Differential C-heterochromatin distribution in two species of freshwater fish, Anabas testudineus (Bloch.) and Puntius sarana (Hamilton.)

Anabas testudineus (2n = 46) had the more conserved pattern of its C-heterochromatin distributed mainly in the centromeric region, whereas Puntius sarana (2n = 50) exhibited a rather unorthodox pattern, many chromosomes showing interstitial, some telomeric and a few chromosomes showing centromeric C-band localization. Further, lateral asymmetry in distribution of heterochromatin was also noted in two pairs of chromosomes in P. sarana. The possible implications of the differential distribution noted in these two species has been discussed.     

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.     

Cyclical changes in the renin-angiotensin-aldosterone system during the menstrual cycle of the baboon (Papio hamadryas)

Abstract: This study characterizes the renin-angiotensin-aldosterone system during the normal menstrual cycle in the baboon. Ten animals received a daily dose of an ACE inhibitor or placebo in a randomized blind cross-over design. Data were obtained during the mid-follicular and early luteal phases of normal non-pregnant menstrual cycles. All examinations and blood collections were performed with ketamine sedation: 7–kg by im injection. Blood pressure was recorded by sphygmomanometer. Serum ACE activity was measured by spectrophotometry. Aldosterone (ALDO), angiotensin I (AI), and angiotensin II (AII) were measured by radioimmunoassay. Plasma renin activity (PRA) was measured by AI generation. The renin-angiotensin-aldosterone system was found to be activated in the follicular phase and suppressed during the luteal phase of the normal non-pregnant menstrual cycle in the baboon.     

Ultrastructure of rat pituitary LH gonadotrophs in relation to serum and pituitary LH levels following repeated LH-RH stimulation

The effects of single and repeated LH-RH injections at 120 min intervals on female rat LH gonadotrophs and on pituitary and serum LH levels were investigated using electronmicroscopy and radioimmunoassay. A temporary stimulation of granule release, of protein and new granule synthesis and of the accumulation of lysosomal structures was found in LH cells after the first LH-RH injection. The temporary stimulations were massively enhanced after the second injection. These consecutive yet in their time-sequence overlapping processes account for the initial depletion of secretory granule content (3--15 min after LH-RH injection), for the subsequent regranulation and accumulation of granules above control levels (60--120 min after injection) and also for the reduction in the number of granules to control levels (150 min after LH-RH injection and thereafter). Increased polymorphic lysosomal structures are believed to be responsible for this reduction of excess granules. The amount of assayable pituitary and serum LH generally corresponds with the morphological changes observed in LH-gonadotrophs, thus further substantiating the above observations. A schema which summarizes the observed morphological and hormonal changes in their time-sequence in response to LH-RH stimulation depicts the short-term regulation of secretory processes in female gonadotrophs.     

Cell volume changes in relation to the cell cycle of differentiating erythroleukemic cells

Murine erythroleukemic cells (MELC) were synchronized by sequential exposure to thymidine and hydroxyurea. Upon removal from hydroxyurea, cells cultured with or without agents that induce erythroid differentiation, such as hexamethylene bisacetamide (HMBA) or dimethylsulfoxide (Me₂SO), proceed through S, G2 and mitosis with the same kinetics: S phase averages 5 h and G2 plus mitosis, 2 h. Cells cultured with HMBA and Me₂SO remain in the subsequent G1 for 5–7 h, compared with an average of only 3 h for cells cultured without inducer. Modal cell volume doubles as the cells proceed from G1 to G2. During the inducer-mediated prolonged G1, MELC retain a small cell volume. In cultures of non-synchronous MELC, inducers also increase the G1 fraction, as well as the proportion of small cells. An Me₂SO-resistant MELC variant (DR10), cultured with Me₂SO, shows little prolongation of G1 and little difference in the modal cell volume compared with cells without inducer. However, HMBA, which induces differentiation of DR10 cells, prolongs G1 and increases the proportion of small cells. These studies indicate that early changes in cell volume associated with induction of MELC to differentiate, in large part reflect alterations in the cell cycle. Evidence is presented which suggests that only one round of DNA synthesis in the presence of inducer may be necessary to initiate differentiation.     

Annual cyclical changes in the testicular activity of an Indian freshwater major carp, Labeo rohita (Hamilton)

Different male germ cells identified on the basis of histological and cytological characteristics in the testicular lobules of Labeo rohita have been grouped into spermatogonia, primary spermatocytes, secondary spermatocytes, spermatids and spermatozoa. The seasonal changes of the testis in L. rohita have been described according to its morphological peculiarities as well as to its variations in gonadal volumes, GSI values and frequency percentages of the different male germ cells occurring in the testicular lobules. Consequently, the entire testicular cycle in L. rohita may be categorised into 4 distinct phases viz., growth, maturation of pre-spawning, spawning, and resting or post-spawning phases.     

Neuropeptide glutamic-isoleucine (NEI) specifically stimulates the secretory activity of gonadotrophs in primary cultures of female rat pituitary cells

The neuropeptide EI (NEI) is derived from proMCH. It activates GnRH neurons, and has been shown to stimulate the LH release following intracerebroventricular administration in several experimental models. The aim of the present paper was to evaluate NEI actions on pituitary hormone secretion and cell morphology in vitro. Pituitary cells from female rats were treated with NEI for a wide range of concentrations (1–400 × 10⁻⁸ M) and time periods (1–5 h). The media were collected and LH, FSH, PRL, and GH measured by RIA. The interaction between NEI (1, 10 and 100 × 10⁻⁸ M) and GnRH (0.1 and 1 × 10⁻⁹ M) was also tested. Pituitary cells were harvested for electron microscopy, and the immunogold immunocytochemistry of LH was assayed after 2 and 4 h of NEI incubation. NEI (100 × 10⁻⁸ M) induced a significant LH secretion after 2 h of stimulus, reaching a maximum response 4 h later. A rapid and remarkable LH release was induced by NEI (400 × 10⁻⁸ M) 1 h after stimulus, attaining its highest level at 2 h. However, PRL, GH and FSH were not affected. NEI provoked ultrastructural changes in the gonadotrophs, which showed accumulations of LH-immunoreactive granules near the plasma membrane and exocytotic images, while the other populations exhibited no changes. Although NEI (10 × 10⁻⁸ M), caused no action when used alone, its co-incubation with GnRH (1 × 10⁻⁹ M), promoted a slight but significant increase in LH. These results demonstrate that NEI acts at the pituitary level through a direct action on gonadotrophs, as well as through interaction with GnRH.     

Localization of sialic acid-containing hormones in GTH cells and ACTH cells of the rat anterior pituitary

Summary The localization of sialic acid-containing substances in the rat anterior pituitary gland has been studied by light and electron microscopy, using a peroxidase-labeled lectin (Limulus polyphemus agglutinin: LPA) which binds specifically to sialic acid residues. LPA stains two types of anterior pituitary cells: (1) round or ovoid cells which are also positively stained with anti-hCG (GTH cell), and (2) small, stellate cells which are unstained with anti-hCG (ACTH cell). All of the LPA-positive cells can be distinguished from TSH cells which are identified by the use of anti-hTSH. On ultrathin sections directly stained with LPA using the postembedding method, the reaction is confined to the secretory granules in GTH cells, and ACTH cells. Of two types of secretory granules in GTH cells, the larger one is intensely stained, whereas the smaller type shows only weak staining with LPA. Since follicle-stimulating hormone (FSH) is known to have high sialic acid contents, the results suggest possible detection of FSH with a technique other than immunohistochemistry. Furthermore, if the sialic acid-containing substances in GTH cells represents FSH, then these results support the hypothesis that LH cells and FSH cells are one cell type.This research was supported by grants from the Ministry of Education of Japan     

Crystalline inclusions in the pituitary gonadotrophs of estrogen-primed, spayed rhesus monkeys (Macaca mulatta)

17-β-Estradiol or cholesterol was subcutaneously implanted in bilaterally oophorectomized rhesus monkeys for 37 days, after which the pituitary glands were analyzed ultrastructurally for evidence of granule disposal. Plasma estrogen returned to a physiologic level of 97.6 ± 2.8 pg/ml in the estrogen-treated animals but not in the cholesterol-treated controls. The hypertrophied gonadotrophs in the controls contained numerous large vesicles and a few secretory granules. The gonadotrophs in the experimental animals showed a reduction in cell size, a diminution of the dilated endoplasmic reticulum, and an accumulation of secretory granules. Moreover, crystalline inclusions composed of parallel and orthogonally arranged fibers were present in some gonadotrophs, most frequently in animals with lower levels of circulating estrogen. These inclusions are thought to be involved in the process of intracellular secretion in monkeys.     

Cyclic changes in the pituitary gland in correlation with the testicular cycle in a hill-stream teleost, Schizothorax richardsonii (Gray and Hard).

In Schizothorax richardsonii, the pituitary gland has been studied in correlation with the phases of the testicular cycle. The cyanophils of the proximal pars distalis show hypertrophy, hyperplasia, granulation, degranulation and vacuolisation in concurrence with the spermatogenic cycle. Secretory activity of the cyanophils controls the maturation of testes, which indicates that these cyanophils are gonadotrops. The formation of sperm mother cells, however, does not appear to depend on the secretory activity of the pituitary. During spawning and regression phases, the azocarminophilous granules in the cytoplasm of the cyanophils show quantitative variation. The migratory chromophil cells show numerical variation, suggesting their involvement in the reproductive cycle. Neurosecretory material also appears to play some part in reproduction.     

Immunoreactivity of gonadotrophs (FSH and LH Cells) and gonadotropin subunit gene expression in the male chub mackerel Scomber japonicus pituitary during the reproductive cycle

The gonadotropins (GtHs), follicle-stimulating hormone (FSH) and luteinizing hormone (LH), are heterodimers composed of a common α subunit (GPα) and a unique β subunit (FSHβ or LHβ); they are synthesized in and secreted from gonadotrophs (FSH and LH cells) in the pituitary. Little is known about the roles of FSH and LH during spermatogenesis in perciform fishes. In this study, we examined immunoreactive changes in FSH and LH cells, and changes in the gene expression of the three gonadotropin subunits in the pituitary of male chub mackerel Scomber japonicus during testicular development. FSHβ-immunoreactive (ir) and LHβ-ir cell area were measured immuno-histochemically based on the FSH and LH cell-occupying area in the proximal pars distalis. The FSHβ-ir cell area increased significantly during spermiation, while FSHβ mRNA levels, already high at the beginning of spermatogenesis, increased further, peaking during spermiation. In contrast, LHβ-ir cell area and LHβ mRNA levels, which were low at the beginning of spermatogenesis, increased significantly during late spermatogenesis, peaking during spermiation. For both FSH and LH, GtHβ-ir cell area and GtHβ mRNA levels decreased until gonadal resting. GPα mRNA levels showed similar changes to LHβ mRNA levels. These results suggest that in the chub mackerel, FSH may play an important role in the early and late phases of spermatogenesis, and that LH may play a role during late spermatogenesis and spermiation. Moreover, our results demonstrate that changes in GtHβ-ir cell area were accompanied by similar changes in the expression of the FSHβ and LHβ genes, both of which increased during testicular development.     

Differential production and regulation of gonadotropins (GTH I and GTH II) in the pituitary gland of rainbow trout,Oncorhynchus mykiss,during ovarian development

Summary Biosynthesis of salmon gonadotropins, GTH I and GTH II, during ovarian development, were examined by means of in situ hybridization histochemistry and indirect immunocytochemistry. In rainbow trout pituitary glands, expression of GTH I- and II-subunit genes appeared separately in distinct cells (GTH I- and GTH II-cells), whereas the GTH -subunit gene was expressed in both cell-types. In the GTH I-cells, coordinated increases in GTh, and I messenger ribonucleic acids (mRNAs) occurred coincident with the onset of vitellogenesis, indicating active synthesis of GTH I during vitellogenesis. In contrast, in the GTH II-cells, both GTH -and II-mRNA signals markedly increased from a later stage of vitellogenesis and persisted throughout oocyte maturation and ovulation, supporting the idea that GTH II is actively synthesized as a maturational GTH. GTH -mRNA levels in the GTH I-cells selectively decreased prior to final oocyte maturation, although I-mRNA levels remained elevated, thus suggesting a decline of biosynthesis of GTH I after vitellogenesis. These findings clarify how the synthesis of GTH I and GTH II are coordinated in the piscine pituitary, and indicate that the expression of GTH subunit genes during gametogenesis is regulated differentially in a cell-specific manner, both temporally and spatially.     

Cyclical changes in the fine structure of bovine endometrial gland cells

Summary Cyclical changes in bovine endometrial gland cells were investigated in six heifers, three at estrus and three at day twelve of the estrous cycle in the luteal phase. The epithelium is generally low at estrus but high in the luteal phase. There are ciliated and non-ciliated cells. The ciliated cells are fewer and lighter and show inconspicuous cyclical changes.The secretory cells show more prominent changes. At estrus, their free border is flat with short microvilli. The conspicuous rough-surfaced endoplasmic reticulum may synthesize protein for later secretion. The Golgi complex seems inactive. The high number of cytosegresomes and dense bodies might express cell regression caused by endocrine changes.In the luteal phase, the cells are lighter with long microvilli. The Golgi apparatus shows vacuoles and immature secretory droplets. Secretory vacuoles with light contents occur in the apical cytoplasm. Some of them appear to discharge their contents into the lumen. This is interpreted as evidence of merocrine secretion. Accumulations of tubular, smooth-surfaced endoplasmic reticulum, masses of glycogen granules, and several fat droplets are present.Some lymphocytes and degranulated granulocytes are seen near the basement membrane, more frequently at estrus.Financial support for this study was received from Anslaget för främjande av medicinsk forskning vid Veterinärhögskolan.The authors express their gratitude to Prof. A. Bane and Dr. J.-E. E. Ringmar, Department of Obstetrics and Gynecology, for their help with the selection and clinical control of the animals and for keeping them in good condition.Post doctoral fellow, No. 43-KO-52 (1968) from the Educational Ministry of Japan.     

Binding and internalization of native gonadoliberin (GnRH) by anterior pituitary gonadotrophs of the rat

Summary To identify anterior pituitary cell types containing GnRH binding sites and to study the internalization process of this peptide by target cells under physiological conditions, autoradiography was performed on rat anterior pituitaries removed at specific time intervals (2–60 min) after intravenous injection of mono-radioiodinated ¹²⁵I-GnRH into intact males. At electron-microscopic level, gonadotrophs and lactotrophs appeared to contain silver grains. Concomitant administration of an excess of unlabeled GnRH with the radioiodinated hormone prevented this localization indicating the specificity of the reaction. The time-course study in gonadotrophs showed that 2 min after injection silver grains could be found over the plasma membrane, secretory granules and nuclear membrane. Similar results were observed 5 and 15 min after injection. Extensive label was observed over the nucleus and nuclear membrane 15 to 60 min after injection. The injection of a radioiodinated GnRH agonist [D-Trp⁶, Pro⁹ (Net), DesGly¹⁰]-GnRH produced comparable results. In contrast, the injection of ¹²⁵I-[D-pGlu¹, D-Phe², Trp^3,6]-GnRH, an antagonist of GnRH, produced positive labeling only at the plasma membrane without internalization. These results indicate that, after binding with receptors on the plasma membrane, GnRH is rapidly internalized, accumulating in secretory granules, and localizing over the nuclear membrane and later, in the nucleus. Association of radioactivity with secretory granules could be related to a specific action of GnRH at this level or to receptor recycling, and presence of label in the nucleus may be related to stimulation of neosynthesis of LH and GnRH receptors.     

Cellular associations of pituitary gonadotrophs in a rodent (Lagostomus maximus maximus) with photoperiod-dependent reproduction

The morphological characteristics and percentage of the cellular associations between gonadotrophs (LH- and FSH-secreting cells) and other cellular types were studied in pituitary pars distalis of adult male viscachas (Lagostomus maximus maximus) by double immunohistochemistry using specific antibodies to LH, FSH, PRL, GH, ACTH, TSH and S-100 protein (by folliculostellate cells; FSC), during long and short photoperiods. Bihormonal gonadotrophs were observed in ventro-medial and dorsal regions, interspersed between monohormonal gonadotrophs, and their number increased in short photoperiod. LH- and FSH-gonadotrophs were found around lactotrophs, enclosed by somatotrophs in the dorsal region, and associated with irregular corticotrophs. Gonadotrophs and thyrotrophs were associated along blood vessels and follicular structures. The cytoplasmic prolongations of FSC were in contact with both gonadotrophs. The percentage of LH–FSH, LH–ACTH, LH–FSC, FSH–LH, FSH–PRL, FSH–GH, FSH–ACTH, FSH–TSH and FSH–FSC associations decreased, whereas LH–PRL increased in short as compared to long photoperiod. The most abundant associations were LH–GH and LH–TSH during long photoperiod, but LH–GH and LH–PRL during short photoperiod. FSH–GH and FSH–PRL were the most numerous associations, and LH–FSC and FSH–FSC were the less abundant ones in both photoperiods. These results provide the morphological evidence for specific cellular associations between gonadotrophs and other cellular types of viscacha pituitary.