The electron microscopic and classic Golgi apparatus

The relationship of the membrane structure, designated in electron microscopy as the Golgi apparatus, to the classic Golgi apparatus in the light microscope were studied withFagopyrum. Comparison of these structures in plant cells with the same or similar structures in animal cells led to the following conclusions: there exist two groups of formations, impregnable with osmium or silver, considered as the classic Golgi apparatus. The first group contains the active membrane structures. These are the dictyosomes and the anastomosing form of the electron microscopic Golgi apparatus. To this group belongs also the endoplasmatic reticulum, which in plant cells forms dense vacuoles, having the appearance of the classic Golgi apparatus, and in animal cells occasionally has a similar arrangement as the anastomosing form of the Golgi apparatus. The second group comprises formation containing reserve and secretion material, i.e. predominantly products of the activity of the electron microscopic Golgi apparatus and of the endoplasmic reticulum (matter of the dense vacuoles, lipochondria, secretory granula etc.). In the plant cells, especially ofFygopyrum, the dictyosomes contained in the structures of the first group are separated from the formations of a reserve character in the second group, formed in the lumen of the endoplasmic reticulum (dense vacuoles). The identity of the dictyosomes with the osmiophilic platelets, considered by some authors in the light microscope as the classic Golgi apparatus, has not been proved up to present, because of the one-sidedness of the methods used nowadays. WithFagopyrum no foundation has been observed for the assumed formation of net-form structures by grouping of the dictyosomes. Structures similar to the net-form of the classic Golgi apparatus in the animal cell form only dense vacuoles. On the basis of the differentiation of both types of formations in the plant cell, the foundations were laid for the characterization of the classic Golgi apparatus in the animal cell. The net-form of the classic Golgi apparatus in the animal cell is obviously not artificial, but reflects the ultrastructural arrangement of the electron microscopic Golgi apparatus or of the endoplasmic reticulum. The problem of the suitability and specification of the name Golgi apparatus in the animal and plant cell was also discussed. In contrast to the opinion of some authors, it does not appear useful to remove the name golgi apparatus, designating the dictyosomes and the anastomosing forms of the smooth membranes.     

ELECTRON MICROSCOPY OF THE VOLVOCACEAE AND ASTREPHOMENACEAE

Lang, Norma J. (U. Texas, Austin.) Electron microscopy of the Volvocaceae and Astrephomenaceae. Amer. Jour. Bot. 50(3): 280-300. Illus. 1963.—Clonal cultures of Gonium sociale, G. pectorale, Pandorina morum, Eudorina elegans, Eudorina sp., Volvulina steinii, V. pringsheimii, Platydorina caudata, Pleodorina illinoisensis, P. californica, Volvox aureus, V. tertius, V. globator, V. barberi, and Astrephomene gubernaculifera representing the Volvocaceae and Astrephomenaceae in the Volvocales were examined with the electron microscope and their ultrastructure compared. The ultrastructure of the various organelles is basically similar in the species studied and no increase in cellular complexity is found to accompany the evolutionary trends evidenced in the Volvocaceae. The ultrastructure of a colonial cell is basically that of Chlamydotnonas. A cytoplasmic membrane having a unit membrane structure encompasses a cell and is continuous with the double-membraned flagellar sheaths. The flagella contain the typical 9 + 2 fibril arrangement with the 2 axial fibrils terminating in a cylinder at the flagellar base and the 9 peripheral pairs continuing into the cytoplasm as a basal body. The organelles comprising the cytoplasm are: mitochondria with plate-like cristae; dictyosomes composed of stacks of agranular cisternae; small, rough or smooth-surfaced vesicles; an endoplasmic reticulum of granule-bearing and agranular tubules, lamellae and broad cisternae; vacuoles which are either contractile, contain fine granular and fibrillar material, or have dense contents probably representing polyphosphate; lipid bodies; and dense granules 100–150 A which have been called ribosomes. The finely granular nucleoplasm is surrounded by a porous, double-membraned nuclear envelope and contains a centric nucleolus composed of dense, spherical granules. The outer membrane of the nuclear envelope bears granules and may have granular extensions into the perinuclear cytoplasm. Each extension appears to encompass one or several dictyosomes and has been termed an “amplexus.” The amplexi are agranular on the surface contiguous to a dictyosome. A double-membraned chloroplast envelope is continuous around the single, cup-shaped chloroplast. The basic chloroplast units are discs closed at each end, occurring in stacks of varying number parallel to the envelope. The presumed proteinaceous matrix of the basal pyrenoid is penetrated by elongated, tubular elements which connect with the lamellar discs. Multiple rows of granules, associated with individual discs, form the anterior stigma within the chloroplast envelope. The colonial matrix is not a structureless, mucilaginous material uniting the cells in colonies, but it has rather a highly complex structure especially around the periphery of the colony and the flagellar channels. The apparent substitution of a fibrillar layer of the colonial matrix for the discrete compact cell wall, such as is found in Chlamydomonas, implies a greater degree of complexity in the evolution of these colonial genera than is generally assumed.     

CHLOROPLAST DEVELOPMENT AND ULTRESTRUCTURE IN THE FRESHWATER RED ALGA BATRACHOSPERMUM1

Chloroplast development and ultrastructure of the freshwater red alga Batrachospermum moniliforme are described. Chloroplasts develop from proplastids which have a double-membraned chloroplast envelope and a parallel double-membraned outer photo-synthetic lamella. Of these 2 double-membraned structures of the proplastid, only the outermost pho-tosynthetic lamella functions in production of further lamellae. The mature chloroplast consists of 2 or more concentric lamellae and a variable number of nonconcentric lamellae. These lamellae are not dense, uninterrupted sheets as described for other red algae, but are largely constructed of tubules, lying side by side, that form interrupted lamellar sheets. The possible physiological significance of lamellar interruptions in providing path-ways for movement of materials in the chloroplast stroma is discussed.     

STRUCTURE AND DEVELOPMENT OF THE CHLOROPLAST IN CHLAMYDOMONAS : I. THE NORMAL GREEN CELL

The cytoplasmic organization of a normal green strain of the alga Chlamydomonas reinhardi has been investigated with the electron microscope using thin sections of OsO₄ fixed material. The detailed organization of the chloroplast has been of special interest. The chloroplast, a cup-shaped organelle, surrounded by a double membrane, consists of: (1) discs about 1 micron in diameter, considered to represent the basic structural unit of the chloroplast, and each composed of a pair of membranes joined at their ends to form a flat closed vesicle; the discs are grouped into stacks resembling the grana of higher plants; (2) matrix material of low density in which the discs are embedded; (3) starch grains; (4) the pyrenoid, a non-lamellar region associated with starch synthesis, and containing tubules which connect with the lamellae; (5) the eyespot, a differentiated region containing two or three plates of hexagonally packed, carotenoid-containing granules, located between discs, and associated with phototaxis. In addition to the chloroplast, the cytoplasm contains various membranous and granular components, including mitochondria, endoplasmic reticulum, and dictyosomes, identified on the basis of morphological comparability with structures seen in animal cells. The nucleus, not investigated in detail in this study, contains a large, granular nucleolus and is surrounded by a nuclear envelope which is provided with pores and exhibits instances of continuity with the endoplasmic reticulum of the cytoplasm.     

AN ELECTRON MICROSCOPE STUDY OF THE CYTOLOGY OF THE PROTOZOAN EUPLOTES PATELLA

1. Structurally the "sensory bristles" in Euplotes patella are typical cilia, but no ciliary rootlets connect their bases. 2. The "neuromotor fibrils" are composed of filaments 21 mµ in diameter. At the point of junction of the filaments with the peripheral ciliary fibrils a granular structure 65 to 90 mµ in diameter is seen which has dense central and peripheral zones separated by a less dense layer. Information on the interconnection of organelles is expanded. 3. A system of subpellicular fibrils is described. The external fibrillar system described by others could not be found. 4. The motorium is shown to be a mass of intertwining rootlet filaments. 5. The micronucleus is shown to have a spongy, dense material in a less dense material, all of which is surrounded by a double-layered membrane. 6. The double-layered macronuclear membrane contains annuli whose outside diameter is 70 mµ; the macronuclear bodies are sometimes closely applied to the membrane. In the macronuclear reorganization bands, the solution plane is a fine network, while the reconstruction plane is devoid of structure at the level of resolution observed. 7. The mitochondria are composed of tubules, only occasionally oriented, usually embedded in a surrounding material of lower density. 8. Microbodies whose diameters are 250 to 350 mµ are frequently observed in close association with mitochondrial surfaces. 9. The food vacuoles, contractile vacuoles, and ciliary vacuoles are bounded by single-layered membranes. In the food vacuoles, the bacteria are surrounded by membranes individually or in small groups. 10. Cytoplasmic rods localized in the oral region, and cytoplasmic granules dispersed at random, are described. No typical ergastoplasm, endoplasmic reticulum, or Golgi material was observed.     

THE ULTRASTRUCTURE OF THE VITELLINE BODY IN THE OOCYTE OF THE SPIDER TEGENARIA PARIETINA

The vitelline body in the mature oocyte of the spider Tegenaria parietina is composed of 4 different zones. 1. The central zone contains granular areas, vesicles, and a few lamellae. 2. The lamellar zone consists of numerous concentric lamellae. These sheets, 45 A in thickness, are stacked in groups. The fine structure and the regular arrangement recall those of myelin sheets, retinal rods, and chloroplasts. Between the stacks of lamellae, finely granular masses and various vesicles are to be found. 3. The "zone of transition" consists of a finely granular substance accumulated in abundant masses. This substance is composed of very closely packed granules about 50 to 60 A in diameter. Very often, near the lamellae, the granules show alignment giving a gradual transition from grains to lamellae. 4. The vesicular zone contains ergastoplasm, dense particles, mitochondria, and Golgi material. It is suggested that the peculiar ultrastructure of these cytoplasmic components may be related to an intense metabolic activity.     

VACUOLE FORMATION IN THE ACTIVELY GROWING ROOT MERISTEM OF BARLEY (HORDEUM SATIVUM)

The subapical meristem of actively growing barley roots produces series of undifferentiated cells, some of which are devoid of vacuoles. At the beginning of their differentiation, the Golgi apparatus gives rise to vesicles and tubules which concentrate hydrolases, acid phosphatase being the typical representative of these enzymes. Some of these structures organize themselves as sequestration vacuoles. Then, the imprisoned fraction is destroyed by the process of autophagy after an alteration of the vacuolar internal membrane. These structures are identical to the “provacuolar apparatus” described by Marty in Euphorbia characias roots. Lytic processes which develop in autophagic vacuoles give rise to the first true meristematic vacuoles. Relations between dictyosomes, provacuoles and vacuoles, and their degree of exclusivity are discussed.     

Cytochemical localization of peroxidase in soybean suspension culture cells and protoplasts: Intracellular vacuole differentiation and presence of peroxidase in coated vesicles and multivesicular bodies

Summary The ultrastructural localization of peroxidase in soybean (Glycine max L.) suspension culture cells and protoplasts is reported. In cells peroxidase is found primarily in the cell wall and at the tonoplast. Protoplasts and cells contain a vacuolar system which is differentiated with respect to peroxidase content since some vacuoles are found which do not contain peroxidase reaction product. The Golgi dictyosomes, coated and smooth vesicles contain peroxidase. Some of the multivesicular bodies have the reaction product as well. The results are discussed in terms of the pathways of sorting of peroxidase between the cell wall and vacuoles of cultured cells.     

ULTRASTRUCTURE OF THE SHOOT APEX OF CHENOPODIUM ALBUM AND CERTAIN OTHER SEED PLANTS

The ultrastructure of cells of the vegetative shoot apices is described for Chenopodium album, Kalanchoë blossfeldiana and K. laxiflora, Bryophyllum daigremontianum, Nicotiana rustica, and N. tabacum (Maryland Mammoth), and Ginkgo biloba. A less intensive study was made of the last three listed. The structures and organelles usually associated with meristematic cells were observed: dictyosomes, plastids (in various stages of development), mitochondria, endoplasmic reticulum (ER), vacuoles, lipid droplets, and plasmalemma. In addition, spherosome-like structures were observed in all zones of the shoot apices. Also, multivesicular bodies were observed in C. album and B. daigremontianum. Ribosome density is greater in cells of the flank meristem. Proplastids, plastids with prolamellar bodies, or grana have a differential distribution in the apex, characteristic for a particular species. Confirmation could not be given to the concept that vacuoles arise as a series of local dilations in long extensions of the so called "smooth ER." The tonoplast and ER are distinguishable at the time of inception of a vacuole, although the tonoplast may arise from the ER. Rapid growth of a vacuole and/or fusion with other vacuoles may result in irregularly shaped prevacuoles. No vacuoles were observed to originate from cisternae of dictyosomes in the species studied.     

LYSOSOMES IN THE RAT SCIATIC NERVE FOLLOWING CRUSH

Peripheral nerves undergoing degeneration are favorable material for studying the types, origins, and functions of lysosomes. The following lysosomes are described: (a) Autophagic vacuoles in altered Schwann cells. Within these vacuoles the myelin and much of the axoplasm which it encloses in the normal nerve are degraded (Wallerian degeneration). The delimiting membranes of the vacuoles apparently form from myelin lamellae. Considered as possible sources of their acid phosphatase are Golgi vesicles (primary lysosomes), lysosomes of the dense body type, and the endoplasmic reticulum which lies close to the vacuoles. (b) Membranous bodies that accumulate focally in myelinated fibers in a zone extending 2 to 3 mm distal to the crush. These appear to arise from the endoplasmic reticulum in which demonstrable acid phosphatase activity increases markedly within 2 hours after the nerve is crushed. (c) Autophagic vacuoles in the axoplasm of fibers proximal to the crush. The breakdown of organelles within these vacuoles may have significance for the reorganization of the axoplasm preparatory to regeneration. (d) Phagocytic vacuoles of altered Schwann cells. As myelin degeneration begins, some axoplasm is exposed. This is apparently engulfed by the filopodia of the Schwann cells, and degraded within the phagocytic vacuoles thus formed. (e) Multivesicular bodies in the axoplasm of myelinated fibers. These are generally seen near the nodes of Ranvier.     

Electron microscopy of the skin of the teleost,Hippoglossoides elassodon

Summary The normal skin of the pleuronectid fish, Hippoglossoides elassodon, is described by light and electron microscopy. The epidermis consists of 5 to 9 layers of cells, the majority of which are squamous cells and the minority mucous cells. The squamous cells are characterized by numerous desmosomes and associated cytoplasmic filaments. The mucous cells accumulate mucous droplets in vacuoles of Golgi origin and are observed apparently in the process of releasing their content at the free surface. The dermis consists of alternating lamellae composed of typical collagen fibers. Pigment cells are of three types: melanophores, iridophores (guanophores), and lipophores.This work was supported by Public Health Service Research Grant CA-08158 from the National Cancer Institute.     

THE GOLGI APPARATUS IN NEURONS AND EPITHELIAL CELLS OF THE COMMON LIMPET PATELLA VULGATA

1. In view of widely diverse views held about the identity and structure of the Golgi apparatus in neurons of Mollusca, particularly gastropods, a study has been made on neurons of the common limpet, Patella vulgata, both by light and electron microscopy. A report is given also of observations made on epithelial cells of Patella by electron microscopy. 2. As revealed by Kolatchev's method, the Golgi apparatus in neurons consists basically of black filaments lying to one side of the nucleus. The filaments generally anastomose to form networks of various complexity. Rarely some cells contain only discrete filaments. Associated with some of the filaments is a weakly osmiophilic substance identified as archoplasm. Kolatchev's method also revealed spheroidal bodies (neutral red bodies, "lipochondria," etc.). 3. It has not been possible to demonstrate the Golgi apparatus using either iron-haematoxylin or Sudan black. 4. Examination of Kolatchev's preparations by electron microscopy has revealed that some of the Golgi filaments consist of chromophilic and chromophobic components. The chromophilic component consists of dense lamellae. 5. After fixation in buffered osmium tetroxide solution and examination by electron microscopy, it has been concluded that (a) the chromophilic component of the Golgi apparatus corresponds to a system of paired membranes (which usually enclose an inner dense substance), (b) the chromophobic component corresponds to a substance lying within small dilations of the paired membrane, and (c) the archoplasm corresponds to numerous small vesicles. 6. The paired membranes branch, anastomose, and can often be traced back to a common source. They are interpreted as lamelliform folds, and occasionally tubular processes, of essentially a single Golgi membrane. In cells containing a Golgi network it is suggested that the membrane extends through the whole of the apparatus in such a way that the substance it encloses may be regarded as being in a continuous phase. 7. Epithelial cells of Patella contain a juxtanuclear Golgi apparatus with an ultrastructure similar to that described for neurons.     

THE FINE STRUCTURE OF ACANTHAMOEBA CASTELLANII (NEFF STRAIN) : II. Encystment

Encysting cells of Acanthamoeba castellanii, Neff strain, have been examined with the electron microscope. The wall structure and cytoplasmic changes during encystment are described. The cyst wall is composed of two major layers: a laminar, fibrous exocyst with a variable amount of matrix material, and an endocyst of fine fibrils in a granular matrix. The two layers are normally separated by a space except where they form opercula in the center of ostioles (exits for excysting amebae). An additional amorphous layer is probably present between the wall and the protoplast in the mature cyst. Early in encystment the Golgi complex is enlarged and contains a densely staining material that appears to contribute to wall formation. Vacuoles containing cytoplasmic debris (autolysosomes) are present in encysting cells and the contents of some of the vacuoles are deposited in the developing cyst wall. Lamellate bodies develop in the mitochondria and appear in the cytoplasm. Several changes are associated with the mitochondrial intracristate granule. The nucleus releases small buds into the cytoplasm, and the nucleolus decreases to less than half its original volume. The cytoplasm increases in electron density and its volume is reduced by about 80%. The water expulsion vesicle is the only cellular compartment without dense content in the mature cyst. The volume fractions of lipid droplets, Golgi complex, mitochondria, digestive vacuoles, and autolysosomes have been determined at different stages of encystment by stereological analysis of electron micrographs. By chemical analyses, dry weight, protein, phospholipid, and glycogen are lower and neutral lipid is higher in the mature cyst than in the trophozoite.     

ON THE SITE OF SULFATION IN THE CHONDROCYTE

As observed autoradiographically in the cartilage of embryonic rats, radiosulfate is bound and concentrated only in vesicles of the juxtanuclear Golgi apparatus of secreting chondrocytes within 3 minutes of its presentation. From this area, vacuoles migrate peripherally and lodge in the subcortex; their sulfated contents are thence discharged via stomata to the extracellular matrix. The label, apparently often associated with microvesicles at 10 and 20 minutes, is subsequently localized in the dense contents of the larger vacuoles. Bound radiosulfate is not detectable in other organelles. It is concluded that the vesicular component of the Golgi apparatus is the actual site of sulfation. Intracellular hyaluronidase-sensitive metachromatic granules are found chiefly at the cell periphery or mantle, rarely juxtanuclear in the main Golgi zone.     

CYTOCHEMICAL DIFFERENCES IN KIDNEYS FROM WINTER-HIBERNATING AND AROUSED BATS (MYOTIS LUCIFUGUS), WITH PARTICULAR REFERENCE TO THE GOLGI ZONE

Kidneys from winter bats (Myotis lucifugus) were removed and fixed in cold formalin-calcium while the animals were in the following states: (a) natural hibernation; (b) arousal from hibernation for 24 hours; (c) laboratory maintained hibernation; and (d) no hibernation since the previous winter. With fixed frozen sections, the lead salt method of Wachstein and Meisel with adenosine triphosphate as substrate (pH 7.2) showed enzymic activity localized in large vacuoles and smaller vesicles or droplets in the Golgi region of distal and proximal tubular epithelial cells of kidneys from hibernating bats. No ATPase activity was detected in the basal lamellae of tubular epithelium from hibernating bats. ATPase activity in the Golgi region was not seen in cells from kidney tubules of bats aroused from hibernation 24 hours previously or of animals that had not hibernated, whereas activity for ATPase was present in the basal infoldings of tubular epithelium from these animals. Inosine di- and triphosphatase and calcium activated ATPase activities were also detected in the Golgi region of hibernating bats but were not present in the basal infoldings of tubular epithelium from active animals. There was little or no activity toward the mono- and diphosphates of adenine, thiamine pyrophosphate, and the di- or triphosphates of guanidine, cytidine, or deoxyadenosine. The loss of enzymic activity from the Golgi region of the tubular epithelium from hibernating bats and its increase in the region of the basal infoldings of tubular epithelium in aroused bats suggests that the Golgi region plays a role in the synthesis of enzymic protein usually identified with the external cell membrane.     

Spermiogenesis in Lumbricus herculeus : An Electron Microscope Study

Small pieces of the sperm sacs of Lumbricus herculeus were fixed for 4 hours in chrome-osmium, embedded in methacrylate, sectioned with a Porter-Blum microtome, and studied with a R.C.A. EMU-2C electron microscope. Each spermatid of a group developing synchronously is attached by a cytoplasmic strand to a common nutrient protoplasmic mass. This mass contains mitochondria and yolk bodies but is anucleate. The proximal centriole, that is, the centriole nearer the nucleus, is at first associated with a small peg which becomes firmly attached to the nuclear membrane. Later these two bodies become separated during the development of the middle-piece which is differentiated in the usual manner from a nebenkern formed by the fusion of 6 or 7 mitochondria. The acrosome develops in relation to the dictyosome (Golgi body), itself composed of 8 or more individual flattened sacs and situated in the cytoplasm opposite the point of attachment of the spermatid to the nutrient mass. Soon after its formation, the acrosome becomes incorporated into a cytoplasmic appendage or acrosome carrier. The carrier moves from its original position, along the lateral border of the elongating nucleus, to the distal margin of the nucleus where the acrosome is deposited. No evidence was found of a centriole located at the point of junction between nucleus and acrosome as suggested by earlier workers.     

ACID PHOSPHATASE ACTIVITY AND ULTRASTRUCTURE OF AGED CELLS OF EUGLENA GRANULATA1,2

Ultrastructure and acid phosphatase activity of aged calls of Euglena granulata are reported. Cells are spherical, quiescent, and nonflagellated. The most conspicuous attribute of aged cells is the accumulation of cyloplasmic vacuoles and lysosome-like structures containing heavily stained, pigmented bodies and membrane fragments. In chloroplasts, portions of whorled lamellae arc abscised and subsequently incorporated into lysosome-like structures; osmiophilic granules increase in number. Membranes surrounding eyespot granules disappear and the granules themselves become diffuse; the usual association with microtubules is not seen in aged cells. Acid phosphatase precipitation accumulates largely at the maturing face of dictyosomes and associated vesicles; there is also activity in multivesicular and lysosome-like vacuoles.     

STUDIES ON THE GOLGI APPARATUS BY ELECTRON MICROSCOPY WITH PARTICULAR REFERENCE TO AOYAMA'S TECHNIQUE

1. Aoyama's silver impregnation method for the Golgi apparatus has been used on exocrine cells of the pancreas of the mouse and studied by electron microscopy in order to determine as precisely as possible where the silver is deposited. Similar cells have also been fixed in buffered osmium tetroxide solution and compared with cells treated by the silver technique. 2. Examination of the Aoyama preparations usually revealed a light deposition of silver in the cytoplasm (hyaloplasm or matrix) and a heavy deposition of silver around a series of closely apposed vacuoles. The heavy deposition of silver was regarded as revealing the chromophilic region of the Golgi apparatus while the vacuoles were identified as the chromophobic component. 3. Comparison of the silver preparations with those fixed in buffered osmium tetroxide solution showed that the silver was primarily deposited in the region of the Golgi membranes.     

Silver-impregnation of the Golgi complex in epididymal epithelial cells of mice

Localization of silver grains detected by the silver-impregnation method, a technique used to detect the classical Golgi apparatus, was examined with light and electron microscopy. Two types of silvered images of the Golgi apparatus were compared; each was obtained by Da Fano 's silver-impregnation method, and one was modified with Caulfield 's fixative during the preliminary fixation. Under ordinary light microscopy the images were very similar and showed the duplex structure of the Golgi apparatus which consists of an argentophil wall and argentophobe core. With electron microscopy, the relationship between the fine structure of the Golgi complex and the silver deposits was obtained in greater detail by the latter technique because the fine structure of the Golgi complex was retained. Many fine silver grains were detected in the cytoplasm adjacent to the Golgi complex, but none were present in the Golgi cisternae. This suggests that the argentophil wall of the duplex structure of the classical Golgi apparatus may be formed from argentophil substances that locate in the cytoplasm adjacent to the Golgi lamellae, and that the argentophobe core may be related to the Golgi lamellae.