A cytochemical and ultrastructural study of the “S.I.F.” cells in cat sympathetic ganglia

Summary According to the hypothesis of Eccles and Libet, the small intensely fluorescent cells (S.I.F. cells) in the sympathetic ganglion would represent an essential element in the inhibition of the principal neuron. As a contribution to the study of this important problem, we have investigated serial sections in superior cervical (S.C.G.) and celiac (C.G.) ganglia of the cat, a species that has not been extensively studied up to now, both by fluorescence and electron microscopy. We have shown that the S.I.F. cells are three times fewer in the cat S.C.G. than in the rat S.C.G. There are five times more S.I.F. cells in the C.G. of the cat than in the S.C.G. of the same species. Moreover we have described two types of S.I.F. cells.Type I is composed of cells characterized by highly polymorphous large dense-cored vesicles. These cells lack processes and are grouped in clusters centered on fenestrated capillaries. They could be endocrine function cells. Type II is formed of isolated cells which exibit long processes and establish synaptic junctions with the dendrites of the principal neurons. In this case, the dense-cored vesicles are very regular and much smaller. These cells could be equivalent to interneurons. Type I very strongly predominates in the S.C.G. and C.G. of the cat where it represents more than 90% of the S.I.F. cell total observed by fluorescence microscopy. A priori such a quantitative and qualitative heterogeneity hardly consistent with Eccles and Libet's hypothesis based on the existence of dopaminergic interneurons only, allows the question to be raised as to the functional significance of the S.I.F. cells in ganglion physiology. The notion of modulation of ganglionic transmission does not seem to be quiered by these new data but could be founded on different forms of action embodied in the broader conception of the neuromodulation phenomenon.     

Cytochemical localization of 5′-nucleotidase in the enteric ganglia and in smooth muscle cells of the guinea-pig

Brain Cell Biology - Cytochemical techniques were used to study the localization of 5′-nucleotidase in the enteric ganglia and in smooth muscle cells of the guinea-pig ileum, iris and vas...     

Rana temporaria metamorph production and population dynamics in the field – Effects of tadpole density,predation and pond drying

This study investigates factors of importance for tadpoles survival and metamorph production in the common frog Rana temporaria. It also assess the importance of this for the population dynamics of the species. Eighteen ponds were studied for up to 8 years. Data collected each year included: number of spawn clumps deposited, tadpole number and metamorph number. The permanency of the ponds was also recorded each year. Measures were taken of predator density. There was no suggestion of density dependence in the survival of tadpoles. In contrast, the number of spawn clumps deposited per pond area was highest for ponds with high survival. Density of predators (sticklebacks, newts and invertebrates) was negatively correlated to tadpole and metamorph survival. This was true both within (among years) and among ponds. Several of the study ponds dried completely before metamorphosis in some years. However, those ponds also were those with the smallest number of predators and in years with successful metamorphosis, these ponds produced more metamorphs than more permanent ponds. An analysis of the year to year dynamics showed that population size (number of deposited spawn clumps) was correlated to that in the previous year, suggesting a fairly high adult survival, but also on the number of metamorphs emerging two or three years before (corresponding to the age of sexual maturity of the species). It is concluded that the aquatic stage is not strongly limiting in these ponds but conservation efforts should be focused on the terrestrial habitat. Also, the study stresses the value of temporary ponds, despite the fact that recruitment often fails totally in these.     

Ultrastructure of sympathetic ganglion cells and granule-containing cells in the paracervical (Frankenhäuser) ganglion of the newborn rat

Summary A study was made of the ultrastructure of the paracervical (Frankenhäuser) ganglion of the newborn rat, using immersion fixation by glutaraldehyde (2.5%) followed by OsO₄ (1%), or KMnO₄ (3%) fixation. The cells containing dense—core vesicles were divided into three groups: (1) primitive sympathetic cells, (2) cells containing some dense-core vesicles 700–1100 Å in size and structurally resembling sympathetic neurons, called principal neurons, and (3) cells containing many dense-core vesicles with a larger, darker dense core, 800–2000 Å in diameter, called granule-containing cells. Using glutaraldehyde-osmium fixation, the principal neurons were further divided into dark and light cells on the basis of electron opacity of the cytoplasmic matrix. The granule-containing cells were believed to correspond to the small, intensely fluorescent cells (SIF-cells) previously described using the formaldehyde-induced fluorescence technique. On the basis of the amount of granules, the granulecontaining cells were classified as mature or maturing SIF-cells and as more primitive SIF-cells, and developing sympathicoblasts. The development of synapses in autonomic ganglia was discussed.Grant: The Finnish Medical Foundation.     

Can 4-chloro-m-cresol be substituted for caffeine as an activator of calcium oscillation in bullfrog sympathetic ganglion cells?

4-Chloro-m-cresol (cresol) and caffeine have been shown to be potent activators of the Ca(2+) release mediated by the ryanodine-sensitive Ca(2+) release channel and therefore increase the cytosolic free calcium concentration in skeletal muscles. To distinguish the effects of cresol and caffeine in neurones, the responses of the intracellular ([Ca(2+)](i)) and intraluminal free calcium concentrations to cresol were investigated using bullfrog sympathetic ganglion cells and then compared with those to caffeine. Cresol generated a gradual rise (slow response) with and without a fast transient rise (fast response) in [Ca(2+)](i). A low extracellular Ca(2+) concentration abolished the slow response but not the fast response, thus indicating that the slow response was caused by a Ca(2+) influx across the cell membrane. The fast response was inhibited by ryanodine, thus confirming that the source may therefore be the Ca(2+) release through the ryanodine-sensitive calcium store. Unlike caffeine, the long-term application of cresol did not cause any calcium oscillation; neither did it cause a decrease in the basal calcium levels.     

Sensory cells of the “rod-” and “cone-type” in the pineal organ of Rana esculenta,as revealed by immunoreaction against opsin and by the presence of an oil (lipid) droplet

Summary The pineal organ of the frog, Rana esculenta, was studied by use of light- and electron-microscopic methods including immunoreaction against opsin. Most of the morphologically classified cone-type outer segments of the pineal photoreceptors reacted with antisera against opsin of the bovine retina that is dominated by rods. Some of the outer segments of pineal photoreceptor cells remained unstained in accord with the reference tissue, the frog retina, where generally the rods were opsin-positive and most of the cones opsin-negative.The opsin-negative outer segments of pineal photoreceptors were found in continuity with inner segments each containing a large oil (lipid) droplet. These oil droplets stained intensely with osmic acid, Sudan III, Sudan Black B or Scharlach R in cryostat sections, and were soluble in lipid solvents. In ultrathin sections of osmicated material, the oil droplets were homogeneous and of varying electron density. Approximately one tenth of the pineal photoreceptors contained oil droplets and at the same time possessed opsin-immunonegative outer segments.Since in the retina oil droplets and a negative immunoreaction against bovine opsin are characteristic of cones, we suggest that in the pineal organ they also mark conetype photoreceptors scattered among rod-type photo-receptors, the latter displaying a positive immunoreaction with the antisera used.Support from the Deutsche Forschungsgemeinschaft (Ok 1/25-3) is gratefully acknowledged     

Lepidoptera rhopalocera in the core of the transboundary biosphere territory “Altai:” Peculiarities of the fauna and population

Migration pathways of butterflies in Southeast Altai (the core of the Transboundary biosphere territory “Altai”) in the spring and summer seasons of 1990 and over the period of 1995–2006 were analyzed. The density of the population was assessed, and specific features of the butterfly distribution among four high-altitude belts (alpine tundra, subalpine, forest, and steppe) were revealed. Eighty-five habitats were examined and butterfly migration pathways were recorded for 359 hours of route surveys with a total length of 875 km. More than 16.5 thousand individuals belonging to 6 families, 66 genera and 107 species were recorded. The total species abundance of butterflies was found to be maximal in the forest belt; the highest population density was recorded in the alpine-tundra belt. The results obtained indicate that the basic differences among the butterfly populations are determined by the thermal conditions related to the altitude and, to a lesser degree, the degree of forestation. The distribution of most butterfly species over the transboundary territory is primarily determined by the recent environmental conditions, in particular the aridity of the tundra-steppe belt.     

An investigation of a sympathetic innervation of the vertebral artery in primates: is there a neurogenic substrate for vasoconstriction?

Vasoconstriction of the vertebral artery may be neurogenic in origin. Although the existence of a perivascular sympathetic plexus of the vertebral artery is not in doubt, no method used to date has conclusively demonstrated a direct sympathetic innervation of the vascular smooth muscle cells and, hence, vasomotor function. It was the aim of this study, therefore, to visualise and localise noradrenergic fibres in the wall of the vertebral artery. Intracranial vertebral artery specimens (10 vervet monkeys and 10 baboon vessels) were sectioned (40 mm serial sections) and treated with anti-tyrosine hydroxylase, anti-dopamine b-hydroxylase, and anti-chromogranin-A antibodies. Some evidence of catecholaminergic fibres in the tunica adventitia but not penetrating the external elastic lamina or tunica media of the vertebral artery wall was seen. These findings were confirmed by electron microscopy. It was concluded that although a perivascular sympathetic plexus exists, the vertebral artery of primates was not shown to have a direct sympathetic innervation and a neurogenic vasoconstrictor function is unlikely.     

Microsatellite variation and population structure in the “Refractario” cacao of Ecuador

Utilization of germplasm for crop improvement is often hampered by absence of information regarding origin, genetic identity and genealogical relationships of germplasm groups or populations. Molecular marker technology offers an efficient tool to verify or reconstruct passport data. Using a high-throughput genotyping system with 15 microsatellite loci, we fingerprinted 482 accessions in 48 putative half-sib families of Refractario cacao (a group of germplasm collected from nine farms in Ecuador). Based on the multilocus profiles, a Bayesian method for individual assignment was applied to verify membership in each half-sib family. Multivariate statistical analysis showed that the Refractario genetic profile was different from other groups tested, except for the “Nacional” cacao from the coastal valley of Ecuador. Hierarchical partitioning of genetic variance in the Refractario cacao showed that 76% of the variation was contributed by intra-family difference, whereas the inter-family and inter-farm difference accounted for 15 and 9% of total variance, respectively. All three sources of variation were highly significant (P < 0.01). Cluster and Principal Coordinates Analyses revealed a population sub-structure in Refractario, which was also highly heterozygous, suggesting hybridization derived from Nacional cacao and multiple other parental varieties, which all shared a similar genetic background. The improved understanding of identities and structure in Refractario cacao will contribute to more efficient conservation and use of this germplasm group in cacao breeding.     

Anatomy of the neurosecretory cells in the cerebral and subesophageal ganglia of the female European corn borer moth,Ostrinia nubilalis (Hübner) (Lepidoptera: Pyralidae)

The anatomy of the neurosecretory cells in the brain-subesophageal ganglion complex of female European corn borer moth Ostrinia nubilalis (Lepidoptera: Pyralidae) was studied using histological and cobalt backfilling techniques. Histological staining revealed the presence of 2 median and one lateral neurosecretory cell groups in the brain. These brain neurosecretory cells are made up of mainly type A cells with a few type B cells in the median group. Three type C neurosecretory cell clusters occupy the apparent mandibular, maxillary, and labial neuromeres at the ventral median aspect of the subesophageal ganglion. Axonal pathways of the neurosecretory cell groups were delineated by retrograde cobalt filling from the corpora cardiaca. Fibers of the 3 brain neurosecretory cell groups merged to form a distinct axonal tract that exits the brain via the fused nervi corporis cardiaci-1 + 2. Cobalt backfilling from the corpora cardiaca filled 4 groups of cell bodies in the subesophageal ganglion. The presence in the subesophageal ganglion of extensive dendritic arborizations derived from the brain suggests interactions between neurosecretory cell groups in the 2 head ganglia.     

The innervation of the hepatic portal vein in the rabbit: Ultrastructural evidence against “purinergic” neurotransmission

Summary The relative density of adrenergic and non-adrenergic nerves in the hepatic portal vein of the rabbit has been determined ultrastructurally. Adrenergic nerves were visualised with the modified chromaffin procedure of Tranzer and Richards (1976). Nearly equal numbers of adrenergic and non-adrenergic nerve profiles were found, indicating a much greater density of innervation by non-adrenergic nerves than that described by Burnstock et al. (1979) using light microscopic histochemical methods. These results imply that part of the argument used by Burnstock et al. (1979) to support purinergic transmission in rabbit portal vein is probably invalid.     

α-ENaC in bullfrog embryo: expression in cement gland, gills and skin

The epithelial sodium channel (ENaC) is involved in Na⁺ responses such as Na⁺ absorption and salt taste. The alpha ENaC subunit (α-ENaC) is expressed in the skin of both the adult and larval (tadpole) bullfrog. α-ENaC expression in the developing bullfrog embryo has not been previously investigated. In this study, the expression of α-ENaC at various stages (Sts.) of bullfrog embryonic development is assessed by western blot and immunofluorescence analysis. Bullfrog α-ENaC (α-fENaC) protein was detected by western blot in embryos at Sts. (Gosner/Shumway) 19, 21 and 25. Immunofluorescence studies indicate that α-fENaC was localized to the embryonic cement glands at St. 18 (muscular response), St. 19 (heart beat) and St. 21 (mouth open and/or cornea transparent), to the external gills at St. 21 and to the outermost cell-layer of the skin at St. 25 (operculum complete). The function(s) of ENaC in these embryonic structures remain to be elucidated.     

Contrasting effects of ACTH and cyanoketone on Δ5-3β-hydroxysteroid dehydrogenase activity in interrenal glands of tadpoles of Rana catesbeiana in vitro

Summary The present communication describes an investigation of stimulation and inhibition of ⁵-3-hydroxysteroid dehydrogenase in interrenal glands of tadpoles of Rana catesbeiana. Frozen sections of interrenal glands, together with kidneys, were prepared histochemically for assay of ⁵-3-HSD activity. Concentrations of 0.01, 0.1, 1, and 10 IU/ml of ACTH or of 0.01, 0.1, 1, and 10 g/ml of cyanoketone were added to the incubation media. The reaction products of the histochemically prepared slides, in terms of absorbance, were scanned at a defined area with a computerized microscope spectrophotometer. The results indicate that ACTH causes a significant dose-response stimulation of ⁵-3-HSD activity in tadpole interrenals; cyanoketone, on the other hand, causes significant dose-dependent inhibition.     

Silvering and gill “mitochondria-rich” cells in the eel,Anguilla anguilla

Gills of typical yellow and silver ells, Anguilla anguilla L., were examined by light and electron microscopy. In both eel types, mitochondria-rich cells were located in the epithelium covering the primary lamellae and consisted ofchloride cells and accessory cells. As compared to yellow eels, the primary gill epithelium of silver eels was thicker and contained larger and more numerous chloride cells with enlarged mitochondria. The accessory cells also increased in number but did not show significant modifications in their size or ultrastructural features. These observations indicate that, as far as mitochondria-rich cells are concerned, the silvering process in eels would be equivalent to smoltification in salmonids. It corresponds to a preparation for seawater life and is probably controlled by hormonal factors.     

“Intracisternal microtubules” in neurons of dog ganglia

Summary The changes which occur in the ultrastructure of the mesothoracic retractor unguis muscle of the cockroach Periplaneta americana as a result of disuse are described. Breakdown of myofibrils, sarcoplasmic reticulum, and mitochondria are all marked, this degeneration only being apparent 9 weeks after the operation.     

Observations on the relationship between “chloride-type” and “pseudobranch-type” cells in the gills of a fish,Oligocottus maculosus

Summary Chloride cells in gill epithelium of Oligocottus maintained in sea water have a much branched system of agranular cytoplasmic tubules, numerous mitochondria, and a prominent apical crypt. The mitochondria are randomly dispersed and do not show preferential orientation with respect to the tubules.After brief exposure of fish to sea water diluted 1/100 with glass distilled water, the mitochondria and tubules become rearranged into parallel arrays and the apical crypts disappear. The appearance of these cells is similar to that of pseudobranch cells in Fundulus heteroclitus (Copeland and Dalton, 1959).These data suggest that chloride cells and pseudobranch cells represent different adaptive forms of a single cell type and that transformation from the chloride cell configuration to that of pseudobranch cells can be induced by osmotic stress.This work was performed in 1963 when the author was enrolled in the Fine Structure Training program of the Department of Biological Structure, University of Washington. Financial aid was provided by the United States Public Health Service (Grant No. 5T1 GM-136).     

Identification and characterization of small cells in the adult pancreas: potential progenitor cells?

In this report we describe the identification of a novel cell type in human and canine pancreas using tissue culture techniques. These cells, representing less than 1% of total islet cells, are of a small size (7-10 microm) and highly quiescent. They display a fairly immature morphology, which is characterized by a weakly developed protein synthesis machinery, a few mitochondria and a small number of neuroendocrine granules. These cells, which we have termed "small cells," are usually organized into small clusters, which can be identified within the islets of predominantly small size. They can also be collected as separate structures from preparations of freshly isolated islets. Immunohistochemically, small cells are positive for PDX-1, synaptophysin, insulin, glucagon, somatostatin, pancreatic polypeptide, alpha-fetaprotein and Bcl-2 and negative for cytokeratin 19 and nestin. Insulin secretion studies demonstrated that these cells secrete insulin in a glucose-responsive fashion, although do not respond to secretagogues such as IBMX and arginine as do mature beta cells. Although this study does not provide evidence of the proliferative and differentiation potential of small cells, their immature morphology, along with a small size and quiescence, let us hypothesize that these cells may serve as progenitors contributing to the islet growth.