residues with 0.4mm periodic acid in approximately 1m hydrochloric acid at 4°C for 1 h (PA^*), have been devised for the simultaneous visualization of neutral sugars and either sialic acid and its side chainO-acyl variants orO-sulphate ester. In the first of these, the selective periodate oxidation—borohydride reduction—saponification—selective periodate oxidation—Thionin Schiff—saponification—borohydride reduction—periodic acid—Schiff (PA^*—Bh—KOH—PA^*—T—KOH—Bh—PAS) technique, sialic acids withO-acyl substituents at C7, C8 or C9 (or which have two of three side chainO-acyl substituents) stain blue while neutral sugars with periodate-sensitivevicinal diols (hexose, 6-deoxyhexose, andN-acetylhexosamine) stain magenta. The second method, the saponification—selective periodate oxidation—Thionin Schiff—saponification—borohydride reduction—periodic acid—Schiff (KOH—PA^*—T—KOH—Bh—PAS), stains all sialic acids blue and neutral sugars magenta. In the third procedure, the selective periodate oxidation—Thionin Schiff—borohydride reduction—periodic acid—Schiff—saponification (PA^*—T—Bh—PAS—KOH) method, sialic acids without side chain substituents (or which have anO-acyl substituent at C7) stain blue and neutral sugars stain magenta. In the fourth method, the saponification-selective periodate oxidation—borohydride reduction—Alcian Blue pH 1.0—periodic acid—Schiff (KOH—PA^*—Bh—AB1.0—PAS) technique,O-sulphate esters stain aquamarine blue and neutral sugars stain magenta. In all of these techniques mixtures of the components stain in various shades of purple. Performance of the KOH—PA^*—Bh—AB1.0—PAS technique without the Alcian Blue pH 1.0 step provides a method for the selective identification of neutral sugars in macromolecules that also contain sialic acids.     

Pituitary cytology of the freshwater teleost Anabas testudineus (Bloch)

In the pituitary of A. testudineus the rostral pars distalis (RPD), proximal pars distalis (PPD) and pars intermedia (PI) are arranged in a rostro-caudal axis. The rostral neurohypophysis (NH) extends dorsal to the pars distalis, while the caudal part vertically penetrates into the PI and ramifies in its component. The RPD mainly consists of erythrosinophils and PgH-positive cells which are comparable to the lactotropes and corticotropes mentioned in the literature. In the nonbreeding season, acidophils are the predominant cells of the PPD and are largely confined to its dorsal aspect. The two types of cyanophils present in the PPD could not be differentiated with the various techniques used in this study. However, those cyanophils which increase in number and are active during the spawning season may be the gonadotropes, while those which are cytologically inactive may be the thyrotropes. The pars intermedia consists of PAS+ and PhH+ cells. Trichrome and tetrachrome staining techniques revealed the presence of acidophils, cyanophils and amphiphils in the PI.     

Sialic acid in colonic mucin: An evaluation of modified PAS reactions in single and combination histochemical procedures

Summary Two new histochemical procedures for detecting sulphated and non-sulphated sialomucin in colonic mucosa were assessed: the saponification—Alcian Blue pH 1—periodic acid—phenylhydrazine—Schiff method (KOH—AB pH 1—PAPS) and the mild periodic acid modification of this (KOH—AB pH 1—mPAS). Using normal colonic mucosa obtained from 11 non-cancer patients, the mPAS and PAPS techniques were tested for specificity and reproducibility for staining sialic acid, either alone or in combination with Alcian Blue. A spectrophotometric method was devised to quantify the uptake of both Schiff and Alcian Blue stain by sections. At low temperature and pH5.5, the mPAS procedure had improved specificity over the PAPS procedure, and after saponification it could be used to stainO-acetyl-substituted sialic acid. When used in combination with Alcian Blue at pH 1, however, underestimation of the sialic acid content occurred owing to interference between Alcian Blue and Schiff dyes. Interference was even greater with KOH—AB pH1—PAPS procedure for both sialic acid and sulphate components. We conclude that caution must be exercised in interpretation of the staining results obtained with these new combination methods and that more accurate information on the sialic acid and sulphate content of colonic mucin is obtained by staining serial sections with the mPAS technique and Alcian Blue pH 1 alone.     

Neurosecretion in the basommatophoran snail Bulinus truncatus (Gastropoda,Pulmonata)

Summary The neurosecretory system of the freshwater snail Bulinus truncatus was investigated. With the Alcian blue-Alcian yellow (AB/AY) staining method at least 10 different types of neurosecretory cells (NSC) were distinguished in the ganglia of the central nervous system. The differences in staining properties of the NSC — with AB/AY the cells take on different shades of green and yellow — are borne out at the ultrastructural level: the NSC types contain different types of neurosecretory elementary granules.The neurosecretory system of B. truncatus is compared to that of Lymnaea stagnalis, the species which has received the most attention among the pulmonates. It appears from the comparison that the systems of both species show many similarities, although some differences are also apparent.     

Identification and localization of eight distinct hormone-producing cell types in the pituitary of male Atlantic halibut (Hippoglossus hippoglossus L.)

The eight distinct hormone-producing cell types in the adenohypophysis of male Atlantic halibut (Hippoglossus hippoglossus L.) were identified and localized using immunohistochemistry and in situ hybridization. Lactotropes either occupied most of the rostral pars distalis (RPD) or they were arranged in follicular structures located along the periphery of the RPD. Corticotropes were confined to a thin layer of RPD cells bordering the pars nervosa (PN). The somatotropes were arranged in multicellular layers bordering the highly convoluted PN penetrating the proximal pars distalis (PPD), while thyrotropes, scattered in small islets in between the somatotropes, were located in the centro-dorsal part of the PPD. Gonadotropes were found throughout the PPD. Immunoreactivity to glycoprotein-alpha and luteinizing hormone beta-subunit was also observed along the periphery of the pars intermedia (PI), indicating that a thin extension of the PPD surrounded the PI. In situ hybridization showed that follicle-stimulating hormone and luteinizing hormone were produced in distinct cells of the PPD. PI contained somatolactotropes bordering the highly convoluted PN, and melanotropes that showed positive immunostaining against both anti-alpha-melanocyte-stimulating hormone and anti-beta-endorphin. The general cellular organization was similar to that of other teleost fish. These results lay the basis for future investigations on Atlantic halibut pituitary physiology.     

Histophysiological and immunocytochcmical study on the nature of the thyrotrops in the pituitary of immature rainbow trout,Salmo gairdneri

Alcian blue- and periodic acid-Schiff-positive "granular basophils" with electron-dense granules +/- 160 nm in diameter, and weakly developed irregular cisternae of the granular endoplasmic reticulum (GER) are present in the rostral pars distalis (RPD) and the proximal pars distalis (PPD) of immature rainbow trout. They all react with antisalmon alpha beta-gonadotropin. However, only granular basophils in the caudal RPD and the rostro-dorsal PPD, often bordering on and sometimes surrounded by neurophypophysial tissue, react with anti-human beta-TSH. These cells, considered as the source of thyrotropin, show degranulation and dilatation of the GER-cisternae in fish treated with potassium perchlorate. The thyroids of the goitrogen-treated animals had relatively numerous small follicles with a high epithelium. The remaining granular basophils are gonadotrops.     

On the specificity of staining by Alcian blue in the study of human,foetal adenohypophysis

Summary The specificity of Alcian blue staining has been investigated on a material comprising the adenohypophysis from 26 human foetuses.Two distinct types of alcianophilic cells were observed. The first cell type appears about 28 mm CRL forming small groups at the epithelial-mesenchymal junction and beneath the anlage of the fibrous capsule. Besides its alcianophilic property the cell shows a pronounced maltase-resistant PAS-positivity. The alcianophilia is due to carboxylgroups — probably of proteins as no sialic acid could be detected. By the performic acid — Alcian blue method the cell seems to show a high content of SH and S-S groups.The second type of cell appears about 70 mm CRL and is mostly located singly. It shows a pronounced alcianophilia — probably due to sulphate groups and to some extent to carboxyl groups.The role of the alcianophilic properties of the cells in typing definite cells of the foetal adenohypophysis was discussed. — Finally the pitfalls caused by different fixatives and by the use of the general polychrome staining methods for the typing of pituitary parenchymal cells were discussed.This work was supported by grants from the Association for the Aid of Crippled Children, New York, and from the Danish State Research Foundation, Copenhagen.     

Immunofluorescent localization of insulin-like material in the median neurosecretory cells of the blowfly,Calliphora vomitoria (Diptera)

Summary Immunocytochemical staining has shown that the median neurosecretory cells (MNC) of the brain of the blowfly, Calliphora vomitoria, contain an insulin-like material which cross reacts with antibodies to bovine insulin. There are 24–26 paraldehyde fuchsin-positive MNC of which only 6–8 show the specific insulin-like immunoreactivity.Dr. M.C. Thorndyke, Department of Zoology, Bedford College, London University, kindly provided us with fluorescence facilities, and we are also grateful to him for helpful discussions on the immunofluorescence procedure     

Pituitary gland of the exotic carp, Carassius carassius Linn., with reference to the histocytology of the adenohypophysis in conjunction to male and female rhythms of reproduction

The pituitaries of the exotic carp (Carassius carassius) are studied at the light microscopic level, for the characterization of the adenohypophysial cell-types with particular emphasis to the gonadotropic potency of the pituitary in relation to the annual reproductive patterns. The gland in the fish is of the cranioleptobasic type. Based on the classical staining methods and localization of the differential cell-types, the adenohypophysis of the fish can be subdivided into the rostral (RPD) and proximal (PPD) pars distalis and the pars intermedia (PI). The pars nervosa (PN) of the fish consists of the neurosecretory (NS) fibres which show varied accumulations of the NS material in concert with the seasonal reproductive cycle. The NS tract is constituted of AF--ve nucleus praeopticus and AF + ve nucleus lateralis tuberis. 2 tinctorially different cell-types, corresponding to corticotrops and lactotrops of other teleosts, are identifiable in the RPD of the present species. In the PPD, 3 cell-types can be distinguished with various granule-staining procedures: they represent somatotrops, thyrotrops, and gonadotrops. Further, 2 gonadotropic cells with apparently different staining properties are discernible. In the PI are dispersed besides a few agranular cells, islets of 2 kinds of cells separable with combined PAS-PbH technique. Seasonal changes in the gonadosomatic index and diameter of the testicular lobules and egg diameter as indices of gonadal maturation are found to be closely interwoven with the morphometric alterations in the basophilic cell area (BCA) chosen as criterion of gonadotropic potency of the PPD. The maximum BCA values are preceded with the time of greatest atresia of the eggs indicating the influence of the pituitary gonadotropins in the follicular resorption in the test fish.     

Pituitary cytology of radiothyroidectomised teleost Heteropneustes fossilis (Bloch.).

The pituitary of Heteropneustes fossilis is divisible into rostral (RPD) and proximal (PPD) pars distalis, and pars intermedia (PI). Neurohypophysis extensively ramifies in the PI. The RPD is formed of acidophilic prolactin cells and lead hematoxylin (H. Pb) positive ACTH cells. The latter is arranged around the neurohypophysial extensions. The PPD is comprised of thyrotrophs (TSH) and gonadotrophs which are stainable with PAS, AF, and aniline blue. Acidophilic somatotrophs (STH) and chromophobes are scattered throughout the PPD. In the pars intermedia PAS positive and H. Pb positive cells are present. The latter is believed to secrete MSH whereas the function of the former is yet to be confirmed. In the partly and completely thyroidectomised H. fossilis the TSH cells exhibited marked hypertrophy. They are located in the middle of the PPD flanked by the gonadotrophs on either side. Hypertrophy of the ACTH cells were also evident. But the other cell types did not show any marked changes as compared to the controls.     

贝氏高原鳅脑垂体显微结构观察

贝氏高原鳅的垂体由神经垂体和腺垂体构成,属于背腹型,但神经垂体稍偏向背部.腺垂体从前到后又依次分为前外侧部(RPD)、中外侧部(PPD)和中间部(PI).前外侧部由ACTH细胞和PRL细胞组成,中外侧部由GH细胞和大小两种嗜碱性细胞组成,中间部分布有MSH细胞和另一种嗜碱性细胞.神经垂体分枝丰富,遍布于腺垂体的各个部分,在中间部尤为丰富.垂体内血管发达.     

The histochemical specificity of High Iron Diamine—Alcian Blue

Summary The specificity of the High Iron Diamine—Alcian Blue pH2.5 (HID—AB 2.5) procedure was examined in tissue sites containing sialogycoproteins alone or differing proportions of sialo- and sulphosialoglycoproteins. Studies with HID in differing final concentrations of hydrochloric acid or sodium chloride confirmed that staining is dependent upon both the pH and the ionic strength of the dye bath and demonstrated a marked heterogeneity in the pKa of the anionic groups of sialosulphoglycoproteins. Use of the sequence High Iron Diamine—Alcian Blue pH 1.0 demonstrated that complete or almost complete staining ofO-sulphate esters occurred when HID was prepared in water (final pH 1.3). However, under these conditions HID—AB 2.5 was shown to be non-specific because only black HID staining was observed in sites containing large quantities of sialic acids. This non-specificity was due either to the masking of Alcian Blue staining by HID and/or the black HID staining of anionic groups other than sulphate. These results may account for some of the conflicting data obtained by different groups of investigators who have studied transitional mucosa in human colonic diseases. Caution should be used in drawing conclusions from the use of HID—AB 2.5 without confirmatory evidence from other more specific procedures.     

Adenohypophysial cell types in the lamprey pituitary: current state of the art

Adenohypophysial cell types in the pituitary of adult sea lampreys, Petromyzon marinus, was localized by means of immunocytochemical and lectin cytochemical techniques. At least four types of adenohypophysial hormone cells are present in the pituitary of adult sea lampreys. The first type of cell is ACTH-like and occupies most parts of the rostral pars distalis (RPD), but a few scattered ACTH-like cells are also present in the proximal pars distalis (PPD). The second type of cell is MSH-like and occupies the whole pars intermedia. The third type of cell is GH/PRL-like and occupies the dorsal half of the PPD. These GH/PRL-like cells were initially detected by heterologous immunocytochemistry using antibodies to salmon GH, salmon PRL and blue shark GH, after hydrated autoclave pretreatment of sections. Later, by use of an antiserum raised against a synthetic peptide corresponding to the partial sequence of lamprey GH/PRL, the same cells as those containing GH/PRL-like immunoreactivity were stained positively. Similarity of the topographic distributions between lamprey GH/PRL-like cells and gnathostome fish GH cells in the pituitary suggests that GH/PRL-like cells in the lamprey may be GH cells. The last type of cell is GTH-like and occupies the ventral half of the PPD. Although GTH has not yet been isolated from the lamprey pituitary, our immunocytochemical data suggest that GTH-like material in the sea lamprey pituitary is more closely related to mammalian-like LH, rather than to FSH or TSH. These four types of adenohypophysial cells occupy most parts of the lamprey adenohypophysis and indeed there is little room for TSH or PRL cells. Thus, the present study further suggests that GH and LH-like GTH are ancestral forms of GH/PRL/SL family and glycoprotein hormones, respectively.     

Immunohistochemical Identification of the Adenohypophysial Cells in the Indian Paddy Field Frog,Rana limnocharis

Cells of the pituitary pars distalis (PD) and pars intermedia (PI) in the frog Rana limnocharis have been identified by an unlabelled antibody enzyme method using antisera developed in rabbit against mammalian hypophysial hormones. On the basis of their immunoreactivity, six types of cells, viz. thyrotropic (TSH), gonadotropic (GTH), prolactin (PRL), growth hormone (GH), corticotropic (ACTH) and melanotropic (MSH), cells have been recognized. GTH and PRL cells are distributed throughout the PD. GH cells usually occur in the anterodorsal and central region of the gland. Immunoreactive TSH cells are fewer in number and are localized in the ventromedian region of the PD. Cells showing immunoreactivity to ACTH 1–24 antiserum are encountered in the rostroventral part of the PD. Cells of the PI also show immunoreactivity to ACTH 1–24 antiserum. PI cells cross-react with α-MSH antiserum at all dilutions up to 1: 50 000. However, when the same antiserum was used at dilutions up to 1: 20 000, the ACTH cells of the PD also showed cross-reactivity.     

Immunocytochemical studies of the gonadotropic cells in the pituitary gland of male mullet, Mugil cephalus, during the annual reproductive cycle in both natural habitat and captivity

Using antiserum specific for the β subunit of coho salmon gonadotropic hormone II (GTH II), an immunocytochemical study of Mugil cephalus (L.) pituitaries was conducted during the annual reproductive cycle of the male in both natural habitat and captivity. The gonadotropic potency of the pituitary gland in general underwent an obvious increase during testicular development, reaching a peak at the time of reproductive maturity. During the testicular cycle of M. cephalus, the GTH cells showed an increase in immunoreactive staining intensity, granulation, hypertrophy and hyperplasia during sexual maturation. However, degranulation, vacuolization, and weakened immunoreactivity of these cells occurred during spawning. The GTH cells in the pituitary gland of M. cephalus males reared in captivity appeared with high synthetic and secretory activity but the reproductive activity declined, as reflected in the form of low values of the gonadosomatic index (GSI) and earlier resorption of the testes.