Scanning electron microscopy of the electric organs of Electrophorus electricus L

Summary The surfaces of the organs of Sachs and Hunter of Electrophorus electricus L. were examined by scanning electron microscopy. Special attention was directed to morphological details of the electrocyte to provide a better understanding of its anterior and posterior faces. Some aspects of the microanatomy of these organs, which differ markedly from those of the main electric organ, provide new information on the structure as revealed previously by light and transmission electron microscopy. The relief, mainly expressed by papillae, is related to the actual membrane area, which is important for calculations of specific resistance and conductance. Information is also presented on the general organization of the tissue, in particular the distribution of the connective elements and external configuration of synaptic terminals. Shrinkage in preparation of tissue was evaluated and correction made whenever necessary. Correction factors for actual membrane area were calculated for anterior and posterior faces of electrocytes from both organs.     

Innervation pattern and electric organ discharge waveform in Gymnotus carapo (Teleostei; Gymnotiformes)

The electrogenic organ (EO) of Gymnotus carapo has two main portions: a posterior region consisting of four bilaterally arranged electrocyte rows; and an anterior portion composed of only two. The lateral row (LR) of the anterior portion contains doubly innervated electrocytes with axon terminals from different nerves on their rostral and caudal faces. The LR is continuous with the most dorsal row of the caudal region. This row also contains doubly innervated electrocytes. The medial row (MR) electrocytes of the anterior region and ventral rows of the caudal region are exclusively caudally innervated. All caudal faces of the anterior or abdominal region are supplied by two nerves which originate from spinal roots VIII to XXI. Roots I to VII give origin to pure rostral nerves whose electromotor axons terminate on the rostral surfaces of the first seven LR electrocytes. A given doubly innervated electrocyte is supplied on its caudal face by a nerve originating several segments (usually seven) posterior to the spinal root supplying its rostral face. Transections of the spinal cord at the level of root VIII isolate the activity of the rostral surfaces of the first electrocytes. The EO discharge (EOD) then appears as a head negative deflection which arises from abdominally located electrocytes. Its monophasic character reveals that the activity remains restricted to the rostral electrocyte surfaces. Damage of the abdominal portion of the EO abolishes the first negative deflection of the normal pulse. Transections of the spinal cord at the level of root XXI isolate the activity of the whole abdominal portion of the EO. Since both doubly and singly innervated electrocytes remain active, the EOD appears biphasic. Comparative studies have shown that the EOD of Hypopomus sp. lacks any early negative wave and correspondingly all its electrocytes are exclusively caudally innervated.     

Caudal Papillae in Romanomermis culicivorax

Distribution of caudal papillae in adult Romanomermis culicivorax was determined by scanning electron microscopy. Ninety eight caudal papillae, each containing one pore, were present in males but absent in females. Papillae were arranged in three longitudinal rows, one ventral, two ventrolateral; the middle ventral row bifurcated anterior to the spicule. The appearance of the papillae was different anterior and posterior to the spicule. The role of the caudal papillae in mediating copulatory behavior was discussed.     

An electron microscopic investigation of the surface coat of the electrocyte of Electrophorus electricus

Summary The surface coat of the electrocyte of the main electric organ of Electrophorus electricus was studied using cytochemical methods (periodic acid-silver methenamine, periodic acid-chromic acid-silver methenamine, periodic acid-thiosemicarbazide-silver proteinate, Concanavalin A — horseradish peroxidase, ruthenium red, Alcian-blue lanthanum nitrate, colloidal iron hydroxide and cationized ferritin). The surface of the electrocyte presents perpendicularly oriented tubular invaginations of the cell membrane. The fibrous coat 50–100 nm thick, penetrates into the lumen of the invaginations. It is also observed in the synaptic clefts existent in the posterior face of the electrocyte. The coating of the surface membrane gives a positive reaction with all techniques used. Binding of colloidal iron hydroxide particles was observed only in the outer layer of the coat. With the Alcian-blue lanthanum nitrate technique, microtubules were observed in the cytoplasm of the electrocyte.The results indicate that the surface coat of the electrocyte contains mucopolysaccharides, glycoproteins, acid mucopolysaccharides and anionic sites detected at low (colloidal iron hydroxyde) and neutral (cationized ferritin) pH.This work has been supported by Conselho Nacional de Desenvolvimento Cientifico e Tecnológico (CNPq), Conselho de Ensino e Pesquisa da UFRJ (CEPG) and Banco Nacional de Desenvolvimento Econômico     

Co-distribution of Tropomyosin and α-Actinin With Actin in Psammobatis extenta Electrocytes Brings Out Their Similarity with Muscle Fiber Cytoplasm

Electric organs of Psammobatis extenta (Rajiformes) electric fish derive from myoblasts of the caudal region [16]. Here we study the presence of muscle proteins, actin and the actin-binding proteins, α-actinin and tropomyosin, in the electrocytes by means of biochemical approaches, scanning electron microscopy and immunocytochemical methods. NBD-phallacidin is employed to detect the filamentous form of actin (F-actin). Immunoblots of actin and α-actinin from P. extenta skeletal and smooth muscle show that the electric organ forms of actin and α-actinin correspond to muscle types. Scanning electron microscopy shows that P. extenta electrocytes are highly polarized cells, semicircular in shape, with an anterior, concave innervated face and a posterior, convex, non-innervated face. The immunofluorescence patterns of α-actinin and tropomyosin distribution are similar to those of actin, in that these epitopes appear to occur throughout the entire electrocyte cytoplasm. F-actin, as revealed by NBD-phallacidin fluorescence, was also found throughout the cytoplasm. This is the first time that evidence is presented to demonstrate the existence of muscle actin in this weak electric fish species electrocyte. The close evolutionary connection to that of muscle cells is discussed.     

Waveform generation in Rhamphichthys rostratus (L.) (Teleostei,Gymnotiformes)

Rhamphichthys rostratus (L.) emits brief pulses (2 ms) repeated very regularly at 50 Hz. The electric organ shows a heterogeneous distribution of the electrocyte tubes and the occurrence of three electrocyte types (caudally innervated, rostrally innervated and marginallycaudally innervated). In the sub-opercular region the electric organ consists of a pair of tubes containing only caudally innervated electrocytes. At the abdominal region the EO consists of three pairs of tubes. Each pair contains one of the described electrocyte types. The number of electrocyte tubes increases toward the tail to reach nine or ten pairs in the most caudal segments. In the intermediate region most tubes contain doubly innervated electrocytes except the ventral pair that contains caudally innervated electrocytes. The caudal 25% contains exclusively caudally innervated electrocytes. The electric organ discharge consists of five wave components (V1 to V5). Electrophysiological data are consistent with the hypothesis that V1 results from the activity of the rostral faces of rostrally innervated electrocytes. V2 results from the activities of rostral faces of marginally-caudally innervated electrocytes while V3 results from the activities of caudal faces of most electrocytes. Curarization experiments demonstrated that V4 and V5 result from action potential invasion and are not directly elicited by neural activity.Abbreviations AEN1 anterior electromotor nerve 1 - AEN2 anterior electromotor nerve 2 - BMB boraxic methylene blue - CIE caudally innervated electrocytes - EMF electromotive force - EO electric organ - EOD electric organ discharge - I current amplitude - MCIE marginally-caudally innervated electrocytes - MT medial tubes - PEN posterior electromotor nerve - R _n internal impedance - RIE rostrally innervated electrocytes - Rl load resistor - SAT short abdominal tubes - V voltage amplitude     

Electric organ morphology of Sternopygus macrurus,a wave-type,weakly electric fish with a sexually dimorphic EOD

In several species of electric fish with a sex difference in their pulse-type electric organ discharge (EOD), the action potential-generating cells of the electric organ (electrocytes) of males are larger and more invaginated compared to females. Androgen treatment of females and juveniles produces a longer-duration EOD pulse that mimics the mature male EOD, with a concurrent increase in electrocyte size and/or membrane infolding. In Sternopygus macrurus, which generates a wave-type EOD, androgen also increases EOD pulse duration. To investigate possible morphological correlates of hormone-dependent changes in EOD in Sternopygus, we examined electric organs from both fish collected in the field, and untreated and androgen-treated specimens in the laboratory. The electrocytes are cigar shaped, with prominent papillae on the posterior, innervated end. Electrocytes of field-caught specimens were significantly larger in all parameters than were electrocytes of specimens maintained in the laboratory. EOD pulse duration and frequency were highly correlated, and were significantly different between the sexes in sexually mature fish. Nevertheless, no sex difference in electrocyte morphology was observed, nor did any parameters of electrocyte morphology correlate with EOD pulse duration or frequency. Further, whereas androgen treatment significantly lowered EOD frequency and broadened EOD pulse duration, there was no difference in electrocyte morphology between hormone-treated and control groups. Thus, in contrast to results from studies on both mormyrid and gymnotiform pulse fish, electrocyte morphology is not correlated with EOD waveform characteristics in the gymnotiform wave-type fish Sternopygus. The data, therefore, suggest that sex differences in EOD are dependent on changes in active electrical properties of electrocyte membranes. © 1992 John Wiley & Sons, Inc.     

Biochemical and cytochemical localization of ATPases on the membranes of the electrocyte of Electrophorus electricus

Summary The localization of (Na⁺-K⁺) ATPase in the intact electrocyte of the electric organ of Electrophorus electricus (L.) and its subcellular fractions was investigated by biochemical and cytochemical methods. The distribution of AChE activity in the subcellular fractions was also comparatively analysed with this enzyme serving as a marker of the innervated membranes of the electrocyte. After application of cytochemical method of Farquhar and Palade to glutaraldehyde-fixed tissue, reaction was observed only at the membranes of vesicles localized at the periphery of the electrocyte. Previously fixed electrocytes, incubated in Ernst's medium showed reaction only at the vesicles whereas in unfixed tissue reaction also appeared at other membranes (surface and invaginations) of the anterior and posterior faces. This reaction was significantly inhibited in the presence of ouabain or in the absence of K⁺. Inhibition of Na⁺-K⁺-ATPase by glutaraldehyde fixation was also confirmed by biochemical analysis.This investigation has been supported by Conselho Nacional de Desenvolvimento Científico e Tecnológico, Conselho de Ensino e Pesquisa da UFRJ and FINEP (FNDCT-375/CT)     

Cuticular structures of the priapulid Halicryptus spinulosus: A scanning electron microscopical study

Summary The surface anatomy and the structures lining the pharynx of Halicryptus spinulosus were examined by scanning electron microscopy (SEM). The structures were compared and contrasted with those reported for other priapulids, particularly those features previously studied with SEM. Buccal papillae and pharyngeal teeth of two types were described. Surface structures observed with SEM were: scalids, abdominal setae, anal papillae, posterior warts and ring papillae. The latter three structures are unique among described priapulids. The anal papillae are composed of several rounded, perhaps pedunculate, structures; the posterior warts are composed of mitriform structures in close association with columnar structures. Both are located in separate depressions in the posterior integument. The ring papillae occur on the annuli close to the posterior end. Halicryptus spinulosus was previously thought to lack these structures.     

Development of the electromotor system of Torpedo marmorata: Distribution of extracellular matrix and cytoskeletal components during acetylcholine receptor focalization

Summary A combination of direct fluorescence and indirect immunofluorescence microscopy has been used to compare the distribution of the acetylcholine receptor with the distribution of major cytoskeletal and extracellular matrix components during electrocyte differentiation in the electric organs of Torpedo marmorata. Laminin, fibronectin and extracellular matrix proteoglycan are always more extensively distributed around the differentiating cell than the acetylcholine receptor-rich patch that forms on the ventral surface of the cell. The distribution of acetylcholinesterase within the ventral surface of the differentiating electrocyte closely resembles the distribution of the acetylcholine receptor. Areas of apparently high acetylcholine receptor density within the ventrally forming acetylcholine receptor-rich patch are always areas of apparently high extracellular matrix proteoglycan density but are not always areas of high laminin or fibronectin density. Desmin levels appear to increase at the onset of differentiation and desmin initially accumulates in the ventral pole of each myotube as it begins to form an electrocyte. During differentiation F-actin-positive filament bundles are observed that extend from the nuclei down to the ventrally forming acetylcholine receptorrich patch. Most filament bundles terminate in the acetylcholine receptor-rich region of the cell membrane. Electronmicroscopic autoradiography suggests that the filament bundles attach to the membrane at sites where small acetylcholine receptor clusters are found. The results of this study suggest that, out of the four extracellular matrix components studied, only the distribution of acetylcholinesterase (which may be both matrix- and membrane-bound at this stage) closely parallels that of the acetylcholine receptor, and that F-actin filament bundles terminate in a region of the cell that is becoming an area of high acetylcholine receptor density.Abbreviations ACHR nicotinic acetylcholine receptor - ACHE acetylcholinesterase - BSA bovine serum albumin - EMPG extracellular matrix proteoglycan fraction - FITC fluorescein isothiocyanate - FN fibronectin - LN laminin - TBS Tris-HCl-buffered saline - SDS PAGE sodium dodecyl sulphate polyacrylamide gel electrophoresis     

Surface ultrastructure of the advanced third-stage larvae of Gnathostoma nipponicum

The surface ultrastructure of advanced third-stage larvae (AL3) of Gnathostoma nipponicum was studied using scanning electron microscopy. The larvae were recovered from the grass snake Rhabdophis tigrina in the Republic of Korea. Parasites had a globular head bulb with a pair of lips at the anterior end and 2 labial papillae and an amphid on each lip. The head bulb was characteristically armed with 3 transverse rows of hooklets, averaging 36, 38, and 43 in number, increasing posteriorly. A total of 213-232 minute unidentate cuticular spines were present along the entire length of the larvae, forming the transverse striations. Two pairs of cervical papillae were located between the 8th and 12th transverse striations, and a pair of body papillae was seen laterally on the posterior third of the body. A pair of caudal phasmids was recognized near the posterior extremity. The surface ultrastructure of AL3 of G. nipponicum is unique compared with that of other species.     

Head morphology and prehensile adaptations in Hedruris spinigera (Nematoda: Spirurida)

Abstract

SEM examination of heads of Hedruris spinigera failed to reveal any posterior pair of papillae as structures that disturb the surface contour of the pseudolabia. In most specimens amphid apertures could not be detected either. All heads examined as transparent en face preparations showed subsurface structures that could be interpreted as ‘papillae’ and amphids. Modifications of the posterior cuticle of both sexes, and associated with the unusual methods of caudal prehension found in this species, are illustrated and discussed.     

A new species of the genus Goezia Zeder, 1800 (Nematoda: Anisakidae) from the fish,Mastacembelus armatus (Lacep.) from West Bengal,India

Goezia moraveci n. sp. is described from light microscope and scanning electron microscope studies of the specimens recovered from the freshwater fish Mastacembelus armatus from West Bengal, India. G. moraveci differs from other species of the genus in having a small body size, the excretory pore posterior to the level of the nerve-ring, a very short, wide intestinal caecum and a long ventricular appendage (ratio 1: 6–15), a different number and arrangement of caudal papillae, and cuticular spines surrounding the bases of the caudal papillae. This represents the first species of the genus from a piscine host in India.     

A pineal ganglion associated with the pineal tract in the domestic fowl

Summary A ganglion-like aggregate consisting of acetyl-cholinesterase-positive neurons was demonstrated in the pineal organ of the domestic fowl by means of light and electron microscopy. This ganglion is located in juxtaposition with the pineal tract at the posterior (caudal) aspect of the pineal stalk. Numerous large and small neurons formed the ganglion in 40-day-old domestic fowl. Some of these nerve cells established direct neuro-neuronal contacts, others were surrounded by satellite cells. These ganglion cells displayed axo-somatic and axo-dendritic synapses. The above-mentioned cluster of nerve cells may be considered as a pineal ganglion. Its central or peripheral nature is open to discussion. Send offprint requests to: Dr. K. Wake, Department of Anatomy, Faculty of Medicine, Tokyo Medical and Dental University, Yushima, Bunkyo-ku, 113, Japan     

Shh signalling restricts the expression of Gcm2 and controls the position of the developing parathyroids

The parathyroid glands originate from the endoderm of the caudal pharyngeal pouches. How these parathyroids are restricted to developing in the caudal pouches is unclear. In this paper we investigate the role of Shh signalling in patterning the vertebrate pharyngeal pouches, and show that Hh signalling may be involved in restricting the expression of the parathyroid marker Gcm2 in the pharyngeal epithelium. In the chick and mouse, Shh signalling is excluded or highly reduced in the posterior/caudal pouches, where the parathyroid marker Gcm2 is expressed, while remaining at high levels in the more anterior pouches. Moreover, though the block of Shh signalling at early developmental stages results in the loss of chick Gcm2 expression, at later stages, it induces ectopic Gcm2 expression domains in the second and first pharyngeal epithelium, suggesting that HH signalling prevents Gcm2 in those tissues. These ectopic domains go on to express other parathyroid markers but do not migrate and develop into ectopic parathyroids. Differences in the expression of Gcm2 in the chick, mouse and zebrafish, correlate with changing patterns of Shh signalling, indicating a conserved regulatory mechanism that acts to define pouch derivatives.     

Observations on the Caudal Cilium of the Tomite of Ichthyophthirius multifiliis Fouquet, 18761,2

Light microscopy of live or silver-impregnated specimens of the fish parasite Ichthyophthirius multifiliis show that the tomites are elongated and claviform with the anterior end broad. The cytostome, indicated by the presence of the organelle of Lieberkühn, is found in the lower part of the broadened anterior third of the tomite. The tapered posterior end bears a rigid, caudal cilium at its pole. Scanning electron microscopy reveals the caudal cilium and associated structures, including the depression from which the cilium protrudes, circumciliary ring, and raised struts on the ring. From these observations it is concluded that a previously reported “apical filament” found on the tomite is actually the posterior caudal cilium described by Canella & Rocchi-Canella in 1976.     

Systematic implications of male caudal morphology in ascaridoid nematode parasites

In view of earlier results, suggesting a considerable stability in the number and distribution patterns of caudal papillae and papillae-like structures in the male of some ascaridoid species, these features are analysed more generally within the superfamily Ascaridoidea. They include the single pair of phasmids on the tail, four pairs of distal papillae some of which are considered in some cases to occur anterior to the level of the cloaca, two pairs of paracloacal papillae, a median papilla or plaque, and numerous pairs of proximal papillae some of which may be situated posterior to the level of the cloaca. The exceptions to this general pattern and the systematic and phylogenetic significance of caudal morphology are discussed.     

Scanning and transmission electron microscope studies on the sensory sucker papillae of the fish parasite Entobdella soleae (monogenea)

Summary The ventral surface of the posterior sucker of Entobdella soleae, a monogenean skin parasite of the sole Solea solea, is covered with more than 800 papillae ranging in size from 2.5–19 m in diameter. The papillae are penetrated by nerves which double over themselves to form a stack-like array of lamellae running parallel to the surface of the haptor. No cilia or associated structures are present within these presumed sense organs and the papillae have no opening to the exterior. The much larger sucker papillae of a related species, E. hippoglossi, have been shown to have a similar ultrastructure. The possibility that the papillae may be contact receptors or strain receptors, providing proprioceptive information assisting the coordination of the attachment organ, is discussed and the papillae are compared with vertebrate touch receptors.I should like to thank Colin Ogden of the British Museum of Natural History for the generous assistance with the scanning electron microscopy and the Director and Staff of the Plymouth Laboratory and Ken Mackenzie and Rod Wootten of the Marine Laboratory, Aberdeen for help in obtaining material.     

Continuity between the ventricular and subarachnoid cerebrospinal fluid in an amphibian,Rana pipiens

Summary Continuity between the ventricular and subarachnoid cerebrospinal fluid has been investigated in Rana pipiens. The structure of the posterior tela, a deficient membrane situated at the extreme caudal end of the roof of the fourth ventricle, has been studied using whole membrane mounts and by light microscopy of resin embedded tissue. The ependymal component consists of columnar and rounded cells which form a regular syncytium enclosing round and oval fenestrations. Small fenestrations are covered on the subarachnoid side by elongated pial cells and thus do not give total continuity between the fourth ventricle and the subarachnoid space. Large fenestrations, on the other hand, are accompanied by equivalent pial fenestrations giving direct access between the fluid compartments. Towards the caudal end the fenestrations break up and the numbers of ependymal and pial cells decrease, the caudal end itself being characterised by a small remaining clump of ependyma and pia or of pia alone.Flow through the tela has been studied using fluorescein-labelled dextran placed in the intraventricular space. Infusion into the lateral ventricle and subsequent localisation by fluorescence microscopy shows the marker to be in the fourth ventricle, in the fenestrations of the posterior tela and in the subarachnoid space overlying the tela. Infusion of the marker followed by freezing and examination of the cut heads on a freezing microtome, shows fluorescence throughout the ventricular system, in the subarachnoid space adjacent to the posterior tela and also along the dorsal subarachnoid space of the spinal cord.