Immunocytochemical localization of somatostatin in the brain of the lizard,Ctenosauria pectinata

Summary The brain of the lizard, Ctenosauria pectinata, was studied light microscopically using an immunocytochemical staining method that is specific for neurohypophysial hormones and somatostatin. It was shown that the telencephalon and particularly the diencephalon contain somatostatin-producing perikarya, while somatostatinergic fibers occur in the entire brain. Similar to the situation in other vertebrates, somatostatin neurons in Ctenosauria pectinata form a population distinct from the neurohypophysial hormone-producing neurons. The small-sized somatostatin neurons were found in the cortex and the hypopallium of the telencephalon, and in two distinct clusters in the diencephalon: (1) ventral from, and partially overlapping with, the classical neurosecretory para ventricular nucleus; and (2) in the region of the infundibular (tuberal) nucleus. Somatostatin fibers were found among the classical neurosecretory fibers of the supraoptico-paraventricular system (tract, median eminence, neural lobe), near to and within the epiphysis, in the septum, in the vicinity of the tectum opticum and the cerebellum, and in the tegmentum.This investigation was supported by a grant from the Belgian Nationaal Fonds voor Geneeskundig Wetenschappelijk Onderzoek     

Immunocytochemical localization of somatostatin in human brain

The distribution of somatostatin (SRIF) was examined in normal human forebrain, using thick vibratome cut sections. The unlabeled antibody enzyme method of immunocytochemistry revealed a widespread distribution of SRIF immunoreactive neurons and fibers throughout the septum, diencephalon and corpus striatum. Within the septum SRIF neurons and fibers were observed in the medial and lateral septal nuclei, the nucleus of the diagonal band, the nucleus accumbens and the bed nucleus of the stria terminalis. SRIF neurons and fibers were found in several hypothalamic and anterior thalamic nuclei as well as all regions of the corpus striatum. An interesting collection of SRIF immunoreactive neurons and processes were observed forming a wide band extending anteriorly from the lateral preoptic area through the lateral hypothalamus and substantia innominata posteriorly. This report on the localization of immunoreactive SRIF in the human forebrain extends previous anatomical findings and lends morphological support to recent biochemical studies.     

Immunocytochemical localization of somatostatin and vasotocin in the brain of the Pacific hagfish,Eptatretus stouti

Summary The brain of the Pacific hagfish, Eptatretus stouti, was studied immunocytochemically using antisera against somatostatin (SRIH), arginine vasopressin (AVP), and adrenocorticotropic hormone (ACTH). SRIH-immunoreactive perikarya were distributed bilaterally in the postoptic nucleus and in the hypothalamic nucleus. Although several short, stained fibers were observed in the vicinity of the perikarya, SRIH-immunoreactivity was not found in the neurohypophysis, nor in other parts of the brain. On the other hand, presumed arginine vasotocin (AVT) perikarya were distributed in an arc-shaped region extending from the posterior part of the preoptic nucleus to the anterior-most end of the hypothalamic nucleus and projected their fibers to the neurohypophysis. Most presumptive AVT perikarya were located close to the paired prehypophysial arteries near the anterior end of the postoptic nucleus. In the neurohypophysis, abundant presumptive AVT-fibers terminated in the posterior dorsal wall, although some fibers terminated in the anterior dorsal wall and only a few fiber endings were found in the ventral wall. No ACTH-positive cells were detected in the hagfish brain or in the pituitary gland.Supported from a grant from the National Science Foundation PCM 8141393     

Immunocytochemical localization of somatostatin in cell bodies of the rat hypothalamus.

Cell bodies of small to moderate-sized neurons in the female rat hypothalamus were stained specifically for somatostatin (SRIF) by means of the unlabeled antibody-peroxidase-antiperoxidase immunocytochemical method. SRIF-positive perikarya were scattered throughout the periventricular nucleus in a limited region extending from the middle of the optic chiasm to the rostral margin of the median eminence. The same neurons were revealed with either rabbit (R) or guinea pig (GP) anti-SRIF antisera. Positive cell bodies were more readily assessed with GP antibodies because nonspecific background staining was much less with these than with R anti-SRIF. Positive perikarya were not observed in other hypothalamic nuclei and ependymal elements were also immunocytochemically negative.     

Ovarian folliculogenesis in the oviparous Mexican lizard Ctenosaura pectinata

The ovarian follicles of Ctenosaura pectinata exhibit a clear seasonal cycle in morphology. Early in development, each oocyte is surrounded by a granulosa composed of a single layer of cuboidal or squamous cells and thin thecal layers. As folliculogenesis progresses, the granulosa becomes multilayered and composed of three distinct cell types. After vitellogenesis begins and active sequestration of yolk into the ooplasm is initiated, the granulosa is reduced to a single cell type. We observed a striking change in the appearance of the ooplasm during folliculogenesis. Early ovarian development is characterized by an ooplasm with homogeneously distributed fine fibrils, but as development progresses, the ooplasm contains dense clumps of fibers aggregated into distinctive bundles. The ooplasm displays further complexity in morphology as previtellogenic growth continues and as different regions exhibit various combinations of fibers and vacuoles. Yolk platelet formation is complex, with distinctive stages generating platelets with varying morphologies. © 1996 Wiley-Liss, Inc.     

Immunocytochemical localization of the glucose-transport protein in mammalian brain capillaries

Summary The endothelial cells of mammalian brain capillaries, which form the anatomical basis of the blood-brain barrier, have been investigated by immunocytochemical methods to determine the distribution of the glucose-transport protein. A monoclonal antibody raised against the intact human erythrocyte glucose-transport protein and polyclonal antibodies raised against a synthetic peptide corresponding to the C-terminal sequence of the human erythrocyte glucose-transport protein were used for immunofluorescent staining of isolated human and bovine cerebral cortex microvessels. The pattern of fluorescence with both antibodies demonstrated the antigen to the distributed throughout the plasma membrane of the capillary endothelial cells. These results provide further evidence for the homology between the human erythrocyte and brain capillary glucose-transport protein, and confirm its abundance in brain capillaries.     

Immunocytochemical localization of d-amino acid oxidase in rat brain

d-amino acid oxidase (d-AAO) is a peroxisomal flavoenzyme, the physiological substrate and the precise function of which are still unclear. We have investigated D-AAO distribution in rat brain, by immunocytochemistry, with an affinity-purified polyclonal antibody. Immunoreactivity occurred in both neuronal and glial cells, albeit at different densities. Glial immunostaning was strongest in the caudal brainstem and cerebellar cortex, particularly in astrocytes, Golgi-Bergmann glia, and tanycytes. Hindbrain neurons were generally more immunoreactive than those in the forebrain. Immunopositive forebrain cell populations included mitral cells in the olfactory bulb, cortical and hippocampal neurons, ventral pallidum, and septal, reticular thalamic, and paraventricular hypothalamic nuclei. Within the positive regions, not all the neuronal populations were equally immunoreactive; for example, in the thalamus, only the reticular and anterodorsal nuclei showed intense labelling. In the hindbrain, immunopositivity was virtually ubiquitous, and was especially strong in the reticular formation, pontine, ventral and dorsal cochlear, vestibular, cranial motor nuclei, deep cerebellar nuclei, and the cerebellar cortex, especially in Golgi and Purkinje cells.     

Immunocytochemical localization and ultrastructural study of gonadotroph cells in the female desert lizard Uromastyx acanthinura

The pars distalis from the pituitary gland of adult female desert lizards (Uromastyx acanthinura), captured during vitellogenesis (late may) and hivernal period, was studied with immunocytochemical methods using specific antisera against human FSH (hFSH) and LH (hLH). The immunostaining with anti-hLH and anti-hFSH allowed the identification of only FSH-like containing cells. The FSH-like immunoreactive cells were affected differently by a physiological stage and showed some heterogenous cytological characteristics. During vitellogenesis, four aspects of rostral FSH-like immunoreactive cells could be recognized. The expression of FSH-like in mainly immunoreactive cells was parallel to an intense synthetic activity and to the presence of ultrastructural features indicating an intense release of the hormone. This release was considerably altered in winter, the immunoreactive cells stored an important amount of secretion granules which increased in size and undergo a crinophagic process.     

Estrogen localization in the brain of the lizard, Anolis carolinensis (1)

Estrogen induces reproductive behavior in lizards by acting on specific areas of the brain. Neural areas that selectively concentrate radioactivity after 3-h-estradiol administration were mapped using thawmount autoradiography. Major accumulations of hormone-concentrating cells occur in the medial preoptic area, basal hypothalamus, amygdala (n. vetromedialis telencephali) and in and around the torus semicircularis. Numerous smaller accumulations are also described and the topographical distribution is compared with that of avian and mammalian species.     

Ontogenetic variation in digestion by the herbivorous lizard Ctenosaura pectinata

I tested the hypothesis that an animal with an ontogenetic diet shift must have different digestive efficiencies for foods that correspond to its diet shift, so that nutrient and energy extraction are maximized. The iguanine lizard Ctenosaura pectinata undergoes an ontogenetic diet shift from eating insects as a juvenile to plants as an adult. When fed six different pure foods from the natural diets of different age classes, C. pectinata assimilated nutrients and energy differently depending on food type and age class. Extraction of energy and nutrients in insect larvae was maximized by juvenile lizards. Calcium, phosphorus, and energy were readily assimilated from flowers and fruit by immature and adult lizards. Magnesium levels were highest in leaves and were extracted by immature and adult lizards, but xenobiotic effects of one plant leaf (Croton suberosus), eaten by adults, killed juvenile lizards. Although juvenile C. pectinata ate some flowers (Senna wislizenii) naturally, they were less efficient at digesting cell walls from these plant parts than were older lizards. Ontogenetic changes in ctenosaur digestive physiology were not the result of a trade-off involving ecological costs of different foods; rather, each age class preferred a diet that maximized its physiological benefit.     

雌激素Beta受体在大鼠脑内表达的免疫组化定位研究

为了探讨雌激素作用于神经系统的机理,采用硫酸镍铵增强显色的免疫组化SP法研究了新的雌激素受体（ER－β）在成年雌雄大鼠脑内的分布。研究证实ER－β免疫阳性物质主要位于神经元的细胞核内,但在个别脑区也可在胞浆甚至突起内检测到。最强的ER－β免疫阳性信号见于前嗅核、大脑皮质、小脑浦肯野细胞、斜角带垂直部、蓝斑和三叉神经运动核等部位;中等强度的染色见于隔内侧核、杏仁外侧核、黑质、中央灰质等部位;较弱的阳性反应见于下丘脑与杏仁复合体的部分核团。在一些部位还存在表达水平甚至细胞内定位模式的性别差异,如前庭上核内的表达只见于雌性;雄性大鼠三叉神经运动核内ER－β蛋白主要表达于胞浆内,细胞核为阴性;而在雌性大鼠该部位ER－β蛋白主要位于细胞核等。以上结果表明ER－β蛋白在大鼠脑内分布广泛并具有一定的性别差异,在与学习记忆有关的脑区如大脑皮质和基底前脑内有很高的表达,提示在脑组织内雌激素可能通过ER－β这一新的信号途径发挥多种重要的调控作用,如学习记忆等。     

Immunocytochemical localization of vasotocin-like immunoreactivity in the brain of the cartilaginous fish,Scyliorhinus caniculus

Summary The distribution of vasotocin-like peptides in the central nervous system of the cartilaginous fish Scyliorhinus canicula was determined by indirect immunofluorescence and peroxidase anti-peroxidase techniques, using a specific antiserum raised in rabbits against synthetic vasotocin. Immunoreactive perikarya were mainly detected in the anterior hypothalamus, within the midcaudal part of the preoptic nucleus. The most rostral positive cell bodies were located in the dorso-lateral parts of the preoptic area, whereas at a more caudal level, they took a ventro-medial position within the deepest layers of the nucleus. Throughout the preoptic region these cells varied in shape according to their location. Occasionally, scattered vasotocin-like immunopositive cells were also identified in the nucleus periventricularis hypothalami. Vasotocin immunoreactivity was detected in numerous varicose nerve fibers of the preopticohypophysial tract. These fibers were seen to course through the medio-basal hypothalamus and caudally, after having passed the hypophysial stem, they reached the neurointermediate lobe of the pituitary. Numerous immunoreactive fibers were also observed within the rostro-medial region of the median eminence. At this level the fibers were in close proximity to the capillary loops. In the preoptic region, some stained cells exibited short processes that appeared to contact non-reactive perikarya. By comparing the distribution of vasotocin- and corticotropin-releasing factor immunoreactivity on adjacent then serial sections, it was revealed that these peptides, in S. canicula, do not coexist in the same perikarya. The present results, are compared with those obtained in other vertebrate groups, and their possible functional implications are discussed.     

Immunocytochemical localization of enkephalin in the lateral septum of the guinea-pig brain

By the use of antisera to met-enkephalin and leu-enkephalin, enkephalin-containing structures were visualized in the lateral septum of the guinea-pig brain. The present results do not reveal immunoreactive perikarya in this area. The immunostaining is exclusively located in numerous nerve fibers and endings mostly encompassing neuronal perikarya, which accounts for the fact that at the light-microscopic level cellular somata appear to be immunostained. The immunoreactive terminals and fibers contain granules approximately 110 nm in diameter and synaptic vesicles. The origin and the functional role of these numerous enkephalin terminals remain to be established.     

Immunocytochemical localization of S-100 protein in the brain of adult rat

Summary The cellular and subcellular distribution of the nervous system-specific S-100 protein has been investigated in the brain of adult rat at the ultrastructural level by the pre-embedding unlabelled antibody PAP method. The protein is found in both fibrous and protoplasmic astrocytes and in the ependymal cells. The neurons, the oligodendrocytes as well as the microglial cells are lacking S-100. The labelled cells show a reaction product diffusely distributed in the cytoplasmic matrix and on specialized membranes, namely plasma membranes, outer mitochondrial membranes and membranes of the endoplasmic reticulum and Golgi apparatus. The astrocytic filaments and the axonemes of the ependymal cilia exhibit a strong immunoreactivity. The reaction product is also present in the nucleoplasm of the astrocytes and ependymal cells but it is absent from the nucleolus and nuclear envelope. This immunocytochemical data on tissue with satisfactory ultrastructural preservation, provides new information on the localization of the S-100 protein, and could contribute to the understanding of the biological role of the protein.     

Immunocytochemical localization of corticotropin-releasing factor (CRF) in the rat brain

The immunocytochemical localization of neurons containing the 41 amino acid peptide corticotropin-releasing factor (CRF) in the rat brain is described. The detection of CRF-like immunoreactivity in neurons was facilitated by colchicine pretreatment of the rats and by silver intensification of the diaminobenzidine end-product. The presence of immunoreactive CRF in perikarya, neuronal processes, and terminals in all major subdivisions of the rat brain is demonstrated. Aggregates of CRF-immunoreactive perikarya are found in the paraventricular, supraoptic, medial and periventricular preoptic, and premammillary nuclei of the hypothalamus, the bed nuclei of the stria terminalis and of the anterior commissure, the medial septal nucleus, the nucleus accumbens, the central amygdaloid nucleus, the olfactory bulb, the locus ceruleus, the parabrachial nucleus, the superior and inferior colliculus, and the medial vestibular nucleus. A few scattered perikarya with CRF-like immunoreactivity are present along the paraventriculo-infundibular pathway, in the anterior hypothalamus, the cerebral cortex, the hippocampus, and the periaqueductal gray of the mesencephalon and pons. Processes with CRF-like immunoreactivity are present in all of the above areas as well as in the cerebellum. The densest accumulation of CRF-immunoreactive terminals is seen in the external zone of the median eminence, with some immunoreactive CRF also present in the internal zone. The widespread but selective distribution of neurons containing CRF-like immunoreactivity supports the neuroendocrine role of this peptide and suggests that CRF, similarly to other neuropeptides, may also function as a neuromodulator throughout the brain.     

Immunocytochemical localization of serotonin in the brain and pituitary gland of the platyfish,Xiphophorus maculatus

Summary Immunoreactive serotonin (ir-5HT) containing cells were localized in the brain and pituitary gland of the platyfish by use of immunoperoxidase procedures. In the brain, ir-neurons were found lining the wall of the third ventricle and in its lateral and posterior recesses. More caudally, ir-perikarya were found in the valvular portion of the cerebellum and in the raphe region. Ir-5HT was also localized within the pineal gland in fish that had been sacrificed before 1:00 p.m. Within the pituitary gland, ir-5HT was localized in periodic acid Schiff-positive cells of the pars intermedia of all fish while, in only a few animals, less intense immunoreactivity was also present in gonadotrops of the caudal pars distalis.     

Immunocytochemical localization of Bombyx-PTTH-like molecules in neurosecretory cells of the brain of the migratory locust,Locusta migratoria

Summary Using a monoclonal antibody directed against a synthetic pentadecapeptide corresponding to the N-terminus of the prothoracicotropic hormone (PTTH) of Bombyx mori, we report the presence of immunoreactive molecules in a large number of median neurosecretory cells of the pars intercerebralis of the migratory locust, Locusta migratoria. These cells correspond to the A₁ cell type which we show to contain also neuroparsins, a family of predominant neurohormones of the migratory locust. In contrast, PTTH-like molecules are absent from A₂ cells of the pars intercerebralis which contain Locusta insulin-related peptide (LIRP). Developmental studies show the presence of PTTH-related substances in neurosecretory cells of Locusta migratoria from late embryogenesis to adult development, including ageing vitellogenic female adults.     

Immunocytochemical localization of aromatase-containing neurons in the rat brain during pre- and postnatal development

the present immunohistochemical study demonstrates the ontogenetic appearance of aromatase-immunoreactive neurons in several discrete regions of the hypothalamus and limbic system in the rat brain, using a purified antibody against human placental aromatase cytochrome P450. Immunoreactive cells were first detected in the preoptic area on the 13th day of embryonic life (E 13), and additionally in the bed nucleus of the stria terminalis on E 15. Labeled cells were also found in the medial amygdaloid nucleus and the ventromedial nucleus on E 16, and some were detected in the arcuate nucleus on E 19. As gestation progressed, the number and the immunoreactivity of these cells gradually increased and peaked within definite periods of perinatal life and there-after declined or disappeared. The immunoreactive cells were also found in the central amygdaloid nucleus and the lateral septal nucleus, and in the ventral pallidum, after the 14th day of postnatal life (P 14) and 30th day (P 30), respectively. The distribution of aromatase-immunoreactive neurons was similar between the sexes, while the immunoreactivity was higher in males than in females after late gestational days. No immunoreaction was detectable in other regions of the telencephalon or midbrain at any time periods studied. The aromatase-immunoreactive neurons in the specific regions may be involved in the sexual differentiation of the brain.