Development of gonadotropes in the chicken embryonic pituitary gland

Although a number of immunohistochemical studies have been carried out on the differentiation of chicken gonadotropes during embryogenesis, the temporal and spatial properties of appearance of gonadotropes are not clear. In this study, we studied the appearance and morphological characteristics of gonadotropes in the embryonic and adult chicken anterior pituitary glands using RT-PCR, in situ hybridization and immunohistochemistry. For this purpose, we raised specific antisera against chicken follicle-stimulating hormone beta-subunit (cFSHbeta) and chicken luteinizing hormone beta-subunit (cLHbeta) based on each putative amino acid sequence. RT-PCR analysis revealed that cFSHbeta mRNA was expressed from embryonic day 7 (E7). Chicken FSHbeta mRNA-expressing (-ex) and -immunopositive (-ip) cells started to appear in the ventral part of the caudal lobe in the anterior pituitary gland at E8. Chicken LHbeta-ip cells were also first observed there at E8, but cLH mRNA expression was confirmed from E4 by RT-PCR analysis. The distribution of these chicken gonadotropin-ex and -ip cells spread from the ventral part to dorsal part in the caudal lobe around E10 and subsequently expanded to the cephalic lobe from E12 to E20. These cells were morphologically classified into two types (round- and club-shaped cells). It was found that the density of gonadotropin-ip cells in the caudal lobe was always higher than that in the cephalic lobe throughout the period of development. To the best of our knowledge, this is the first report focusing on the differentiation of chicken gonadotropes by assessment of both protein and mRNA of chicken gonadotropin.     

Increased proliferative activity and programmed cellular death in the turkey hen pituitary gland following interruption of incubation behavior

Incubation behavior or broodiness in turkey hens is characterized by ovarian regression, hyperprolactinemia, and persistent nesting. Nest-deprivation of incubating turkey hens results in disruption of broodiness accompanied by a precipitous decline in plasma prolactin (PRL) concentrations. The objective of the present study is to examine cellular changes in the pituitary gland associated with nest-deprivation for 0, 1, 2, 3, 4, or 7 days. Bromodeoxyuridine (BrdU) was administered prior to kill to study proliferative activity. Pituitary tissue sections were immunostained using turkey growth hormone (GH) antibody, and/or chicken PRL peptide antibody, and BrdU antibody. Plasma PRL concentrations declined significantly following nest-deprivation for 1 or more days. The midsagittal pituitary area immunoreactive (ir) to GH was significantly increased while that of PRL was significantly decreased following nest-deprivation for 2 or more days. Terminal deoxy-UTP nick end labeling and PRL-immunostaining revealed an abundance of apoptotic nuclei in both cephalic and caudal lobes of the anterior pituitary gland, suggestive of programmed cellular death of lactotrophs in the pituitary gland of hens nest-deprived for 2 or more days. Mammosomatotrophs were abundant in hens nest-deprived on Day 0 but were absent in hens nest-deprived for 1 or more days. Proliferating (BrdU-ir) cells were significantly abundant in the pituitary cephalic and caudal lobes following nest-deprivation for 1 or more days but were absent on Day 0 or in laying hens. Dual-labeling studies indicated that most of the BrdU-ir nuclei in the caudal lobe were not colocalized in somatotrophs in hens nest-deprived for 1-4 days but did colocalize with GH following 7 days of nest-deprivation. In conclusion, nest-deprivation of incubating turkey hens results in 1) a precipitous decline in plasma PRL concentration, 2) programmed cell death of lactotrophs, 3) disappearance of mammosomatotrophs, 4) increased proliferative activity of pituitary cells, and 5) recruitment of somatotrophs arising primarily from mitosis of nonsomatotrophic cells.     

Expression of the common α-subunit mRNA of glycoprotein hormones during the chick pituitary organogenesis, with special reference to the pars tuberalis

The expression of a common alpha-subunit mRNA of glycoprotein hormones was examined in the pituitary of chick embryos at various stages of development by in situ hybridization with a digoxigenin-labeled quail alpha-subunit cRNA probe. As a comparison with the expression of alpha-subunit mRNA, the onset of luteinizing hormone (LH) immunoreactivity was examined by immunohistochemical staining with a chicken LH antiserum. Both alpha-subunit mRNA and LH immunoreactivity began to appear in the basal-posterior region of the Rathke's pouch at embryonic day (E) 3.5. At E4.5 when the cephalic and caudal lobes of the pars distalis could be distinguished in the Rathke's pouch, intense signal for alpha-subunit mRNA was restricted to the cephalic lobe, consisting of a high columnar epithelium. At E6, gonadotrophs that were ovoid in shape, expressed intense signal for alpha-subunit mRNA, and revealed intense immunoreactivity for LH, were first detected in the cephalic lobe. At this stage, alpha-subunit mRNA expression became weak in the undifferentiated columnar cells of the cephalic lobe. At E8, the pars tuberalis primordium located close to the median eminence was formed at the lateral-apical end of the cephalic lobe. The primordium expressed intense signal for alpha-subunit mRNA. Gonadotrophs showing immunoreactivity for LH were densely distributed throughout the cephalic and caudal lobes in 8-day-old embryos. The pars tuberalis primordium expressing alpha-subunit mRNA progressively extended along the median eminence with embryonal age and reached the rostoral end by E14. Thus, both primordia of the pars distalis and pars tuberalis expressed intense signal for the common alpha-subunit mRNA. This subunit may play a role in the cytodifferentiation of the adenohypophysis.     

Hypophyseal corticosteroids stimulate somatotrope differentiation in the embryonic chicken pituitary gland

Although it is known that glucocorticoids induce differentiation of growth hormone (GH)-producing cells in rodents and birds, the effect of mineralocorticoids on GH mRNA expression and the origin of corticosteroids affecting somatotrope differentiation have not been elucidated. In this study, we therefore carried out experiments to determine the effect of mineralocorticoids on GH mRNA expression in the chicken anterior pituitary gland in vitro and to determine whether corticosteroids are synthesized in the chicken embryonic pituitary gland. In a pituitary culture experiment with E11 embryos, both corticosterone and aldosterone stimulated GH mRNA expression and increased the number of GH cells in both lobes of the pituitary gland in a dose-dependent manner. These effects of the corticosteroids were significantly reversed by pretreatment with mifepristone, a glucocorticoid receptor (GR) antagonist, or spironolactone, a mineralocorticoid receptor (MR) antagonist. Interestingly, an in vitro serum-free culture experiment with an E11 pituitary gland showed that the GH mRNA level spontaneously increased during cultivation for 2 days without any extra stimulation, and this increase in GH mRNA level was completely suppressed by metyrapone, a corticosterone-producing enzyme P450C11 inhibitor. Moreover, progesterone, the corticosterone precursor, also stimulated GH mRNA expression in the cultured chicken pituitary gland, and this effect was blocked by pretreatment with metyrapone. We also detected mRNA expression of enzymes of cytochrome P450 cholesterol side chain cleavage (P450scc) and 3β-hydroxysteroid dehydrogenase1 (3β-HSD1) in the developmental chicken pituitary gland from E14 and E18, respectively. These results suggest that mineralocorticoids as well as glucocorticoids can stimulate GH mRNA expression and that corticosteroids generated in the embryonic pituitary gland by intrinsic steroidogenic enzymes stimulate somatotrope differentiation.     

Immunohistochemical characterization of chicken pituitary cells containing the vasotocin VT2 receptor

Research in mammals has demonstrated a variety of regulatory effects of vasopressin and oxytocin on endocrine functions of the anterior pituitary gland. Less evidence is available regarding the hypophysiotropic action of arginine vasotocin (AVT) comprising vasopressic and oxytocic activities in birds. Some hypophysiotropic effects of AVT may result from its interactions with brain circuits controlling pituitary functions, whereas others are caused by its direct affect on pituitary cells. Use of an antiserum to the vasotocin receptor VT2 (VT2R) has revealed numerous immunoreactive cells in the anterior pituitary gland of the chicken. The objective of the present study has been to identify endocrine phenotypes of chicken pituitary cells containing VT2R by means of immunohistochemical labeling. VT2R immunoreactivity has been found in all cells immunoreactive for adrenocorticotropin and alpha-melanotropin. Approximately 10% of labeled lactotropes are also immunoreactive for VT2R and lie around the anatomical boundary dividing the cephalic and caudal lobes. In both corticotropes/melanotropes and lactotropes, immunoreactive VT2R is present in a narrow layer outlining cell bodies. Immunoreactive VT2R is not found in gonadotropes, thyrotropes, or somatotropes. These results provide evidence for the important role of VT2Rs in mediating effects of AVT on endocrine secretion from corticotropes and, partially, from lactotropes.     

Immunohistochemistry of the hypothalamic neuropeptides and anterior pituitary cells in the Japanese quail

The pars distalis of the avian adenohypophysis consists of well-defined cephalic and caudal lobes which are distinct in their cellular constituents. Immunocytochemical investigations on the pituitary hormones of the pars distalis of the Japanese quail reveal five types of secretory cells, adenocorticotropin (ACTH) cells, prolactin (PRL) cells, thyroid-stimulating hormone (TSH) cells, growth hormone GH (STH) cells, and FSH/LH (gonadotropic) cells. The ACTH cells, TSH cells, and PRL cells are restricted to the cephalic lobe, and GH (STH) cells are confined to the caudal lobe, while FSH/LH cells are distributed throughout the cephalic and caudal lobes. The median eminence of birds has distinct anterior and posterior divisions, each with different neuronal components. The avian hypophysial portal vessels also consists of two groups, anterior and posterior. The peculiar arrangement and distribution of the avian hypophysial portal vessels are possibly related to the distribution of neuropeptides in the two divisions of the median eminence and to the cytological and functional differentiation of two lobes of the pars distalis. The localization of perikarya and fibers containing luteinizing hormone releasing hormone (LHRH), somatostatin, vasotocin, mesotocin, corticotropin-releasing factor (CRF), vasoactive intestinal polypeptide (VIP), glucagon, metenkephalin, and substance P in the hypothalamus and median eminence of the Japanese quail has been investigated by means of immunohistochemistry using antisera against the respective neuropeptides. LHRH-, somatostatin-, VIP-, met-enkephalin-, and substance P-immunoreactive fibers are localized in the external layer of the anterior and posterior divisions of the median eminence, while CRF- and vasotocin-reactive fibers are demonstrated only in the external layer of the anterior division of the median eminence. The metenkephalin fibers are thicker in the anterior median eminence but the substance P fibers are more abundant in the posterior division. Mesotocin fibers occur only in the internal layer of the median eminence and neural lobe.     

Cytodifferentiation of the adenohypophysis of the domestic fowl

Summary In the chick embryo the first membrane-bound secretory granules occur in the cytoplasm of occasional cells in the cephalic lobe of pars distalis at the 7th day of incubation. On the 8th day most of the cells in both the cephalic and caudal lobes contain secretory granules that are variable in size, form and density.On the 9th day at least two types of glandular cells are distinguishable in the cephalic and in the caudal lobes; however, these cells are not comparable with those of the adult gland. Differentiation of acidophils and basophils occurs, apparently simultaneously, in 11-day embryos.The cells of the cephalic and caudal lobes are morphologically distinct from their first appearance. Thus it is concluded that these two lobes develop independently and differently from an early stage of ontogenesis.The secretory granules are formed in the Golgi area of the hypophysial cells after the 8th day of incubation. However, secretory material may be synthesized also by a process not involving the Golgi apparatus.Nerve fibers containing granules first appear in the superficial layer of the median eminence on the 8th embryonic day and by the 12th day three types of granules and two types of clear vesicles are identifiable.The investigation reported herein was supported by grant from Japan-U.S. Cooperative Science Program of Japan Association for Science Promotion to Professor Mikami and by U.S.-Japan Cooperative Science Program Grant No. GF-33334 to Professor Farner.     

Immunocytochemical localization of secretogranin III in the anterior lobe of male rat pituitary glands.

Secretogranin III (SgIII) is one of the acidic secretory proteins, designated as granins, which are specifically expressed in neuronal and endocrine cells. To clarify its precise distribution in the anterior lobe of the rat pituitary gland, we raised a polyclonal antiserum against rat SgIII for immunocytochemical analyses. By immunohistochemistry using semithin sections, positive signals for SgIII were detected intensely in mammotropes and thyrotropes, moderately in gonadotropes and corticotropes, but not in somatotropes. The distribution pattern of SgIII in the pituitary gland was similar to that of chromogranin B (CgB), also of the granin protein family, suggesting that the expressions of these two granins are regulated by common mechanisms. The localization of SgIII in endocrine cells was confirmed by immunoelectron microscopy. In particular, secretory granules of mammotropes and thyrotropes were densely and preferentially co-labeled for SgIII and CgB in their periphery. Moreover, positive signals for SgIII were occasionally found in cells containing both prolactin and TSH in secretory granules. These lines of evidence suggest that SgIII and CgB are closely associated with the secretory granule membrane and that this membrane association might contribute to gathering and anchoring of other soluble constituents to the secretory granule membrane.     

Somatotrophs and lactotrophs: an immunohistochemical study of Gallus domesticus pituitary gland at different stages of induced moult

The objective of this study was to determine the distribution of somatotrophs and lactotrophs and conduct a morphometrical analysis of immunoreactive somatotrophs and lactotrophs in the pituitary glands of White Leghorn Hens (Gallus domesticus) during the period of induced moult. We divided the periods of induced moulting into three phases viz. 7, 14 and 21 days. The labeled alkalinephsphatase method with anti-GH (growth hormone) and anti-PRL (prolactin) as a primary antibody was used to detect somatotrophs and lactotrophs, in the midsagital sections of chicken adenohypophysis. Immunohistochemistry showed that somatotrophs are not only confined to the cephalo-caudal axis but can also be found in the caudal lobe; while lactotrophs were distributed in both lobes of the anterior pituitary gland at all stages of moulting (7, 14 and 21 days). Lactotrophs were of different shapes but somatotrophs were oval to round in morphology. At the given stages of induced moulting, some hypertrophied lactotrophs were also present after 7 days of induced moult in the anterior pituitary gland. However, there were moulting-related changes: from 7 to 21 days of induced moulting the immunoreactive-PRL cell population decreased, while the mean lactotroph size was more than that of somatotrophs. Basic quantitative and morphological information relating to somatotrophs and lactotrophs during the period of induced moult in laying hens is reported here and the changes brought about by induced moulting are restricted to PRL positive cells rather than GH positive cells.Key words: Moult, pituitary gland, somatotrophs, lactotrophs, chicken.     

Circular polysomes predominate on the rough endoplasmic reticulum of somatotropes and mammotropes in the rat anterior pituitary

We have studied the shape and size distribution of membrane-bound polysomes in somatotropes and mammotropes, which are the sources, respectively, of growth hormone and of prolactin in the rat pituitary. The observations were made in conventional electron micrographs of these cells in situ, where occasional surface or en face views of the rough endoplasmic reticulum allow the polysomes to be seen as rows of ribosomes arranged in distinctive patterns on the membranes. It is possible by this means to characterize the shape and number of ribosomes for the total population of bound polysomes in the respective cell types. The great majority of membrane-bound polysomes in these two cell types (81% in somatotropes, 78% in mammotropes) have an approximately circular shape and contain an average of 6.8 (somatotropes) or 6.5 (mammotropes) ribosomes, which is an appropriate size for translation of the polypeptide hormones produced by these cells. About 17% of the membrane-bound polysomes in somatotropes and 20% in mammotropes have a spiral shape, resembling somewhat the letter "G," and contain about eight to nine ribosomes in both cell types. The preponderance of circular polysomes on the rough endoplasmic reticulum of somatotropes and mammotropes suggests the possibility that ribosomes (or the 40S ribosomal subunit) may recycle on the polysome after the translation of growth hormone or of prolactin.     

Immunohistochemical evidence that follicle-stimulating hormone and luteinizing hormone reside in separate cells in the chicken pituitary.

As is the case in other tetrapod species, the chicken gonadotropins LH and FSH consist of a common alpha subunit and a hormone-specific beta subunit. Gonadotrophs containing LH were shown earlier to be distributed throughout both the caudal and cephalic lobes of the chicken anterior pituitary, but the cellular distribution of FSH in avian species is still uncertain. The purpose of this study was to determine the cellular distribution of FSH-containing chicken gonadotrophs by use of FSH-specific monoclonal antibodies (mAbs). Three new mAbs toward chicken FSH were proven hormone specific by immunodetection of purified hormones on dot blots and by dual-label immunohistochemistry (IHC) on sagittal sections of chicken pituitaries. A rabbit antibody was used to detect chicken LH. Results showed that LH-containing gonadotrophs were densely distributed throughout the anterior pituitary, whereas gonadotrophs containing FSH were much less numerous; in addition, while also present in both lobes, FSH-positive cells were largely absent from the outer margin of the gland. Dual-label IHC revealed that LH and FSH reside almost exclusively in separate gonadotrophs. The identity of FSH-containing cells was further confirmed through use of an antibody to the chicken alpha subunit, which showed that FSH immunoreactivity was always colocalized with the alpha subunit. Our results suggest the possibility that production and secretion of LH and FSH may be regulated differently in chickens than in most other species studied to date.     

Localization of fertility factor SP22 to specific cell types within the anterior pituitary gland

Sperm protein 22 (SP22) was recently identified in the anterior pituitary gland (AP) of male Golden Syrian hamsters using ion trap mass spectrometry. SP22 has been implicated in apoptosis, androgen receptor function, fertility, and ontogeny of early-onset Parkinson's disease. However, the role of SP22 in the pituitary has not been investigated. We cloned the cDNA for full-length SP22 from AP and posterior lobe (posterior pituitary and intermediate lobe) of the pituitary gland in adult male rats and Golden Syrian hamsters, confirming the presence of SP22 mRNA in the AP and posterior lobe. Because gonadal steroids are important regulators of AP function, and SP22 is associated with androgen receptor function, we used Western blots to compare SP22 in the AP of intact and orchidectomized male rats given placebo or a low or high dose of testosterone. SP22 did not differ with treatment, indicating that AP SP22 concentration was not regulated by testosterone. To localize SP22 to specific cells of the AP, mirror-image paraffin sections were labeled against SP22 and either luteinizing hormone (LH)beta, thyroid-stimulating hormone (TSH)beta, prolactin, adrenocorticotropic hormone (ACTH), or growth hormone (GH) using peroxidase-conjugated secondary antibody. Additional sections were colabeled with SP22 and one of the AP hormones using fluorescent secondary antibodies. SP22 colocalized in somatotropes and thyrotropes in rat and hamster. We identified SP22 in a small percentage of corticotropes, gonadotropes, and lactotropes. This is the first report that SP22 mRNA is present specifically in the AP, and SP22 is localized primarily in somatotropes and thyrotropes. SP22 may help regulate AP function and be particularly important for the control of GH and TSH secretion.     

Light- and electron-microscopic studies on the secretory cytology of the adenohypophysis of the Japanese quail,Coturnix coturnix japonica

The effects of thyroidectomy, adrenalectomy, and castration on the pars distalis of male Japanese quail, and of injection of LH-RH on sexually inactive females, were investigated by light and electron microscopy. Correlation between light and electron microscopy was attained by use of alternate thin and thick sections. Six types of secretory cells were identified and the ultrastructural characteristics described. Putative endocrine functions have been designated on the basis of responses to experimental interventions and on other criteria. The putative STH cells are characterized by the presence of large dense secretory granules (250-300 nm) that are stained with orange-G by the trichrome method. They occur only in the caudal lobe and appear to be unchanged by castration, thyroidectomy, adrenalectomy and LH-RH injection. The putative prolactin cells are characterized by large (400-600 nm), spherical or polmorphic, dense secretory granules stainable with acid fuchsin and aniline blue; prominent Golgi apparatus and well developed endoplasmic reticulum with densely packed, regularly parallel lamellae. They are found mainly in the cephalic lobe. The prolactin cells develop some vacuolization after adrenalectomy and undergo some degeneration after castration. The ACTH cells, which are restricted to the cephalic lobe, are identified by the dense, spherical granules (250-300 nm) that are stained with acid fuchsin. After adrenalectomy, they lose their secretory granules and are transformed into large, chromophobic adrenalectomy cells. TSH cells are so designated by their response to thyroidectomy including loss of their fine secretory granules and transformation to large, vacuolated thyroidectomy cells. We have found TSH cells and thyroidectomy cells only in the cephalic lobe. Basophilic cells, considered to be gonadotropes, occur in both the cephalic and caudal lobes. The gonadotropes of the cephalic lobe appear to have slightly larger (120-200 nm) granules than the caudal lobe (120-150 nm). However, after castration, the gonadotropes in both lobes become hypertrophied and vacuolated and are transformed into mutually indistinguishable castration cells. Twenty minutes after injection with LH-RH, the gonadotropes of both lobes increase in size and number, degranulate, develop vacuoles in the cytoplasm, and appear very similar to castration cells.     

Expression of neural cell adhesion molecules,NCAMs, and their polysialylated forms,PSA-NCAMs,in the developing rat pituitary gland

Neural cell adhesion molecules (NCAMs) can undergo post-translational modifications, such as the addition of polysialic acid chains, thus generating PSANCAMs, which are expressed mainly during development. Since polysialylation considerably modifies NCAM adhesivity, expression of NCAMs and PSANCAMs has been investigated in the developing hypophysis by immunohistochemistry. At embryonic day 13 (E13), an antibody against NCAM outlined all cellular profiles in the entire Rathke's pouch; this labelling persisted until adulthood. NCAM expression increased in all lobes during development and concerned all pituitary cell types. In contrast, at E13, PSA-NCAMs were only detected in the neural lobe, solely constituted of pituicytes at this stage, and the tuberal lobe, the only lobe expressing hormonal mRNA at the same stage. PSA-NCAMs expression increased in the neural lobe at E17 with the arrival of the neurosecretory fibres and persisted into adulthood. In the anterior lobe, PSA-NCAMs appeared at E15 where their distribution was similar to that of the differentiating corticotrophic cells; at subsequent stages, their expression extended to the whole anterior lobe. Only two cell types, corticotrophic and somatotrophic cells, remained labelled in the adult gland. In the intermediate lobe, melanotrophic cells never expressed PSA-NCAMs but these were expressed on folliculo-stellate cells at birth, preceding the onset of innervation. These results suggest that NCAMs and PSA-NCAMs play a role in pituitary histogenesis, cell differentiation and neurointermediate lobe innervation.     

Anterior and posterior groups of portal vessels in the avian pituitary: incidence in forty nine species.

The hypophyseal portal vessels were studied in forty nine species of birds. The primary capillary plexus in the median eminence is single or divided into an anterior and a posterior plexus. Irrespective of whether the primary capillary plexus is single or divided, distinct, non-interconnected anterior and posterior groups of portal vessels are present in all the species investigated. The anterior group of portal vessels originates in the anterior region of the median eminence and breaks up into capillaries in the cephalic lobe of the pars distalis; the posterior group of portal vessels originates in the posterior region of the median eminence and breaks up into capillaries in the caudal lobe of the pars distalis. This type of regional distribution of portal vessels appears to be of general occurrence in the avian pituitary. The median eminence in the species investigated shows an AF-positive anterior region and an AF-negative posterior region. The pars distalis is differentiated into histologically distinct cephalic and caudal lobes. The arrangement of the portal vessels into anterior and posterior groups provides morphological basis for the view that the functions of the cephalic lobe may be controlled by the anterior median eminence, whereas those of the caudal lobe may be controlled by the posterior median eminence. However, experimental data available to date do not suggest a physiological significance to the widespread incidence of the regional distribution of portal vessels in the avian pituitary.     

Antiapoptotic Factor Humanin Is Expressed in Normal and Tumoral Pituitary Cells and Protects Them from TNF-α-Induced Apoptosis

Humanin (HN) is a 24-amino acid peptide with cytoprotective action in several cell types such as neurons and testicular germ cells. Rattin (HNr), a homologous peptide of HN expressed in several adult rat tissues, also has antiapoptotic action. In the present work, we demonstrated by immunocytochemical analysis and flow cytometry the expression of HNr in the anterior pituitary of female and male adult rats as well as in pituitary tumor GH3 cells. HNr was localized in lactotropes and somatotropes. The expression of HNr was lower in females than in males, and was inhibited by estrogens in pituitary cells from both ovariectomized female and orquidectomized male rats. However, the expression of HNr in pituitary tumor cells was not regulated by estrogens. We also evaluated HN action on the proapoptotic effect of TNF-α in anterior pituitary cells assessed by the TUNEL method. HN (5 µM) per se did not modify basal apoptosis of anterior pituitary cells but completely blocked the proapoptotic effect of TNF-α in total anterior pituitary cells, lactotropes and somatotropes from both female and male rats. Also, HN inhibited the apoptotic effect of TNF-α on pituitary tumor cells. In summary, our results demonstrate that HNr is present in the anterior pituitary gland, its expression showing sexual dimorphism, which suggests that gonadal steroids may be involved in the regulation of HNr expression in this gland. Antiapoptotic action of HN in anterior pituitary cells suggests that this peptide could be involved in the homeostasis of this gland. HNr is present and functional in GH3 cells, but it lacks regulation by estrogens, suggesting that HN could participate in the pathogenesis of pituitary tumors.     

Mammotrope heterogeneity in the pituitary gland of European ferret,Mustela putorius furo: An immunoelectron-microscopic study

Employing the superimposition technique of electron-microscopic immunocytochemistry ultrastructural heterogeneity of the mammotropes in the pituitary gland of the European ferret,Mustela putorius furo,was studied. On the basis of the size of their secretory granules, the mammotropes were classified into three subtypes, type-I, type-II and type-Ill, which may correspond to different developmental or physiological states of a single cell type. Simultaneous study of mammotropes and somatotropes in several pairs of serial semithin sections demonstrated the occasional occurrence of bihormonal somatomammotropes /mammosomatotropes which may represent a transitional stage of the progenitor stem-somatotrope during its differentiation into mammotrope; alternatively it may be a functional intermediate during the cross-transformation of somatotrope into mammotrope or vice versa.     

An immunocytochemical study of human pituitary mammotropes from fetal life to old age.

The objectives were to (a) describe the cytology and distribution of mammotropes in the human pituitary gland, (b) determine whether the mammotrope is a distinctive secretory cell type and (c) ascertain when it first appears in the fetal hypophysis. Identification of mammotropes was based primarily on the Sternberger peroxidase-antiperoxidase immunocytochemical method used with an antiserum to human prolactin. Hypophyses from 25 male and 6 female adults, and 21 fetuses ranging in gestational age from 6 to 23 weeks were studied. In the adult two morphological forms of mammotropes were observed. Mammotrope I possessed a small perikaryon that commonly was located centrally in parenchymal cell cords. From the perikaryon long cytoplasmic processes extended toward neighboring capillaries. Mammotrope I reached its highest incidence in the posterolateral zones of the pars distalis. Mammotrope II possessed a larger perikaryon with short processes; cells of this form were fewer and occurred chiefly in the anteromedian zone. Mammotropes with intermediate morphological features that prevented classification into categories I or II were common in some hypophyses. Both forms of mammotropes were present prepuberally (one 6-week and one 9-year-old male) and in adult males and females. Mammotropes were only slightly more prominent in females than males. Regression of mammotropes was evident in old age. Mammotropes were distinctly different from somatotropes, corticotropes, gonadotropes and thyrotropes. In the fetal hypophysis mammotropes appeared first at 14 weeks of gestational age and remaind few through 16.5 weeks. Their number increased greatly at 23 weeks.     

Contrasting effects of pituitary adenylate cyclase activating polypeptide (PACAP) on in vivo and in vitro prolactin and growth hormone release in male rats.

Pituitary adenylate cyclase activating polypeptide (PACAP) is produced by hypothalamic neurons which terminate within the median eminence suggesting that it may be a hypophysiotropic hormone. However, little endocrine activity has been ascribed to the peptide. Therefore we studied the effects of PACAP on prolactin (Prl) release from dispersed cultivated rat pituitary cells in vitro using conventional cultures as well as the reverse hemolytic plaque assay (RHPA). Furthermore the effects of the peptide on in vitro GH release were assessed. In addition, the activity of the peptide on in vivo release of Prl and GH was studied in hypothalamus-lesioned animals. PACAP dose dependently inhibited Prl release form dispersed pituitary cells in both, monolayer cell cultures and the RHPA, whereas GH secretion was not affected. In hypothalamus-lesioned rats which have high Prl levels due to the absence of hypothalamic dopamine, PACAP further stimulated Prl release. Serum GH increased more than 20 fold in response to the intravenous PACAP infusion. Thus in vitro (inhibition of Prl release, no effect on GH release) and in vivo (stimulation of both hormones) experiments yielded contradicting effects of PACAP on pituitary hormone release. We suggest that PACAP may stimulate the release of a paracrine, yet unknown factor which in the intact pituitary overrides the direct inhibitory action of PACAP on the lactotropes. The same or another paracrine factor may also enhance in vivo GH release. In cell culture the paracrine factor is diluted by the medium. Therefore the peptide never reaches effective concentrations which are present within the intact pituitary tissue.