Turnover of asymmetric unit membranes in the transitional epithelial superficial cells of the rat urinary bladder.

Asymmetric thick unit membranes were observed on the luminal surface, fusiform vesicles, and multivesicular bodies of superficial cells of rat transitional epithelium. When HRP-labeled Ricinus communis lectin (RCA-I) was injected into the rat urinary bladder, RCA-I was deposited along the luminal cell membrane and in some multivesicular bodies, but not in the fusiform vesicles either before or after contraction. When the bladder was sliced by Vibratome and stained with HRP-labeled RCA-I after fixation, RCA-I was observed in many cell organelles, including fusiform vesicles and multivesicular bodies as well as the luminal surface. When small pieces of tissue were stained en-bloc with HRP-labeled RCA-I, RCA-I was found along the luminal cell surface but not in the fusiform vesicles nor the multivesicular bodies. When HRP alone was injected into the bladder, HRP was observed in some multivesicular bodies after contraction but not in the fusiform vesicles. Various lysosomes were observed by electron microscopy. Some were wrapping multivesicular bodies in ringlike fashion, and some contained asymmetric unit membranes. These findings suggest that the asymmetric unit membranes are carried to the luminal cell membrane via the fusiform vesicles and that old luminal cell membranes are removed via the multivesicular bodies to be degraded by lysosomes.

Development of human cerebellar nuclei. Morphometric study.

The morphometric development of the human cerebellar nuclei was examined in 9 fetuses (16-40 weeks of gestation; WG), an infant (2 months old) and 2 adults (16 and 63 years old). With the morphological observation of serial sections of the brain containing the cerebellar nuclei, the authors measured sections to get several morphometric parameters: the volume of nuclear column and number, packing density and cell body area of neurons. Each nucleus (dentate, emboliform, globose and fastigial nucleus) was recognized even at 16 WG. Nerve cells containing Nissl bodies were observed in all nuclei after 23 WG. Degenerative changes were detected in some neurons for every nucleus at 21 and 23 WG. Three stages were observed in the developmental course of nuclear volume and neuronal packing density: the primary or undifferentiated stage at 16 WG, the secondary stage with variability at 21-32 WG and the tertiary stage with monotonous increase (nuclear volume) or gradual decrease (neuronal packing density) after 35 WG. No significant correlation between neuronal number and gestational age was noticed for every nucleus. The analysis of cell body area (neuronal size) demonstrated that the dentate neurons developed after the intermediate or fastigial neurons. It is concluded that there is a critical period between slightly before 20 WG and slightly after 30 WG, matched with the secondary stage in the development of the cerebellar nuclei.

Modification of diurnal changes in the ultrastructure of synaptic ribbons of the turtle.

Synaptic ribbons of rod cells of the red-eared turtle under light-dark (LD) cycle, continuous dark (DD) and continuous light (LL) were examined by electron microscopy. In the LD cycle, several stick-shaped (plate-shaped, three-dimensionally) ribbons formed multilayered ribbon complexes at 1000 hr, and they turned into aggregates of rounded ribbons at 1900 hr (1 hr after light-off). In DD, stick-shaped ribbons were situated singly near presynaptic membranes. Large ribbon complexes consisting of 7-8 parallel layers, as shown in the LD cycle, were not observed. Aggregates of rounded ribbons, or club-shaped ones could not be observed. In LL, aggregates of irregular ribbon fragments were noted. None of the ribbons in this regimen showed sharp contours. Cyclic LD illumination is necessary to form multilayered ribbon complexes and rounded ribbons. Internal circadian rhythm contributes little part to the diurnal changes in the structure of the synaptic ribbons.

Derivation and termination of fusiform vesicles in the transitional epithelium of the rat urinary bladder.

Ultrastructural behavior of fusiform vesicles with an asymmetrical unit membrane in the rat transitional epithelium was investigated by in situ injection of gold colloidal particles and gold-labeled Ricinus communis lectin, and by section staining with the lectin. These experiments suggest that the fusiform vesicles are not formed from the luminal cell membrane by contraction of the urinary bladder, and that old luminal cell membranes are removed via multivesicular bodies.

Ultrastructural abnormalities in the skeletal muscle of children with chronic cholestasis and a long-term vitamin E replacement.

We investigated the effects of long-term oral and intramuscular vitamin E repletion in children with chronic cholestasis. Clinical improvement or suppression of neuromuscular involvement after adequate vitamin E repletion was demonstrated. Light and electron microscopic abnormalities of the skeletal muscle, however, did not completely disappeared despite the correction of the biochemical abnormalities for more than 3 years. The muscle fibers showed less variety of pathologic features than before vitamin E repletion. Inclusions observed in the skeletal muscle fibers before vitamin E treatment were still observed in subsarcolemmal cytoplasm and the perinuclear regions. They were more irregularly curved and consisted of various substances. Similar inclusions were also observed in Schwann cells, perineural cells, fibroblasts, pericytes, endothelial cells and smooth muscle cells of intramuscular vessels. Although the external lamina was not disrupted, separation of the external lamina from the plasma membrane and multilayered external lamina were often observed. The nerves among muscle fibers still showed degenerative features. Morphological changes of the skeletal muscle during vitamin E therapy have not so far been reported in cases of chronic cholestasis. We discuss the relationship of these findings to vitamin E replacement in children with chronic cholestasis.

Serotonergic efferent nerve fibers in the retinal plexiform layer of the abalone.

Serotonin (5-HT)-immunohistochemistries at light- and electron-microscopic levels, using rabbit anti-5-HT serum (#1234), were applied to the whole head and only to the eye of the abalone, respectively. Peroxidase-antiperoxidase and fluorescein isothiocyanate methods were used for the light-microscopic immunohistochemistries. Many immunoreactive nerve fibers were demonstrated in the outer zone of the retinal plexiform layer, small optic nerve fiber bundles, the optic nerve trunk and the cerebral ganglion. Immunoreactive somata were observed only in the cerebral ganglion. Accordingly the immunoreactive fibers in the retinal plexiform layer are considered to be efferent. Cored vesicles in the retinal plexiform layer demonstrated by both conventional chemofixation and a rapid-freeze-substitution method showed strong immunoreactivities localized within their limiting membrane. The same fibers also contained small clear vesicles. They are considered to be different from larger clear vesicles in non-immunoreactive fibers reportedly containing acetylcholine. The function of the efferent fibers remains to be elucidated.

Acyltransferase and acid hydrolase activities of the abalone photoreceptor cell.

In order to study the synthesis and degradation processes of the photoreceptor membranes in the abalone, Nordotis discus, the localization of acyltransferase and acid hydrolase activities, respectively, were determined at the electron-microscopic level. Acyltransferase activity was localized on the cytoplasmic sides of thick (greater than 10 nm) membranes of the following organelles: a few cisternae at the trans (or concave) side of Golgi apparatus, Golgi and probably related vesicles, short tubules, curved pentalaminar disks and limiting membranes of the phagosomal multivesicular bodies; all organelles were scattered in the peri- to supranuclear cytoplasm. The phospholipids, which are major components of the photoreceptor membrane, are considered to be synthesized by these membranes. Acid phosphatase activity was localized in the lumina of Golgi cisternae and vesicles, lysosomes, and smaller multivesicular and related bodies, but not in multilamellar bodies. The matrices of the larger multivesicular bodies and of the pigment granule complexes showed arylsulfatase activity. Vesiculated and autophagocytosed photoreceptor microvilli seemed to be degraded by acid hydrolases, forming multivesicular and related bodies. Supporting cells also showed acyltransferase and acid hydrolase activities.

Ultrastructural changes in skeletal muscle of a patient with familial intrahepatic cholestasis associated with vitamin E deficiency.

The skeletal muscle from a patient with familial intrahepatic cholestasis associated with vitamin E deficiency was studied by electron microscopy. The muscle fibers showed a variety of pathologic features including degenerative, necrotic and regenerative changes. Granule-like inclusions found in our biopsy specimens were similar in structure to those observed in vitamin E deficient animals. These inclusions were noted not only in the skeletal muscle fibers but also in endothelial cells of the intramuscular capillaries, Schwann cells and perineural cells. Disruption and disappearance of the plasma membrane, separation, disruption and pleats formation of the external lamina, and multilayered external laminae were observed in muscle fibers most frequently. The nerves among muscle fibers also showed degenerative features. These severely degenerative alterations of the muscle fiber have not so far been reported in vitamin E deficient patients. We discuss the process of cell damages caused by vitamin E deficiency.

Diurnal changes in synaptic ribbons of rod cells of the turtle.

Diurnal morphological changes in synaptic ribbons of the rod cells of the turtle were revealed by electron microscopy with serial ultrathin sections. Freshwater turtles (Pseudemys) were maintained under the light-dark cycle with lights on from 0600 to 1800 hr. Retinas around the posterior pole of the eyeball were fixed in 2.5% glutaraldehyde and 1% osmium tetroxide. In total, 124 rod cells and several hundred cone cells were observed. At 0000 (midnight), ribbons are situated close to and perpendicular to the presynaptic membrane. They show single stick-shaped profiles on thin section. From midnight toward noon, the stick-shaped ribbons grow into large multilayered ribbon complexes composed of several sticks arranged parallel to each other. Then, the ribbon complexes begin to disintegrate into irregular fragments from noon toward night. At 1900, aggregates of rounded or club-shaped ribbons are predominant.

Endothelial alterations of skeletal muscle capillaries in childhood myopathies.

Intramuscular blood capillaries in thirteen biopsied specimens obtained from eleven patients with various childhood myopathies were investigated ultrastructurally. The patients were classified into 4 different groups according to the percent distribution of various stages of endothelial alteration; normal endothelial type (E0 Type), slightly abnormal type (E1 Type), moderately abnormal type (E2 Type) and markedly abnormal type (E3 Type). Endothelial types in Duchenne dystrophy progressed from E0 to E1 and further to E2 Type, which kept pace with muscle degeneration. The endothelial types in Werdnig-Hoffmann disease and central core disease were E0 and those in multicore disease and Fukuyama type congenital muscular dystrophy were E1. Two cases of mitochondrial myopathy showed E3 Type, which was characterized by markedly swollen endothelial cells and narrowed lumens. The endothelium in these two cases showed more marked changes than in other childhood myopathies.

Ultrastructural alterations of Paneth cells in infants associated with gastrointestinal symptoms.

Paneth cells containing abundant and large inclusion bodies found in patients with acrodermatitis enteropathica have been considered as a consistent and characteristic abnormality of this disease. We examined 34 children with gastrointestinal symptoms due to a wide variety of basic diseases and found similar abnormal Paneth cells in most of them, particularly in infants under the age of one year. This abnormality, according to our findings, is attributed to the secondary effect of zinc deficiency due to malnutrition.

Relationship between granule-containing cells and blood vessels in the rat autonomic ganglia.

Granule-containing (GC) cells and related blood vessels in the superior cervical ganglion and the pelvic plexus of the rat were examined by light and electron microscopy of serial thin sections. In the superior cervical ganglion, GC cells formed many clusters of more than 20 cells. These clusters were supplied with dense networks of fenestrated capillaries, while the ordinary ganglion cells had continuous capillaries distributed more sparsely. Several continuous capillaries diverged from the networks in the GC cell clusters to join with capillaries around the ganglion cells. In the pelvic plexus, continuous capillaries with well-developed pericytes were distributed similarly around the GC cells and the ganglion cells. Dense networks of fenestrated capillaries in the clusters were never seen. Based on these observations, the physiological significance of the GC cells was discussed.

Differential effects of medial, central and basolateral amygdaloid lesions on four models of experimentally-induced aggression in rats.

To clarify whether various nuclei of the amygdaloid complex play different roles in aggressive behavior including muricide, 4 types of aggression were experimentally induced in rats. These include olfactory bulbectomy (OB rats), midbrain raphe lesions (Raphe rats), administration of delta 9-tetrahydrocannabinol (THC rats) and long-term isolation (Iso rats). Rats which exhibited muricide following these treatments were subjected to bilateral lesions of either the medial (AME), central (ACE) or basolateral (ABL) amygdaloid nuclei. Both muricide and hyperemotionality in the OB rat were markedly inhibited by AME lesions. Those of the Iso and THC rats were moderately inhibited. However, in the Raphe rat, aggressive behavior was not inhibited by AME lesions. Furthermore, ACE or ABL lesions caused no significant changes in all 4 models of aggression. These results suggest that the AME plays a facilitatory role in aggression of OB, Iso and THC rats, but aggression in Raphe rat is independent of amygdaloid activity.

A simple method for whole-cell preparation in electron microscopy.

A simple method for whole-cell preparation without using gold or platinum grids as substrata for culture is described. Cells were cultured on formvar film over round pores, each 3 mm in diameter, of Thermonox coverslips. The cells on the formvar coated coverslip are fixed, stained, dehydrated in situ, and introduced into a critical point drying apparatus. A small quantity of 0.2% mesh-cement is applied to slot grids, and they are laid onto the formvar film over the pores of the coverslip. After the grids are removed from the plastic substratum, they are ready for observation under the electron microscope.

Ultrastructural studies on congenital generalized fibromatosis regressed spontaneously.

Cellular components of a subcutaneous nodule from a male infant with congenital generalized fibromatosis consisted of myofibroblasts, fibroblasts, and their various intermediate types. The myofibroblasts were characterized by the presence of bundles of microfilaments and dense bodies similar to those in smooth muscle cells. Membrane-bound vesicles containing dense material were abundant in the extracellular space. They were probably lysosomes released by disintegration of cellular components. A possible role of the myofibroblasts and extracellular lysosomes in the regression of nodular lesions was discussed.

Freeze fracture study on three types of synapses in the Xenopus spinal cord.

Three types of synapses (S-, F- and C-types) were identified in the thin-sectioned Xenopus spinal cord and their structure was analyzed with the freeze-fracturing technique. All three types of synapses showed similar specializations of the presynaptic membrane. This finding suggests that the three types of synapses may release their transmitters by a similar mechanism. By contrast, the three types of synapses revealed different specializations of the postsynaptic membrane. The E-face of the S-type postsynaptic membrane was characterized by a dense aggregate of large intramembrane particles, 12 to 15 nm in diameter. An aggregate of small particles, 8 to 9 nm, was evident on the E-face of the postsynaptic membrane of the F-type synapse. In the C-type synapse, there was a striking aggregate of intramembrane particles, 10 to 14 nm in diameter, on the P-face of the postsynaptic membrane. These characteristic features in the distribution of particles in the three types of postsynaptic membranes may reflect differences in the type of transmitter released or transmitter action on the postsynaptic neuron. The overall size of the area of aggregated particles on the P-face of the C-type postsynaptic membrane was coextensive with the underlying subsurface cistern (SSC) which showed partial occlusion of the lumen. This fact supports the view that the SSC is closely related to the C-type synaptic action which might be distinct from the other synaptic types.

Diurnal changes in the fine structure of photoreceptors in an abalone, Nordotis discus.

In order to examined a diurnal (24 h) rhythm in the fine structure of photoreceptors of abalone, Nordotis discus, the animals were kept on a daily light-dark cycle (light period: 6:00 a.m.--6:00 p.m., 2--5 lx) for 10 days; subsequently their eyes were fixed at nine specified times of the day. At 5:30 a.m. the photoreceptors consisted of numerous terminal microvilli growing on the surface of a dome-shaped apical cytoplasmic process (AP). Illumination induced breakdown of the photoreceptive microvilli into many vesicles. These vesicles were densely aggregated on almost the entire surface of enlarged APs at 7:00 a.m. After 1 h they markedly decreased in number. The cell at 10:00 a.m. possessed sparse, short and irregularly-contoured microvilli. The microvilli gradually increased in number from the middle through the later half of the light period. Only within 1 h after deprivation of light shorter but characteristic terminal microvilli were observed. By midnight they became similar in fine structure to those at 5:30 a.m.

Autonomic innervation of the muscles in the wall of the bladder and proximal urethra of male rats.

The muscular coat of the body of the rat bladder is innervated almost exclusively by cholinergic endings:adrenergic endings are rare. In the inner longitudinal muscle layer of the proximal urethra, 53% of 310 autonomic nerve endings observed in close relation to the smooth muscle cells were adrenergic and the remaining 47% cholinergic. The middle circular muscle layer of the proximal urethra was innervated predominantly by adrenergic endings: in this layer 86% of the total of 335 endings examined wre regarded as adrenergic. A similar predominantly adrenergic innervation was noted in the outer longitudinal layer of the proximal urethra. A number of striated muscle fibres arose from the outermost striated muscle layer of the proximal urethra and intruded deeply into the outer and middle smooth muscle layers. These intruding striated muscle fibres also received direct autonomic (mostly adrenergic) innervation. The significance of these findings in relation to the physiology of the lower urinary tracts is discussed.

The occurrence of electron dense intercellular materials and gap junctions in the human intestinal epithelium.

Electron microscopy of biopsy specimens from the small intestines in infants and children with various gastrointestinal disturbances revealed that the intercellular space of the mucosal epithelium was often filled with electron dense materials. Here apposed cell membranes were arranged almost parallel with each other. The dense materials also filled the space just subjacent to the epithelium to form a thick, dense layer in which the basal lamina was completely buried. The gap junctions were especially well developed in the epithelia filled with these materials. Long-period fasting of patients seemed to facilitate the occurrence of the dense materials and the formation of gap junctions.