Histological characteristics of gills and dorsal skin in ambystoma leorae and ambystoma rivulare: morphological changes for living at high altitude / Características histológicas de la piel del dorso y las branquias en ambystoma leorae y ambystoma rivulare: cambios morfológicos para vivir a gran altitud

SUMMARY: Vertebrates exhibit structural changes in their cardiovascular and gas exchange systems in response to hypoxic conditions in high altitude environments. In highland neotenic mole salamanders, as other amphibians, the majority of gases exchange is carried out for skin and gills. But, in high altitude environments, the available oxygen is lower than it is in the air thus, the scarcity of oxygen limits the survival of organisms. Many studies on this subject have focused on understanding the hematological mechanisms that amphibians exhibit in response to hypoxia. However, little is known about possible morphological changes in respiratory structures that may permit increased gas exchange during respiration in high altitude amphibians like Ambystoma leorae and A. rivulare, two threatened Mexican salamander species. The aim of the present study was to describe and compare the histological characteristics of the gills and dorsal skin of A. leorae and A. rivulare from populations at low and high altitudes. We found that, in comparison to lowland organisms, highland ones exhibited more pronounced skin folds, greater numbers of secondary branches in the gills, thinner dorsal and gill epidermises, and greater quantity of melanin surrounding the gill blood vessels. These differences permit a greater capacity for gas exchange and also increase thermoregulatory capacity in high altitude environments.

RESUMEN: Los anfibios que viven en ambientes de altitud se enfrentan a factores abióticos que limitan la vida, tales como la disminución de la presión barométrica con la consecuente disminución de la presión parcial de oxígeno (O2). Conocer los mecanismos que optimizan la obtención del O2 en estos animales es de gran importancia para entender las diferencias en la sensibilidad a la hipoxia de las diferentes especies. Ambystoma rivulare y A. leorae son anfibios endémicos del Estado de México que viven en ambientes de alta altitud por lo que se cree presentan estrategias fenotípicas para asimilar eficazmente el O2 y poder subsistir en los ambientes de altitud. El objetivo de este trabajo fue analizar las características histológicas de branquias y piel cefálica (que son las principales estructuras que se encargan del intercambio gaseoso) provenientes de tres poblaciones con diferente altitud. Nuestros resultados muestran que los organismos que habitan a mayor altitud tienden a aumentar la superficie de intercambio gaseoso, como es el caso de pliegues epidérmicos y ramas branquiales secundarias. Las diferencias histológicas de branquias y piel cefálica tanto interespecíficas como intraespecíficas respecto a la altitud parecen apoyar la idea de que los organismos modifican sus estructuras para contrarrestar las limitantes de la vida en ambientes de altitud.

Adenosine suppresses exocytosis from cone terminals of the salamander retina.

In the retina, adenosine is released in the dark and has been shown to inhibit Ca2+ influx through voltage-gated Ca2+ channels in cones. Therefore, we tested whether adenosine can inhibit exocytosis from isolated cone photoreceptors. Simultaneous measurements of membrane exocytosis and Ca2+ were made from cones using the activity-dependent dye, Synaptored-C2, and the Ca2+ indicator dye, Fluo-4. Adenosine suppressed exocytosis in cones, indicating that transmitter release is also reduced from cone terminals, and further supports an inhibitory mechanism for modulating transmitter release onto second-order neurons. Furthermore, this raises the possibility that adenosine might be neuroprotective for photoreceptors and second-order neurons by suppressing Ca2+ levels in cones and reducing exocytosis of L-glutamate, respectively.

Paedomorphosis in Ambystoma talpoideum: effects of initial body size variation and density.

Facultative paedomorphosis is the ability of a salamander to either metamorphose into a terrestrial, metamorphic adult or retain a larval morphology to become a sexually mature paedomorphic adult. It has been hypothesized that density and initial body size variation within populations are instrumental in cueing metamorphosis or paedomorphosis in salamanders, yet few studies have adequately tested these hypotheses in long-term experiments. Beginning in the spring of 2004, 36 experimental ponds were used to manipulate three body size variation levels (low, medium, high) and two density levels (low, high) of Ambystoma talpoideum larvae. Larvae were individually marked using visible implant elastomers and collected every 2 weeks in order to measure snout-vent length and mass. Bi-nightly sampling was used to collect new metamorphs as they appeared. Analysis revealed significant effects of density, size variation and morph on body size of individuals during the summer. Individuals that metamorphosed during the fall and following spring were significantly larger as larvae than those becoming paedomorphic across all treatments. These results support the Best-of-a-Bad-Lot hypothesis, which proposes that the largest larvae metamorphose in order to escape unfavorable aquatic habitats. Large larvae may metamorphose to leave aquatic habitats, regardless of treatment, due to the colder climate and lower productivity found in Kentucky, which is in the northern-most part of A. talpoideum's range. By maintaining a long-term experiment, we have provided evidence for the transition of both larvae and paedomorphs into metamorphs during fall and spring metamorphosis events. Furthermore, the production of similar morphs under different environmental conditions observed in this research suggests that the ecological mechanisms maintaining polyphenisms may be more diverse that first suspected.

Developmental and evolutionary origins of the vertebrate dentition: molecular controls for spatio-temporal organisation of tooth sites in osteichthyans.

The rainbow trout (Oncorhynchus mykiss) as a developmental model surpasses both zebrafish and mouse for a more widespread distribution of teeth in the oro-pharynx as the basis for general vertebrate odontogenesis, one in which replacement is an essential requirement. Studies on the rainbow trout have led to the identification of the initial sequential appearance of teeth, through differential gene expression as a changing spatio-temporal pattern, to set in place the primary teeth of the first generation, and also to regulate the continuous production of replacement tooth families. Here we reveal gene expression data that address both the field and clone theories for patterning a polyphyodont osteichthyan dentition. These data inform how the initial pattern may be established through up-regulation at tooth loci from a broad odontogenic band. It appears that control and regulation of replacement pattern resides in the already primed dental epithelium at the sides of the predecessor tooth. A case is presented for the developmental changes that might have occurred during vertebrate evolution, for the origin of a separate successional dental lamina, by comparison with an osteichthyan tetrapod dentition (Ambystoma mexicanum). The evolutionary origins of such a permanent dental lamina are proposed to have occurred from the transient one demonstrated here in the trout. This has implications for phylogenies based on the homology of teeth as only those developed from a dental lamina. Utilising the data generated from the rainbow trout model, we propose this as a standard for comparative development and evolutionary theories of the vertebrate dentition.

Expression of calcium transporters in the retina of the tiger salamander (Ambystoma tigrinum).

Changes in intracellular calcium concentration, [Ca2+]i, modulate the flow of visual signals across all stages of processing in the retina, yet the identities of Ca2+ transporters responsible for these changes are still largely unknown. In the current study, the distribution of plasma membrane and intracellular Ca2+ transporters in the retina of tiger salamander, a model system for physiological studies of retinal function, was determined. Plasma membrane calcium ATPases (PMCAs), responsible for high-affinity Ca2+ extrusion, were highly expressed in the salamander retina. PMCA isoforms 1, 2, and 4 were localized to photoreceptors, whereas the inner retina expressed all four isoforms. PMCA3 was expressed in a sparse population of amacrine and ganglion neurons, whereas PMCA2 was expressed in most amacrine and ganglion cells. Na+/Ca2+ exchangers, a high-capacity Ca2+ extrusion system, were expressed in the outer plexiform layer and in a subset of inner nuclear and ganglion layer cells. Intracellular Ca2+ store transporters were also represented prominently. SERCA2a, a splice variant of the sarcoplasmic-endoplasmic Ca2+ ATPase, was found mostly in photoreceptors, whereas SERCA2b was found in the majority of retinal neurons and in glial cells. The predominant endoplasmic reticulum (ER) Ca2+ channels in the salamander retina are represented by the isoform 2 of the IP3 receptor family and the isoform 2 of the ryanodine receptor family. These results indicate that Ca2+ transporters in the salamander retina are expressed in a cell type-specific manner.

Goalpha labels ON bipolar cells in the tiger salamander retina.

By using double-label immunocytochemistry and confocal microscopy, we studied rod and cone synaptic contacts, photoreceptor-bipolar cell convergence, and patterns of axon terminal ramification of ON bipolar cells in the tiger salamander retina. An antibody to recoverin, a calcium-binding protein found in photoreceptors and other retinal neurons in various vertebrates, differentially labeled rods and cones by lightly staining rod cell bodies, axons, and synaptic pedicles and heavily staining cone cell bodies and pedicles. An antibody to G(oalpha) labeled most ON bipolar cells, with axon terminals ramified mainly in strata 6-9 and a minor band in stratum 3 of the inner plexiform layer (IPL). Stratum 10 of the IPL was G(oalpha) negative, and previous studies showed that axon terminals of rod-dominated ON bipolar cells are monostratified in that stratum. The axonal morphology of G(oalpha)-positive cells resembled that of the cone-dominated (DBC(C)) or mixed rod and cone ON (DBC(M)) bipolar cells. The G(oalpha)-positive dendritic processes made close contact with all cone pedicles and superficial contact with some rod pedicles, consistent with the idea that G(oalpha) subunits are present in DBC(C)s and DBC(M)s. The size and density of these cells were analyzed, and their spatial distributions were determined. To our knowledge, this is the first study to characterize photoreceptor inputs and axon terminal morphology of a population of ON bipolar cell with the use of a G(oalpha) antibody as an immunomarker in the salamander retina.

Pharmacological characterization of P2Y receptor subtypes on isolated tiger salamander Müller cells.

Müller cells express a variety of neurotransmitter receptors that permit them to "sense" the extracellular environment within the retina. We have used a battery of agonists and antagonists to characterize the purinergic receptor subtypes expressed on isolated tiger salamander Müller cells. Changes in intracellular calcium ion concentration ([Ca(2+)](i)) in Müller cells were measured using the Ca(2+) indicator dye Fura-2 and digital imaging microscopy. ATP, 2-methylthio-ATP, 2-methylthio-ADP, ADP, UTP, UDP, deoxyATP, and 3'-O-(4-benzoyl)benzoyl ATP evoked increases in [Ca(2+)](i) in both the presence and absence of extracellular Ca(2+). Therefore, the increases we observed were likely due to intracellular Ca(2+) release mediated by G-protein-coupled P2Y receptor activation, rather than Ca(2+) influx via P2X receptor channels. The P2Y(1) receptor agonists 2-methylthio-ATP, 2-methylthio-ADP, and ADP evoked increases in [Ca(2+)](i) that were inhibited by the P2Y(1) receptor antagonists adenosine 3'-phosphate 5'-phosphosulfate and 2'-deoxy-N(6)-methyleneadenosine-3',5'-bisphosphate. Responses to ADP were not completely inhibited by the P2Y(1) receptor antagonists. The residual response to ADP could be mediated by P2Y(13) receptors. UTP evoked an increase in [Ca(2+)](i) that was partially inhibited by suramin, suggesting that Müller cells express P2Y(2) and P2Y(4) receptors. The P2Y(6) receptor agonist UDP, and the P2Y(11) receptor agonists deoxyATP, and 3'-O-(4-benzoyl)benzoyl ATP, evoked increases in [Ca(2+)](i) in Müller cells. We conclude that isolated tiger salamander Müller cells express P2Y(1), P2Y(2), P2Y(6), P2Y(11), and possibly P2Y(4) and P2Y(13) receptors. Therefore, the physiological release of ATP, ADP, UTP, and UDP and/or their accumulation in the retina under pathological conditions could stimulate increases in [Ca(2+)](i) in Müller cells.

Expression of Axwnt-8 and Axszl in the urodele, axolotl: comparison with Xenopus.

In both the urodele axolotl and the anuran Xenopus, Wnt-8 is expressed in posterior lateral plate mesoderm (LPM) in neurula and tailbud stages. In contrast to Xenopus, expression in axolotl is more prominent in gastrula endoderm, is not initiated in mesoderm until late gastrulation, and is present in the tailbud and in the brain at tailbud stages. Sizzled is expressed in axolotl in the ventral region, similar to its pattern in Xenopus. In axolotl, the Wnt-8-expressing LPM remains relatively dorsal through tailbud stages, while ventral blood island (VBI) markers appear in a wide ventral arc.

Differential modulation of rod and cone calcium currents in tiger salamander retina by D2 dopamine receptors and cAMP.

Synaptic transmission from vertebrate photoreceptors involves activation of L-type calcium currents (ICa). Dopamine is an important circadian neuromodulator in the retina and photoreceptors possess D2 dopamine receptors. We examined modulation of ICa by dopamine and cAMP in retinal slices and isolated cells of larval tiger salamander. Results show that dopamine and a D2 agonist, quinpirole, enhanced ICa in rods and red-, blue- and UV-sensitive small single cones but inhibited ICa in red-sensitive large single cones. A D1 agonist, SKF-38393, was without effect. Quinpirole effects were blocked by pertussis toxin (PTx) pretreatment indicating involvement of PTx-sensitive G-proteins. Like dopamine, inhibition of cAMP-dependent protein kinase (PKA) by Rp-cAMPS enhanced ICa in rods and small single cones, but inhibited ICa in large single cones. In contrast, forskolin and Sp-cAMPS, which stimulate PKA, inhibited ICa in rods and small single cones but enhanced ICa in large single cones. Sp-cAMPS also occluded effects of quinpirole. These results suggest that D2 receptors modulate ICa via inhibition of cAMP. Differences among the responses of photoreceptors to cAMP are consistent with the possibility that small single cones and rods may possess different Ca2+ channel subtypes than large single cones. The results with dopamine and quinpirole showing inhibition of ICa in large single cones and enhancement of rod ICa were unexpected because previous studies have shown that dopamine suppresses rod inputs and enhances cone inputs into second-order neurons. The present results therefore indicate that the dopaminergic enhancement of cone inputs does not arise from modulation of photoreceptor ICa.

Histologia de Ixodidae: I. Ovário e traquéia de Amblymma tigrinum (Acari) / Histology of Ixodidae: I. Ovary and trachea of Amblyomma tigrinum (Acari)

I was studied the histology of Amblyomma tigrinum (Ixodidae) an ixodid fick. Three A. tigrinum engorged females were dissected, collected ovary and tracheae, and fixed in Bouin's aqueous fixative during 24 hours; the organs were cut at a thickness of 6-7 um and stained by hematoxylin-eosin. Ovary of A. tigrinum (Fig. 1) showed two cellular layers, similar to the ovary of D. andersoni and B. microplus, being one outer (oocyts) and the inner layer (epithelial cells). Trachea (Fig. 2) showed a cellular layer and a chitions spiral, similar to D. andersoni

Spermathecal cytology of Ambystoma opacum (Amphibia: Ambystomatidae) and the phylogeny of sperm storage organs in female salamanders.

Sperm storage glands, spermathecae, were examined from mated female Ambystoma opacum during the breeding season. No differences occur in the spermathecal ultrastructure of individuals sacrificed prior to oviposition and those sacrificed within 3 days of removal from tended clutches of recently oviposited eggs. The simple tubuloalveolar glands produce two types of secretory vacuoles. Apical secretory vacuoles contain glycosaminoglycans for export into the lumen to bathe stored sperm, perhaps providing the chemical/osmotic environment necessary for sperm quiescence. The other type of secretory vacuole contains an unsaturated lipid that is produced for export into the connective tissue surrounding the spermathecae. The role of this secretion may involve the contraction of myoepithelial cells, resulting in sperm expulsion. Some sperm undergo degradation in the spermathecal epithelium, and an interepithelial leukocyte was observed in one specimen. Apical secretory vacuoles and sperm are absent from the spermathecae of a specimen sacrificed 62 days after removal from a tended egg clutch. This is the first report on the spermathecal cytology of a salamander from the Ambystomatidase, and comparisons with salamanders from other families provide a morphological basis for considering spermathecae polyphyletic within the Caudata.

Peripheral projections of nervus terminalis LHRH-containing neurons in the tiger salamander, Ambystoma tigrinum.

The peripheral projections of the nervus terminalis (NT) have been difficult to examine due to the weak immunoreactivity of the processes to various antibodies. We performed two experimental manipulations in the tiger salamander in an attempt to increase the luteinizing hormone-releasing hormone-immunoreactive (LHRH-ir) labelling in the peripheral processes of the NT:1) the NT was sectioned centrally, or 2) a 100 mg melatonin pellet was embedded subcutaneously for 3 days prior to sacrifice. Following these manipulations, animals were sacrificed and tissue was processed with standard immunocytochemical techniques for the analysis of the distribution of LHRH-ir processes. In the nasal cavity, LHRH-ir fibers were observed projecting 1) into the rostral olfactory epithelium, 2) to Bowman's glands in the lamina propria of the rostromedial olfactory mucosa and ventrolateral mucosa between the main nasal cavity and Jacobson's organ, 3) into the naris constrictor muscle, and 4) along the palatine nerves and ganglia. These lesion and hormone manipulations have enabled the detection of peripheral projections of the NT not observed previously with immunocytochemical procedures alone. The wide distribution of LHRH-ir NT processes in the nasal cavity and cranium suggests that this nerve may influence many different cranial structures during appropriate pheromonal or neuroendocrine events.

Immunocytochemical localization of a galanin-like peptidergic system in the brain of two urodele and two anuran species (Amphibia).

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.

Neuroepithelial endocrine cells in the lung of Ambystoma mexicanum.

Neuroepithelial endocrine (NEE) cells were for the first time identified in the lung of the entirely aquatic urodele, Ambystoma mexicanum, by using light and electron microscopy, histochemistry, and immunocytochemistry. In the basal part of the ciliated epithelium and, less often, in the respiratory portion of the lung, NEE cells were found to occur both solitarily and in small clusters. No typical neuroepithelial bodies could be found. Using the method of Fernandez Pascual, some NEE cells were found to be argyrophilic. Microspectrofluorimetric analysis of formaldehyde-induced fluorescence and immunocytochemistry revealed the presence of 5-hydroxytryptamine. With antibodies to neuron-specific enolase only a few NEE cells exhibited a faint immunostaining. Electron-microscopically, the NEE cells are provided with distinctive cytoplasmic membrane-bound dense granules of variable size, which gave a positive argentaffin reaction. The images of emiocytotic granule release are indicative of a secretory function. In the tracheal epithelium. NEE cells seem to occur only solitarily. They bear the same ultrastructural characteristics as the intrapulmonary NEE cells but here, the dense granules are larger and associated with numerous bundles of microfilaments. Intraepithelial nerve endings were observed near the airway lumen. Between nerve terminals and NEE cells, synaptic complexes with aggregations of clear-centered vesicles close to the presynaptic membrane thickenings were observed. In addition, some nerve endings from "reciprocal synapses" with NEE cells. A receptosecretory function for NEE cells in the lung of A. mexicanum is supposed.

Immunocytochemical localization and spatial relation to the adenohypophysis of a somatostatin-like and a corticotropin-releasing factor-like peptide in the brain of four amphibian species.

The distribution of somatostatin (SRIF) - and corticotropin-releasing factor (CRF)-like - immunoreactive material was studied in the brain of four amphibian species (Ambystoma mexicanum, Pleurodeles waltlii, Xenopus laevis, Rana ridibunda) by use of immunocytochemistry. A wide network of SRIF-immunoreactive fibers and numerous perikarya were observed in all amphibians examined, with a dense accumulation of nerve endings in the external layer of the median eminence (ELME). In the representatives of the four amphibian species the CRF-like system was more circumscribed. Immunoreactive perikarya were present in the preoptic area, mainly in a ventrobasal position, and in the interpeduncular nucleus. The tract running along the ventral part of the tuber cinereum ends in the ELME facing the rostroventral lobe of the pars distalis that contains corticotrophs. CRF fibers were scarce or absent in the neural lobe. In all species studied in the present work, CRF fibers end in the area of the ELME close to the pituitary lobe containing corticotrophs. This correlation is similar to that reported for the Japanese quail and several teleosts.

Amphibian terminal nerve: distribution revealed by LHRH and AChE markers.

Immunocytochemical and histochemical studies in the tiger salamander and bullfrog demonstrated the presence of luteinizing hormone-releasing hormone-like immunoreactive (LHRH-ir) material and acetylcholinesterase (AChE) in the terminal nerve (TN). Immunoreactive perikarya and processes were found within the olfactory, vomeronasal and trigeminal nerves and in the nasal epithelium. Central TN projections consisted of fibers terminating in the olfactory bulb and bundles that projected to another group of LHRH-ir perikarya in the preoptic region. Up to 4 weeks following hypophysectomy, the labeling intensity and number of TN-immunoreactive neurons were not altered. Acetylcholinesterase histochemistry in the salamander revealed two distinct groups of neurons associated with the TN: a lightly labeled group of fusiform perikarya was located in the olfactory nerve proper and a more heavily labeled group of larger oval perikarya was found within AChE-positive trigeminal fascicles in the ventral mucosa. This study has demonstrated that the amphibian TN follows olfactory, vomeronasal and trigeminal nerves to reach peripheral targets in the nasal mucosa. The projection of TN fibers to discrete olfactory bulb glomeruli, especially evident in the bullfrog, suggests that the TN functions in odor processing. The TN projection to the preoptic region in both of these amphibians implicates the TN in reproductive processes.

A scanning electron microscopy and histological study on the effects of the mutant eyeless (e/e) gene upon the hypothalamus in the Mexican axolotl Ambystoma mexicanum Shaw.

A scanning electron microscopy, histological, and immunochemical investigation examined the effects of the mutant gene (e) upon hypothalamic development in the Mexican axolotl. The adult eyeless mutant is sterile. Previous studies indicated that this reproductive defect was due to the mutation's effect upon the hypothalamus. The present study demonstrated the pleiotropic effects of the eyeless gene upon development of the hypothalamus. Scanning electron microscopy studies looked at the early ontogeny of the hypothalamohypophyseal system. The major morphological difference observed in the hypothalamus of normals compared to eyeless mutants was the reduced nature or complete lack of a preoptic recess in eyeless mutants. Early embryonic tissue movements also differed when normal siblings were compared to eyeless mutant axolotls. Histological examination looking for paraldehyde-fuchsin-positive secretory neurons revealed a paired nucleus preopticus in both normals and eyeless mutants, but this region lacked the emanating paraldehyde-fuchsin-positive fiber tracts in eyeless mutants. The neurohypophysis of the eyeless mutants was atrophied and contained far less paraldehyde-fuchsin-positive material when compared to normal axolotls. Immunochemical studies were done to look at the distribution of immunoreactive luteinizing-hormone-releasing hormone (ir-LHRH) in brains of eyed and eyeless mutant axolotls of different stages. This study detected deficiencies in ir-LHRH in the anterior hypothalamus of eyeless mutants. In general in the eyeless mutant axolotl, the observed anterior hypothalamic deficiencies are comparable to those observed in anurans which have had their optic vesicles removed. These observations suggest a possible utility of the eyeless mutant axolotl for studies concerned with endocrine development in the absence of hypothalamic modulation.

The pattern of lateral-line afferents in urodeles. A horseradish-peroxidase study.

The organization of posterior and anterior afferents of the lateral-line system was studied in several species of urodeles by means of transganglionic transport of horseradish peroxidase. The afferents of each lateral-line nerve form distinct fascicles in the medullary alar plate. Each of the two branches of the anterior lateral-line nerve is organized in two long and one short fascicles. The posterior lateral-line afferents form only two long fascicles. Each ordinary neuromast is supplied by only two afferents, which run in the two ventral medullary fiber bundles. It is suggested that afferents to hair cells displaying one type of polarity form together one bundle, but those contacting hair cells polarized in the opposite way form the second ventral bundle of one lateral-line branch. Thus, the lateral-line afferents may be organized in a directotopic fashion. The short dorsal fascicle formed only by the anterior lateral-line afferents receives fibers exclusively from small pit organs. Each pit organ is supplied by only one afferent. Anatomically, these pit organs resemble in many respects the electroreceptive ampullary organs of certain fish. Neurons labeled retrogradely via the anterior lateral-line nerve afferents have been attributed to the nervus trigeminus or facialis. In addition to the posterior lateral-line afferents, only few centrifugally projecting neurons were labeled. These neurons are discussed as efferents to the posterior lateral-line neuromasts.

Ultrastructure of lateral line organs in aneurogenic amphibian larvae (Ambystoma).

An ultrastructural study was performed to determine the ability of presumptive lateral line tissue in A. maculatum embryos to migrate and differentiate in the absence of all sensory innervation. In experimental animals, all presumptive neural tissue was surgically removed at Harrison's stage 24, leaving only the posterior portion of the post-auditory placode. At later stages, completely differentiated aneural lateral line organs were found in the midbody region of the larvae. These were morphologically similar to those of control larvae. No signs of atrophy or degeneration of the neuromasts were seen even after several weeks of nerve-free conditions.

Apical secretion from taste bud and other epithelial cells in amphibians.

Taste buds of the axolotl, Ambystoma mexicanum, contain cells, previously undescribed in this species, which have a long apical process, and are similar to the Type III cells of mammalian taste buds, and to the gustatory cells in fish. In the supporting cells, there is evidence of periodic decapitation, in addition to secretion by exocytosis. Bilaminar fragments, which are leaf-shaped bodies formed of two dense laminae separated by a lucent gap, protrude from the apical microvilli of the supporting cells and are found detached in the extracellular secreted layer. Their form and dimensions suggest that they represent secreted lipo-protein material. Similar bilaminar fragments have been seen, in much smaller numbers, on some other epithelial cells in amphibians, and in fish. A unique case, in which rough endoplasmic reticulum was found in the extracellular layer of the axolotl oral epithelium, is reported; it had apparently been ejected from the cell. It is suggested that the axolotl produces a copious secretion at the taste bud pore, in order to wash the surface, and that the bilaminar fragments represent material aiding this cleansing process. The condition in the axolotl is compared with that in some other species, particularly Rana temporaria.