Embryonic development of the bovine pineal gland (Bos taurus) during prenatal life (30 to 135 days of gestation)

The ontogenesis of the pineal gland of 30 bovine embryos (Bos taurus) has been analysed from 30 until 135 days of gestation by means of optical microscopy and immunohistochemical techniques. For this study, the specimens were grouped into three stages in accordance with the most relevant histological characteristics: Stage 1 (30 to 64 days of prenatal development); Stage 2 (70 to 90 days) and Stage 3 (106 to 135 days). In the cow, it is from 30 days of gestation that the first glandular outline becomes differentiated from the diencephalic ependyma of the third ventricle. This differentiation includes the phenomena of proliferation and multiplication of the ependymal cells that form the epithelium of the pineal outline in development. At 82 days of intrauterine life, in the interior of the pineal parenchyma, we witnessed some incipient pseudoglandular structures that at 135 days were well differentiated. The pineal parenchyma displays a cytology made up of two cellular types of structurally distinct characteristics: pinealoblasts and interstitial cells. Both cellular types begin differentiation at 70 days of embryonic development, the pinealoblasts being greater in number than the interstitial cells. The glandular stroma is formed from the capsular, trabecular and the perivascular connective tissue, filling the interparenchymal space. A dense network of capillaries, which drive across the trabecular connective tissue towards the central glandular zone where their density increases and their calibre is reduced, complete the glandular structure. GFAP positive cells were observed in the embryonic pineal parenchyma in stage 3. At 135 days of gestation, NPY positive fibers entered the pineal gland through the pineal capsule occupying a perivascular localization. Morphological studies of this nature are vital for future use as parameters, indicative of the functional activity of the bovine pineal gland during embryonic development.     

Histology of the venom gland of Trachinus draco (Actinopterygii,Trachinidae)

The structure of the venom gland of Trachinus draco was studied by histological techniques using light microscopy. New structures in the large glandular cells were detected, including concentric cytoplasmic laminae, basal vesicles, silver nitrate staining cytoplasmic granules, and a high affinity for wheat‐germ agglutinin lectin in the cytoplasm of mature large glandular cells.We also describe morphologic changes of the venom gland during thermal test. At low temperatures, the large glandular cells and their nuclei become enlarged, whereas at high temperatures the large glandular cells are smaller and their nuclei irregular in shape. Similarly, samples from fish captured during two opposite seasons showed differences in their nuclei, which were smaller in summer than winter. The larger growth of supporting cells at low temperatures, together with their aggregation during conditions of stress and their vacuoles secretion, suggest that supporting cells could be played several roles.     

Cells expressing preproenkephalin mRNA in the rat pineal gland are not serotonin-producing pinealocytes: Evidence usingin Situ hybridization combined with immunocytochemistry for serotonin

Summary 1. Preproenkephalin (PPEnk) mRNA expressing cells have been identified in rat pineal gland using radioactivein situ hybridization histochemistry. 2. Approximately 7% of the cells in the pineal gland (7.5±0.86, mean ± 95% CI) express PPEnk mRNA. These cells are distributed throughout the pineal as either scattered single cells or small groups of cells with large round or oval nuclei. 3. Usingin situ hybridization combined with ABC immunocytochemistry for serotonin (5-HT) in the same pineal sections, the PPEnk mRNA labeling cells are found not to be serotonin-immunoreactive cells. These data indicate that the PPEnk mRNA is expressed in a certain discrete subpopulation of cells in the rat pineal gland and these cells are not serotonin-producing pinealocytes. 4. The physiologic role of PPEnk-derived peptides in the pineal remains unknown. It is possible that these peptides either are synthesized and secreted as hormones or act as pineal paracrine signals.     

Morphologic changes in the pineal gland of rats exposed to continuous darkness

We examined the pineal structure of rats exposed to constant darkness (DD) at light microscopic level. Two groups of rats were exposed to 12:12 light/dark cycle (LD) or DD from their prenatal ontogenesis and then for 3 months after birth. The gland structure of DD rats was observed to have an active appearance. Some of the observed pinealocytes with light nuclei from DD rats were determined to contain double nucleoli. Nuclear area and perimeter of both dark and light types were greater in rats kept in DD than in LD. Rats exposed to DD had more cells with light nuclei and lesser cells with dark ones than rats kept in LD. No significant differences in nuclear characteristics of intermediate type were found between rats kept in LD and those kept in DD. The activity of mammalian pineal can be altered by light conditions to which the animal is exposed.     

Ultrastructure of male accessory glands of Chrysomya megacephala (Fabricius) (Diptera: Calliphoridae)

The ultrastructure of the male accessory glands of the blow fly, Chrysomya megacephala (Fabricius), was presented using light microscopy (LM), scanning electron microscopy (SEM), and transmission electron microscopy (TEM). A pair of accessory glands was separated at opposite sites. Morphometric results using LM yield evidenced no significant difference in the median of either length or width of the left and right glands. A significant increment in both length and width was seen to plateau between three to six days. SEM observation showed that the surface of the glands revealed a faint irregular groove pattern throughout, and it was occasionally penetrated by tracheoles. Each gland was a slender, elongated sac‐like tubule having apical rounded ends, with a slight constriction at the sub‐apical part of the gland being observed occasionally. TEM analyses of three‐day‐old males showed that the glands consisted of external capsular cells with a basement membrane underneath, glandular cells, and gland lumen. The capsular cell was flat and contained a nucleus with electron dense material in the nuclear envelope. The glandular cell, appearing as columnar, consisted of a vacuolated component that contained a large oval nucleus centrally or sub‐basally located, with dense mitochondria, numerous rough endoplasmic reticulum, and secretory vesicles containing electron‐lucent materials. In the gland lumen, the cross‐section through the middle portion revealed dense secretory materials, characterized by electron‐dense materials. Some sections revealed a large lumen where secretion accumulates within the delicate sac. The seven‐day‐old glands exhibited a remarkable change in the lumen, where the whole space contained a large amount of secretory materials, with the electron‐dense materials being characterized as similar to those observed in three‐day‐old glands. About four prominent types of secretions were observed on the basis of difference in electron‐density.     

Morphology of adenocarcinoma in situ and microinvasive adenocarcinoma of the uterine cervix. A cytologic and ultrastructural study

Adenocarcinoma in situ (AIS) and microinvasive adenocarcinoma of the uterine cervix and normal endocervical columnar epithelium were studied by cytology, morphometry and electron microscopy to identify differentiating features and to ascertain the cellular origin of cervical adenocarcinoma. Smears from AIS showed the characteristic cytology, consisting of glandular rosettes, palisading and crowded sheets; most nuclei had a relatively uniform oval shape. Smears from microinvasive adenocarcinoma showed more crowded sheets, with enlarged, round and irregular-shaped nuclei and prominent oval nucleoli. These nuclear features were confirmed by the morphometric results. Ultrastructurally, reserve cells in the normal tissues contained tonofibers and secretory granules and showed squamous and adenomatous features. The ultrastructural features of microinvasive adenocarcinoma were similar to those of well-differentiated invasive adenocarcinoma. The cells from both contained tonofibers and secretory granules. These findings suggested that the reserve cell is the cell of origin for cervical adenocarcinoma.     

Pituitary adenylate cyclase-activating polypeptide-immunoreactive (PACAP-IR) nerve fibers in the pig pineal gland

The present study demonstrates the occurrence of PACAP-immunoreactive (PACAP-IR) nerve fibers in different compartments of the pig pineal gland, including glandular capsule (where they form a very dense network) and subependymal tissue close to the pineal recess (moderate to dense meshwork of varicose fibers). Furthermore, several varicose fibers penetrate from the capsule into the connective tissue septa and then into the parenchyma, where they form unequally distributed, fine network and, in some cases, basket-like structures around pinealocytes. Some of the PACAP-IR nerve fibers, observed both in the habenular and posterior epithalamic areas, extend to the pineal gland. PACAP-IR cells could be demonstrated neither in the pineal gland, nor in epithalamic areas.     

Morphofunctional research on the interrenal gland of the frog Rana temporaria following arginine vasotocin administration

Studies have been made of the effect of injections of hypothalamic nonapeptide neurohormone, arginine vasotocin, on functional condition of the interrenal gland in mature frogs. In unoperated, sham-operated and in animals 10 days after hypophysectomy, single and especially three subsequent injections of arginine vasotocin (5 x 10(-9) M per 1 kg of the body weight) result in evident activation of glandular cells of the interrenal gland which is manifested in the increase of the volume of their nuclei and cytoplasmic area, as well as in the dilatation of the blood vessels. Activation of the interrenal gland in hypophysectomized frogs, which lack endogenous ACTH, indicate the direct para-adenohypophyseal influences of nonapeptide hypothalamic hormones on the activity of glandular cells in the peripheral endocrine glands, in particular, the interrenal gland of the grass frog.     

Prenatal Entrainment of Rat Testicular Malate Dehydrogenase Activity Circadian Rhythm

In mammals the photoperiodic synchronization of circadian system starts before birth. During fetal and neonatal period mothers relay the photoperiodic information to their litter. The maternal pineal melatonin 24 h cycle acts as a synchronizing signal. We have studied the effect of pineal maternal sympathetic denervation and administration of melatonin to mothers denervated during gestation on the prenatal synchronization of testicular malate dehydrogenase (MDH) activity circadian rhythm of the offspring 25 days after birth. When mothers were denervated at the 7th, 10th or 11th day of gestation, pups showed disruption of testicular MDH activity circadian rhythms. In contrast, no disruptive effect was observed when the mothers were denervated on the 12th or 14th day of gestation. When denervated mothers (7th day of gestation) were treated with a daily dose of melatonin from the 11th to the 14th day of gestation, pups showed a MDH activity circadian rhythm. The hormone failed to impose a daily phase when administered from the 9th to the 12th day of gestation. Results suggest that prenatal synchronization in the rat occurs very early in the development, before suprachiasmatic nuclei morphologic arrangement and functional activity begin.     

Demonstration of nerve fibers immunoreactive to peptide N-terminal histidine C-terminal isoleucine (PHI) and vasoactive intestinal peptide (VIP) in the pig pineal gland

The pineal functions are modulated by some neuropeptides including PHI and VIP. The presence of PHI-immunoreactive and VIP-immunoreactive nerve fibers in the pineal gland has been shown in several mammalian species. Both peptides influence the pineal serotonin N-acetyltransferase activity and melatonin synthesis. The aim of the present study was to examine the localization of PHI- and VIP-immunoreactive nerve fibers in the pig pineal gland. Four three-month old female pigs housed in natural light conditions, with free access to food and water, were used in the study. The pineals were fixed by perfusion with 4% paraformaldehyde in 0.1 M phosphate buffer. An immunohistochemical ABC streptavidin-biotin-complex method was used for the demonstration of PHI and VIP. PHI- and VIP-immunopositive nerve fibers were found in the pineal gland as well as in the habenular and posterior commissural areas. In the pineal gland, the density of PHI-immunoreactive nerve fibers was considerably higher than that of the fibers containing VIP. PHI- and VIP-immunopositive nerve fibers were more abundant in the cortical than in the medullary part of the gland. The nerve fibers formed bundles in the pineal capsule, from where they penetrated to the connective tissue septa and formed a dense meshwork surrounding blood vessels. In the parenchyma, PHI- and VIP-immunoreactive nerve terminals created baskets around clusters of pinealocytes. No PHI- or VIP-immunopositive cells were found in the pig pineal gland.     

Prenatal Entrainment of Rat Testicular Malate Dehydrogenase Activity Circadian Rhythm

In mammals the photoperiodic synchronization of circadian system starts before birth. During fetal and neonatal period mothers relay the photoperiodic information to their litter. The maternal pineal melatonin 24 h cycle acts as a synchronizing signal. We have studied the effect of pineal maternal sympathetic denervation and administration of melatonin to mothers denervated during gestation on the prenatal synchronization of testicular malate dehydrogenase (MDH) activity circadian rhythm of the offspring 25 days after birth. When mothers were denervated at the 7th, 10th or 11th day of gestation, pups showed disruption of testicular MDH activity circadian rhythms. In contrast, no disruptive effect was observed when the mothers were denervated on the 12th or 14th day of gestation. When denervated mothers (7th day of gestation) were treated with a daily dose of melatonin from the 11th to the 14th day of gestation, pups showed a MDH activity circadian rhythm. The hormone failed to impose a daily phase when administered from the 9th to the 12th day of gestation. Results suggest that prenatal synchronization in the rat occurs very early in the development, before suprachiasmatic nuclei morphologic arrangement and functional activity begin.     

Expression of IGF-I and -II mRNA in the brain and craniofacial region of the rat fetus

Insulin-like growth factors (IGF-I and -II) are present in the brain during development, with high levels of both being also found in the periphery particularly in the embryo. IGFs in the brain are believed to stimulate the proliferation of neuronal and glial precursors and their phenotypic differentiation. Using in situ hybridization, we have investigated the distribution of cells producing IGF-I and -II in the rat fetus during the second half of prenatal development with special emphasis on the peripheral and central nervous system. High levels of IGF-I mRNA were found in the olfactory bulb and in discrete neurons of the cranial sensory ganglia, notably in the trigeminal ganglion, as early as 13 days of gestation, in the pineal primordium of 18 day old fetuses, and in discrete groups of cells in the cochlear epithelium located laterally outside the forming spiral organ, in day 13 to 21 fetuses. High levels of IGF-II mRNA in the brain, besides the choroid plexus and the leptomeninges, were detected in hypothalamus, in the floor of the 3rd ventricle at all stages studied, in the pineal primordium at 18 days and in the pars intermedia of the pituitary or in the Rathke's pouch epithelium from which it is derived, with progressive fading towards the end of the gestation. In the peripheral nervous system the IGF-II mRNA was only found in association with the vascular endothelia of the ganglia. IGF-II mRNA in the nervous system was found in highly vascularized areas, meninges, blood vessels and choroid plexuses. It is thus associated with structures involved in the production of extracellular fluids and/or substrate transport and supply in the nervous tissues. A more specific role in the differentiation or fetal endocrine function should be considered for IGF-II in cells producing melatonin and melanocyte stimulating hormone (MSH) in the pineal and pituitary glands, respectively. The presence of IGF-I mRNA in the nervous system could be associated with fiber outgrowth and synaptogenesis in the cases of olfactory bulb and developing iris. The role of IGF-I in restricted populations of cells of the cochlear epithelium and in the pineal gland is unclear and requires further investigations including a search for IGF-I receptors in possible target cells. In the sensory ganglia, the presence of high levels of IGF-I mRNA eventually corresponds to the production, by post-translational processing, of the amino-terminal tripeptide of IGF-I, which might represent a neurotransmitter for these sensory neurons.     

Presence of a pineal nerve in sheep and rabbit fetuses

The presence of a nerve located just caudal to the pineal gland in the midsagittal plane is demonstrated in sheep and rabbit fetuses. This nerve lies freely in the subarachnoid space and extends from the pineal gland to a region of the CNS located dorsal to the rostralmost part of the subcommissural organ (SCO). In rabbit fetuses the nerve is obsered on days 23 and 24 of gestation; we suggest that it is an ontogenetic equivalent to the pineal nerve of anuran amphibians. The developmental fate of the mammalian fetal pineal nerve is dicussed.     

Time of origin of the rat pineal gland cells. A bromodeoxyuridine immunohistochemical study

The immunohistochemical detection of bromodeoxyuridine (BrdU) was used to study the time of origin of the cells in the pineal gland of the rat. A study was made involving 17 groups of 4 rats each, administered with a single dose of bromodeoxyuridine (BrdU, 25 mg/kg) in 7 phases of the embryonic period (E15 to E21) and in 10 postnatal phases (between P0 and P30), followed by determination in each rat of the number of visible immune-labeled cells in the pineal gland 60 days after birth. The results show that approximately 60% of the pineal cells underwent the last division(s) prior to differentiation in the prenatal period between E18 and E21. The rest of the pineal cells originated after birth, particularly in the first 5 postnatal days.     

Monoclonal antibody (SBU-1 and SBU-3) identification of cells dissociated from the sheep placentomal trophoblast.

We studied the localization of alpha-keratin in the sheep placenta using an alpha-keratin-specific monoclonal antibody (MAb) SBU-1, and examined the feasibility of using this MAb as a marker for determining the purity of isolated uninucleate cells from the placentomal trophoblast. At about 30-50 days of gestation the placentomal and interplacentomal uninucleate cells and some binucleate cells were stained by SBU-1, whereas only the apical region of the syncytial cytoplasm was stained with this MAb. Other cells stained included the uterine and endometrial glandular epithelial cells and fibroblast-like cells in the endometrium and chorionic villi. At about 100-130 days of gestation only the trophoblast uninucleate cells were stained by SBU-1. Approximately 60% of cells isolated from placentomes at 100-130 days of gestation were stained by SBU-1, and they had similar morphological features to the trophoblast uninucleate cells. The number of binucleate cells present was confirmed by their affinity for MAb SBU-3. These results show that MAb SBU-1 is an excellent marker for trophoblast uninucleate cells from placenta of sheep at the later stages of pregnancy.     

Calcitonin cells and unusual follicles of the thyroid of the indian grey mongoose, Herpestes edwardsi (Geoffroy).

Histological preparations of thyroid, parathyroid and thymus glands of Herpestes edwardsi were examined for calcitonin cells. They reveal that (1) the thyroid calcitonin cells are oval, rounded and rarely elongated in shape; these cells and their nuclei are distinctly larger than those of the follicular cells and their nuclei; (2) calcitonin cells are unevenly distributed in the thyroid, with the result that certain portions of the thyroid are completely devoid of these cells; (3) on an average, calcitonin cells are in a ratio of 10-15 cells/100 follicular cells; (4) the parathyroid and thymus glands do not display calcitonin cells, and (5) the thyroid gland displays unusual follicles of two categories, (a) follicles with ciliated epithelial cells and (b) follicles with squamous epithelium.     

Embryonic development of pineal gland vesicles: a morphological and morphometrical study in chick embryos.

Pineal cell aggregates in 5, 10 and 15 day-old chick embryos have been studied. Cell aggregates were classified into rosettes or vesicles and spheroid and ellipsoid vesicles distinguished. The number of pineal vesicles per unit of surface (vesicle density) was determined in three pineal portions: apical, anterior and posterior. By day 5, only cellular rosettes were found, mainly in the apical portion. After 10 and 15 days, the presence of rosettes was occasional. The posterior wall showed only small spheroid vesicles, while in the apical and anterior areas ellipsoid vesicles were also observed. Moreover, the spheroid/ellipsoid vesicle ratio increased from the 10th to the 15th day of incubation. The vesicle density decreased between the 10th and 15th day because of the increase in both vesicle and pineal size, without changes in the total number of vesicles. The results suggest that changes in vesicle morphology and density could be related to the functional activity of the pineal gland during embryonic development.     

Prenatal development of the human submandibular gland

The prenatal development of the human submandibular gland has been investigated in 26 fetuses from the 10th week of gestation to full term. At 10-12 weeks, the glandular elements (primitive ducts and acini) were immature and surrounded by a loose mesenchyme. The acinar cell population increased gradually till the age of 32 weeks, and the rate of increase was diminished thereafter. At 16 weeks, intercalated and striated ducts were distinguished and their number increased till the age of 32 weeks when their number seemed to be stabilized. The development of the granular convoluted tubule cells from the proximal segments of striated ducts occupied the later stages of development. They appeared around the age of 20 weeks and proceeded till full term. At birth, the gland appeared devoid of mucous acini and fat cells and the secretory end-pieces were of the serous type. During the second trimester, periodic acid-Schiff- and alcian blue-positive secretory materials appeared in the epithelial cells of both ducts and acini, and in their lumina. This secretory activity was transitory and disappeared around the age of 28 weeks. The possible function of these secretory products is discussed.     

Regional distribution and cellular expression of tryptophan hydroxylase messenger RNA in postmortem human brainstem and pineal gland

The characterization and cellular localization of tryptophan hydroxylase mRNA in the human brainstem and pineal gland were investigated by using northern blot analysis and in situ hybridization histochemistry. Northern analysis of human pineal gland revealed the presence of two mRNA species that were absent in RNA isolated from human raphe. In situ hybridization experiments revealed very dense hybridization signal corresponding to tryptophan hydroxylase mRNA in cells throughout the pineal gland. In contrast, tryptophan hydroxylase mRNA was heterogeneously distributed in neurons in the dorsal and median raphe nuclei. Within the dorsal raphe, the ventrolateral and interfascicular subnuclei contained the greatest number of tryptophan hydroxylase mRNA-positive neurons. Also, the cellular concentration of tryptophan hydroxylase mRNA varied widely within the dorsal and median raphe. Comparison of the cellular concentration of tryptophan hydroxylase mRNA between the pineal gland and the raphe nuclei revealed an 11- and 46-fold greater average grain density of tryptophan hydroxylase mRNA positive cells in the pineal gland compared with the dorsal and median raphe, respectively. These findings are the first to demonstrate the cellular localization of tryptophan hydroxylase mRNA in the human brain and pineal gland as well as heterogeneity in the cellular concentration within and between these tissues.