[Intensity of respiration and oxidative phosphorylation in the ontogeny of tailless amphibia (Anura, Amphibia)].
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The phylogenetic relationship among toad, frogs and newt from Japan was investigated by means of immunoelectrophoresis. Homologous, heterologous and non-identity reactions between rabbit anti-Bufo bufo antiserum and antigen from each of 13 species and 2 subspecies of Japanese amphibia were examined. The number and positions of arcs appeared in these reactions were compared. The relationship observed in the comparison well coincided with the classification of the upper taxa of amphibia; viz 1) Hyla arborea is more intimately related to B. bufa, 2) members of Ranidae and Rhacophoridae have equal intimacy for B. bufo, and 3) Cynops pyrrhogaster is most distantly related to B. bufo. By contrasting arc obtained in non-identity reaction and those obtained in homologous reaction, it was found that toad, B. bufo carried 22 antigens, and among these antigens, one was observed through all amphibians tested, 12 were carried solely by toad, and some of the remainders were common to antigens carried by various frogs. Newts carried only one antigen common to toad. Based on these results, the phylogeny of Japanese amphibia was discussed.
Studies have been made of the distribution of mercury and its occurrence as methylmercury in the organs of amphibia collected from different sites mainly in Slovenia, Yugoslavia, including the area around the mercury mine at Idrija. Liver accumulates the highest amounts of mercury, up to 2 ppm in apparent background areas, with values up to 0.5 ppm in muscle, where virtually all mercury is present in the methyl form. Results are reported for some other trace elements in liver. Amphibia may provide useful monitors of the occurrence and spread of mercury contamination.
In the diencephalon of two species of Gymnophiona (Amphibia) two neurosecretory nuclei were examined with histological (Alcian Blue, Aldehyde Fuchsin, Brookes Trichrome stain) and enzyme histochemical techniques (acid phosphatase, alpha-naphthyl acetate esterase, acetylcholinesterase (AChE)). In the preoptic nucleus two categories of secretory neurons were distinguished: large and medium sized neurons. The perikarya of both cell types contain very little neurosecretory material. The Alcian Blue method stained the medium sized neurons faintly but selectively. The tractus praeopticohypophyseus is marked by the presence of Herring bodies, which, however, are relatively scarce. The neurohypophysis, in contrast, contains large amounts of neurosecretory material. Both cell types of the preoptic nucleus are characterized by their very strong AChE and alpha-naphthylacetate esterase activity. The AChE also marks the tractus praeoptico-hypophyseus. In the large neurons acid phosphatase is present around the nucleus; in the medium sized neurons this enzyme is concentrated close to the origin of the axon. In the dorso-caudal hypothalamus a small group of neurons is stained with Alcian-Blue. These neurons, which also contain AChE, are located immediately under the ependyma which seems to be specialized in this region.
A cytogenetic study performed on a population of the South American leptodactylid frog Eleutherodactylus maussi revealed multiple sex chromosomes of the X1X1X2X2 female/X1X2Y male (= XXAA female/XXAY male) type. The diploid chromosome number is 2n = 36 in all females and 2n = 35 in most males. The multiple sex chromosomes originated by a centric fusion between the original Y chromosome and a large autosome. In male meiosis the X1X2Y (= XXAY) multiple sex chromosomes form a classical trivalent configuration. E. maussi is the first species discovered in the class Amphibia that is distinguished by a system of multiple sex chromosomes. Only one single male was found in the population with 2n = 36 chromosomes and lacking the Y-autosomal fusion. This karyotype (XYAA male) is interpreted as the ancestral condition, preceding the occurrence of the Y-autosome fusion.
Galanin-like immunoreactivity was localized in the brain of Urodela (Ambystoma, Pleurodeles) and Anura (Bufo, Xenopus) by immunocytochemistry with anti-porcine galanin antiserum. In the four species, immunoreactive perikarya were observed in the telencephalon (striatum, amygdala), diencephalon preoptic area mainly along the anterodorsal wall of the preoptic recessus, suprachiasmatic nucleus, lateral hypothalamus, ventral and dorsal infundibular nuclei, paraventricular organ, and rhombencephalon (nucleus of the solitary tract). Galaninergic fibres extended in similar regions and in the medial septum, ventral telencephalon, ventral hypothalamus, median eminence, and various mesencephalic and rhombencephalic regions. Contacts with the cerebrospinal fluid cavity occurred along the preoptic recessus (Ambystoma) and the ventral infundibular wall (all species). Fibres were scarce in the neurohypophysis. The distal and intermediate lobes of the pituitary were virtually devoid of immunoreactivity. The galaninergic system appeared more developed in adult amphibia than in young animals, suggesting the stimulating influence of sex steroids on the expression of galanin as previously described in Anguilla. The extensive distribution of the galanin-like immunoreactive neurons in amphibian brains suggests that this peptide may act as a neuromodulatur and/or neurotransmitter.
The metabolism of histamine has been compared in Rana ridibunda and Discoglossus pictus, both Amphibia anura. An intralymphatic injection of 14C-histamine (5 microCi) was administered to the animals and the total radioactivity excreted in the urine was measured 24, 48, and 72 hours later. The metabolites excreted in the urine at 24 hours were analyzed by thin-layer chromatography and subsequent autoradiography. In both species approximately 60--70% of the injected dose was found in the urine by 72 hours. In Rana ridibunda two histamine metabolites were found: imidazoleacetic and methylimidazoleacetic acid. The present results suggest that the catabolism of histamine differs in the two species.
We have developed sensitive and reliable radioimmunoassays for T4 and T3 in amphibian plasma and have used these procedures to measure plasma T4 and T3 levels in spontaneously developing Rana catesbeiana tadpoles at various stages of metamorphosis. During premetamorphosis circulating levels of both T4 and T3 were below the limits of detection of the RIA procedures (T4 less than 50 ng/100 ml, T3 less than 5 ng/100 ml). A gradual rise in plasma T3 and T4 became apparent during prometamorphosis, and at the onset of metamorphic climax the levels of both T4 and T3 increased sharply. Peak levels for both hormones were observed in the middle of metamorphic climax (stage XXIII). The circulating T3 level reached a mean peak of 78 ng/100 mg, at least 15 times greater than the level during premetamorphosis. The peak T4 level was 0.5 microgram/100 ml, about a 10-fold increase over the premetamorphosis level. The surge in thyroid hormone secretion lasted only for several days, and during the latter half of metamorphic climax there was a fairly rapid decrease in plasma T4 and T3. By 2 days post-climax the levels had declined to about 20% of their peak values. Free T4 and T3 levels in plasma followed the same general pattern as the total hormone levels during the various stages of tadpole development. In adult R. catesbeiana, plasma T4 and T3 levels were surprisingly low. Similarly low values were observed in Bufo marinus and in Rana pipiens. The very low levels of circulating T4 and T3 both in premetamorphosis tadpoles and in adults suggest that thyroid hormones in anuran Amphibia may be of importance only during the period of metamorphosis.
Transmission electron microscopy was used to study the interface between the ureter and the mesonephric blastema. When the blastema is induced in a functional organ, numerous pseudopodial protrusions of the ureter and of the mesonephric cells lead to a close appostion between the two structures. The presence of squamous material, perhaps of a mucopolysaccharidic nature, with a network of collagen fibres is always found on this interface. These particular processes seem to play a role as a morphological basis for the phenomenon of induction. The importance of the extracellular matrix is discussed. Neither the occurrence of zones of close contact, reported by several authors, in other species, nor the fusion of membranes has ever been observed in Amphibia.
In the brain of the Caecilian species Chthonerpeton indistinctum the following enzymes have been demonstrated by means of histochemical techniques: acid phosphatase, alpha-naphthylacetate esterase, acetylcholin esterase. Acid phosphatase occurs in the cytoplasm of the neurons in 4 different types of localization. Its activity in the ventral parts of the brain is markedly higher than in the dorsal ones. Of particularly high activity are: the motor neurons in the tegmentum, the nucleus mesencephali trigemini, individual large neurons in the marginal zone of the grey matter of the telencephalon, which seems to be a special character of the Caecilians among the Amphibia. The ependyma exhibits local differences in respect of acid phosphatase activity. alpha-Naphthylacetate esterase marks in particular the secretory neurons of the hypothalamus, the large perikarya of the nucleus mesencephali trigemini and the motor neurons of the tegmentum. In the telencephalon the alpha-naphthylacetate esterase activity corresponds to that of acid phosphatase. Acetylcholin esterase marks--with certain restrictions--cholinergic neurons. These predominate in Chthonerpeton in the caudal parts of the brain. In the telencephalon amygdala, septal area striatum and the mitral cells are of comparatively high activity. The neurosecretory neurons of the hypothalamus are particularly rich in this enzyme. As an anurans the cholinergic fasciculus retroflexus as asymmetric. The tectum opticum is of secondary simplicity and does not exhibit a clearly recognizable stratification.
Cloacae were examined from male and female salamanders representing 12 genera and 22 species in the Salamandridae. All female salamandrids possess numerous sperm storage glands, spermathecae, in the roof of the cloaca, but intergeneric variation exists in the occurrence of additional cloacal glands. Pleurodeles and Tylototriton possess both vent and anterior ventral glands, and secondary loss has occurred of vent glands in all other genera and anterior ventral glands in Chioglossa, Cynops, Paramesotriton, and Triturus The most highly derived cloaca occurs in Euproctus asper, in which the cloacal tube extends through a conical projection, and ventral glands secrete onto the dorsolateral surface of the projection rather than into the cloaca. Marked intergeneric variation occurs in males in conformation of the cloacal cavities and in extent of the dorsal gland. In Cynops, Euproctus, Pachytriton, Paramesotriton, Taricha, and Triturus, the pseudopenis (a broad, posteriorly projecting evagination of the dorsal roof) fills much of the cavity of the anterior cloacal chamber. In most salamandrids, distal ends of the dorsal glands occur lateral to pelvic glands in the anterior end of the cloaca, and dorsal gland tubules descend to secretory sites at the posterior end of the vent. Salamandra and Mertensiella possess a unique, bifurcated dorsal gland in which distal ends of tubules lie dorsal to the other cloacal glands, and proximal ends curve ventrally in the anterior end of the cloaca to secretory sites along the cloacal orifice. Cladistic analyses indicate that the variation in presence of anterior ventral glands is due to homoplasy. The occurrence of female vent glands, bifurcated dorsal glands, and the pseudopenis supports a phylogeny based upon non-cloacal characters.
Light and electron microscopy of the lungs of Ambystoma tigrinum (Urodela) revealed a relatively complex pattern of the neuroendocrine (NE) cells. In the apical parts of smaller septa single NE cells not associated with nerve fibres were covered and surrounded by pneumocytes. The larger septa possessed small areas of ciliated epithelium, in which the NE cells were grouped in a form of neuroepithelial bodies (NEB) consisting of 3-5 cells and covered by goblet cells. NE cells possessed a large nucleus with patches of condensed chromatin, clear cytoplasm, and membrane-bound vesicles of variable morphology and size, containing an electron dense interior surrounded by a lucent space. The size of these dense core vesicles (DCV) ranged from 70-140 nm, while rarely the larger ones exhibited a diameter of 300-600 nm. In some NEB a second type of NE cells was observed for the first time in an amphibian species: these cells communicated with the air space and exhibited on their surface microvilli and a single modified cilium with a 8 + 1 microtubule arrangement. Their cytoplasm contained two types of DCV: dense core granules with a diameter of 140-260 nm and vesicles 320-700 nm in diameter with a moderately electron dense interior. The NEB were associated with intracorpuscular, sensory nerve terminals morphologically afferent and efferent. By immunocytochemistry, the NE cells revealed the presence of serotonin, met-enkephalin, and leu-enkephalin. A paracrine and chemoreceptor role is proposed for NEB of Ambystoma tigrinum.
The innervation of the musculature of the tongue and the hyobranchial apparatus of caecilians has long been assumed to be simple and to exhibit little interspecific variation. A study of 14 genera representing all six families of caecilians demonstrates that general patterns of innervation by the trigeminal, facial, glossopharyngeal, and vagus nerves are similar across taxa but that the composition of the "hypoglossal" nerve is highly variable. Probably in all caecilians, spinal nerves 1 and 2 contribute to the hypoglossal. In addition, in certain taxa, an "occipital," the vagus, and/or spinal 3 appear to contribute fibers to the composition of the hypoglossal nerve. These patterns, the lengths of fusion of the contributing elements, and the branching patterns of the hypoglossal are assessed according to the currently accepted hypothesis of phylogenetic relationships of caecilians, and of amphibians. An hypothesis is proposed that limblessness and a simple tongue, with concomitant reduced complexity of innervation of muscles associated with limbs and the tongue, has released a constraint on pattern of innervation. As a consequence, a greater diversity and, in several taxa, greater complexity of neuroanatomical associations of nerve roots to form the hypoglossal are expressed.
Histology of the cloacae of Rhyacotriton olympicus and representative species from the genera Ambystoma and Dicamptodon was examined by light microscopy. Females of Ambystoma possess sperm storage glands, the spermathecae, as well as ventral glands and dorsal glands, both of uncertain function. Females of Ambystoma examined from the subgenus Linguaelapsus differ from those in the subgenus Ambystoma by possessing more extensive ventral gland clusters and a shorter cloacal tube. Females of Dicamptodon possess spermathecae and ventral glands, but differ in cloacal conformation from females of Ambystoma and lack the dorsal glands. Females of R. olympicus possess more extensive epidermal lining in the cloaca than that found in females of Ambystoma and Dicamptodon, and the only glands present are spermathecae, which cluster around a tube in the dorsal roof. Males of Ambystoma, Dicamptodon, and R. olympicus possess five types of cloacal glands (dorsal pelvic glands, lateral pelvic glands, anterior ventral glands, posterior ventral glands, and Kingsbury's glands) that function in spermatophore formation, and vent glands that may produce a courtship pheromone. In Ambystoma and Dicamptodon, vent glands secrete along the medial borders of the cloacal orifice. Males of A. opacum and A. talpoideum differ from males of other species examined from the two genera by possessing more extensive vent glands. Males of R. olympicus possess unique vent glands in which tubules secrete onto the surface of vent lobes lateral to the posterior end of the cloacal orifice, and distal ends of the glands pass anteriorly, superficial to the fascia enclosing the other cloacal glands. The results from analysis of cloacal anatomy support other data indicating that Ambystoma and Dicamptodon are sister groups, and that Rhyacotriton olympicus is not closely related to either of the other two genera and merits placement in a separate family.
Microcorrosion casts of the renal vascular system of tadpoles of the Clawed Frog, Xenopus laevis, were observed by scanning electron microscopy. Glomerular differentiation was studied qualitatively and quantitatively during developmental stages 56-66 (metamorphic climax). The general structure of the renal vascular system corresponds to the pattern commonly found in anurans; however, the arterial supply has conspicuous connecting vessels that supply groups of glomeruli. In the dorsal part of the kidney, qualitative differentiation of glomerular structures precedes quantitative growth. The ventral part of the kidney has larger, well-developed renal corpuscles of nearly adult appearance. Four developmental stages of glomerulogenesis are distinguished morphologically and their glomerular and vascular growth is analyzed.
The neuronal typology in the hypothalamus of the frog and the crested newt was studied by the Golgi technique. In the newt, piriform, multipolar or cerebrospinal fluid (CSF)-contacting neurons of relatively primitive type, according to the classification of Ramón-Moliner, are encountered in the preoptic area. Moreover, magnocellular neurons are impregnated. In the frog the preoptic area shows a more varied typology. The posterior hypothalami of the frog and the newt exhibit mainly bipolar CSF-contacting and piriform neurons. These latter are generally "tufted", but some bipolar of multipolar cells are encountered, especially in the frog. The simple anatomical organization of the amphibian hypothalamus corresponds well with the pattern of "generalized" integrative area where multimodal sensory inputs converge--including visceral information from cerebrospinal fluid by means of hypothalamic CSF-contacting sensors--to regulate the neuroendocrine outflow.
The uterine epithelium of the viviparous Salamandra atra and the ovoviviparous Salamandra salamandra was studied in non pregnant and ovulating females and in females during different stages of pregnancy. The epithelium of both species is organized in a monolayer. The epithelial cells are characterized by a moderate secretory activity, a variable amount of apical granules which include PAS-positive material and by some apical and basal exo- or endocytotic vesicles. Adjacent cells are joined by junctional complexes. The lateral surfaces form a tortuous boundary with adjoining cells which suggest that the epithelium is involved in transport. Sporadic light cells possess highly variable cytoplasmic inclusions and are not joined with neighbouring cells. Possibly they represent migratory cells. The entire epithelium, except for a small cranial portion of the uterus in S. atra, undergoes no remarkable morphological changes during the different physiological stages examined except that flattened cells seem to be more numerous in pregnant females. The results are discussed with regard to the possible supply of the developing young by the mother.
The angioarchitecture of the neural stalk and the encephaloposthypophysial portal system of the hypophysis of the toad, Bufo bufo (L.), was studied using three different methods. The neural stalk is mainly supplied by branches of the arteria infundibularis superficialis which form a widemeshed vascular network. Dorsally this network continues into the plexus of the pars nervosa. The vascularization of the pars nervosa is made up of the encephalo-posthypophysial portal system. This portal system consists of a hypothalamic branch (=portion), a mesencephalic and a mesencephalicbulbar branch (=portion). The hypothalamic branch was found to drain the pars ventralis of the tuber cinereum as well as more dorsal regions of the diencephalon. The mesencephalic-bulbar trunk enters the hypothalamic branch. The resulting common stem of the encephalo-posthypophysial portal vein the curves around the retroinfundibular communicating artery, crosses its ventral side and runs caudally. The secondary capillary plexus of the pars nervosa is characterized by well defined capillary plexus of the pars nervosa is characterized by well defined capillary networks which are located at the periphery of the parenchyma of the pars nervosa, thus forming a rostral, dorsal and ventro-caudal net. The central region of the parenchyma of the pars nervosa is supplied only by main branches of the encephalo-postpophysial portal vein. The venous drainage of the pars nervosa is via the vena hypophysea transversa.