Non-isotopic in situ hybridization at the ultrastructural level

Non-isotopic in situ hybridization techniques are becoming increasingly widely used at the ultrastructural level, permitting rapid localization of nucleic acid targets with a high degree of resolution. Technical considerations dictate that the great specificity of the method cannot be matched by a similar degree of sensitivity; the value of non-isotopic ultrastructural in situ hybridization lies in its unique ability to localize nucleic acid targets in relation to submicroscopic cellular structures. This article presents an overview of non-isotopic ultrastructural hybridization methods and applications.     

Distribution and synaptic organization of dopaminergic axons in the lateral geniculate nucleus of the rat

In the present study, immunocytochemistry with an antiserum against dopamine (DA) revealed hitherto unknown terminal fields of DA axons in the lateral geniculate nucleus (LGN) of the rat. The innervation of all subdivisions of the LGN is achieved by a common set of afferent fibers that branch to form terminal fields of uneven density. The ventral lateral geniculate nucleus (LGv) receives slightly more DA axons than the dorsal lateral geniculate nucleus (LGd), whereas within the latter, DA afferents innervate the lateral part of the nucleus slightly more densely. Labeled axon terminals and varicosities, examined in single and serial ultrathin sections, were found in the extraglomerular neuropil in the LGd and in the neuropil of the LGv characterized by relatively simple synaptic relationships. They formed predominantly asymmetrical synaptic contacts with dendritic profiles. Occasionally, the postsynaptic elements were found to be presynaptic dendrites of presumptive interneurons. Some of the possible roles of this newly demonstrated DA afferent system in the physiology of the LGN and in the pathophysiology of diseases associated with impairment of dopaminergic activity are discussed.     

Prenatal development of the sheep pineal gland: An ultrastructural study

Abstract: The ultrastructure of the pineal gland of 32 sheep embryos was studied from day 54 of development through birth. Embryos were arranged in four age-groups, defined in terms of the most relevant histological features: group 1 (54 to 67 days of prenatal development), group 2 (71 to 92 days), group 3 (98 to 113 days), and group 4 (118 to 150 days). A primary cell type, designated the pinealoblast, was observed from 54 days until birth; ultrastructurally, this cell was found to contain all the organelles required for hormone synthesis. A second cell population, classified as interstitial cells by virtue of their location among pinealoblasts, appeared at 78 days gestation and persisted until birth. Interstitial cells were scarce and exhibited tropism for the perivascular space. From 118 days gestation until birth, a third cell type, termed the pigmented cell, was visible. Pigmented cells, whose ultrastructural characteristics differed from those of pinealoblasts, contained a large number of pigment granules of varying size and shape. The pineal gland of developing sheep embryos showed considerable innervation and abundant vascularization; this, together with certain ultrastructural characteristics, suggests that the gland has a secretory function in uterine life.     

Immunoglobulin Synthesis in Hairy Cell Leukaemia

3 cases of hairy cell leukaemia were studied with ultrastructural immunocytochemical methods using an anti human Ig HRPO-Fab fragment. Ig were detected on the cell surface, in the perinuclear cisterna and endoplasmic reticulum of hairy cells. Evidence of Ig in these sites demonstrates a B-lymphoid differentiation of the leukaemic cells.     

Extra-synaptic localization of α-bungarotoxin receptors in cultured chick ciliary ganglion neurons

Surface [¹²⁵I]α-bungarotoxin receptors were localized on dissociated cultures of embryonic chick ciliary ganglion neurons by electron microscopic autoradiography. Grain density was 4–5-fold higher on cell body membrane versus neuritic process membrane, and there was no apparent concentration of grains at morphologically identifieble inter-neuronal synapses.     

Interest of simultaneous ultrastructural characterization of morphology, cytochemistry and immune phenotype in a case of putative hybrid acute leukaemia

A case of acute leukaemia is reported in which blast cells expressed some B-related antigens (namely the CALLA antigen) and no peroxidase activity at the optical level; however, some mature granular cells contained Auer rods. Simultaneous characterization of ultrastructural morphology, cytochemistry and immune phenotype was performed. There was an apparent mutual exclusion in the expression of myeloperoxidase activity and the CALLA antigen, and a heterogeneity in the CALLA expression among the blastic population. These results disagree with the hypothesis of a true biphenotypic leukaemia and demonstrate a complete heterogeneity between the lymphoblastoid cells and the myeloid ones. The interest of such a simple combined method in a case of putative hybrid acute leukaemia is emphasized.     

Light and Electron Microscopic Study of Dalén-Fuchs Nodules in Sympathetic Ophthalmia

A light and electron microscopic study was undertaken in an effort to establish the origin of the “epithelioid” cells in Dalen-Fuchs nodules from an eye enucleated because of sympathetic ophthalmia. The nodules were visible as minute (130–160 µm), round, grayish-white mounds elevating the retinal pigment epithelium. Bruch's membrane appeared intact in all the sections examined. By electron microscopy the “epithelioid” cells had round to oval nuclei with abundant, relatively lucent cytoplasm containing parallel profiles of rough-surfaced endoplasmic reticulum, prominent Golgi lamellae, clusters of polyribosomes, and scattered mitochondria. Many interdigitations of the plasma membranes, some of which exhibited fascia adherens type attachments, were observed. Some cells within the nodules showed large membranebound phagosomes containing laminated structures. Other “epithelioid” cells displayed moderately electron dense membrane-bound granules that appeared to be early precursors of lipofuscin granules. Examination of the nodules under ultraviolet light showed myriad autofluorescent yellowish-orange dots consistent with lipofuscin. Additionally, a montage of electron micrographs from the edge of the nodule, coupled with the above findings, provided support to the concept proposed by Ishikawa and Ikui (1972) that the “epithelioid” cells in Dalén-Fuchs nodules represent transformed retinal pigment epithelial cells forming a cage-like framework within the nodule. This study also emphasizes the remarkable capabilities for differentiation by the retinal pigment epithelial cells.     

Identification des terminaisons vestibulaires dans les noyaux oculomoteurs communs chez le chat par radioautographie en microscopie electronique

The projection of vestibular pathways to the oculomotor nuclei was investigated by electron microscopic radioautography. Unilateral injection of tritiated amino acids into the rostral vestibular complex was used in order to characterize the location and to identify the different types of labeled synaptic terminals involved in these pathways.In the normal oculomotor nuclei, 4 types of synaptic boutons were identified. Following the labeling of the vestibular synapses, in the ipsilateral oculomotor nucleus, types I and II boutons are the most prominent group and make up 75% of the total number of this labeled synaptic population. Large boutons with small diameter synaptic vesicles, they are distributed on the cellular soma and the large dendrites of the oculomotor neurons. In contrast, in the contralateral oculomotor nucleus, type III boutons which are smaller and have larger diameter synaptic vesicles were predominant; they are prevalent on the distal part of the dendritic tree.From the results obtained, a relationship between the present anatomical findings and previously published physiological studies is established. The following conclusion is suggested: the inhibitory vestibular inputs probably terminate on the oculomotor neurons by these large types I and II boutons and the excitatory vestibular inputs by the smaller type III boutons. Also discussed is the complexity of the pattern of afferentation and the functional arrangement of the oculomotor nuclei.     

Les différentes formes des inclusions nucléaires et cytoplasmiques de la leucoencéphalite sclérosante subaiguë en microscopie ultrastructurale

Summary Enzyme histochemical as well as electron microscopic studies were made in a typical case of SSLE. From the enzyme histochemical point of view the lesions were not specific, and the observed increase of oxydative enzymes is identical with that found in every reactive macro- and microgliosis. No intranuclear inclusions of microtubuli with a diameter of 170–230 Å resembling certain mixoviruses could be demonstrated by electron microscopy. By light microscopy, nuclear inclusions of the Cowdry type could not be demonstrated neither; however, they are not always found in SSLE. The microtubuli inclusions, observed for the first time by one of us, seem to be the most specific element in SSLE. On the other hand, the intracytoplasmatical particles with a virus-like aspect which have been described byGonatas in SSLE and by ourselves in periaxial diffuse encephalitis of the Schilder type were also found in the present case. Our impression is that these particles are nonspecific and great caution has to take place in classifying them as viral elements. They might be secretory granules arising in the glia in certain pathological conditions, or perhaps phagocyted particles, possibly stemming from myelin.

Ce travail a été réalisé en partie grâce aux subsides de la Fondation de Recherche Scientifique Nationale de Belgique et de la Fondation Engels Born pour l'Histochimie.