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PubMed1993

Ultrastructure of the glomus cells in the carotid body of chronically hypoxic rats: with special reference to the similarity of amphibian glomus cells.

The ultrastructural characteristics of the glomus cells in the rat carotid body exposed to extremely long-term hypoxia (10-12 weeks) were investigated. The glomus cells could be classified into four distinct types according to the shape of dense-cored vesicles in the glomus cell cytoplasm: 1) small vesicle cells (SVCs, 50 nm in mean diameter), 2) large vesicle cells (LVCs, 80 nm in mean diameter), 3) dilated eccentric vesicle cells (EVCs, 400-800 nm in diameter), and 4) mixed vesicle cells (MVCs, large and eccentric vesicles). Many clusters of glomus cells were found to contain all four categories of cell types. The appearance of EVCs was a unique and common characteristic of glomus cells in this long-term hypoxia model. We also noted other ultrastructural features with chronic hypoxia which are characteristic of the amphibian carotid labyrinth glomus cells: 1) incomplete covering of glomus cells with the supporting cell missing over a wide area, 2) long thin cytoplasmic projections in the intervascular stroma, and 3) intimate apposition of the glomus cells and pericytes (g-p connection), endothelial cells (g-e connection), plasma cells, and fibrocytes. Because arterial PO2 is generally low in amphibia, these may be general features of hypoxic adaptation and facilitate both uptake of oxygen from blood and release of catecholamine into the blood. The g-p and g-e connections may take part in the regulation of the microcirculation in the enlarged carotid body.

Amphibians↗
PubMed2001

New methods for the isolation of skeletal muscle sarcolemma and sarcoplasmic reticulum allowing a comparison between the mammalian and amphibian beta(2)-adrenergic receptors and calcium pumps.

New methods were established for the rapid and simultaneous isolation of multiple sarcolemmal and sarcoplasmic reticular fractions from very small amounts (0.25-2.0 g) of skeletal muscle. Thebeta(2)-adrenergic receptor and calcium transport systems were used as indices of purity and functional integrity as well as being the focal points of the study. These methods were found to be suitable for the special needs of small tissue samples, allowed rapid preparation and were appropriate for skeletal muscle from various species, frogs to mammals. The sarcolemmalbeta(2)-adrenergic receptor was expressed in frogs and mammals at similar levels of expression (336-454 fmol. x mg(-1)). The calcium pump was also present in sarcolemmal and sarcoplasmic reticular fractions in all species but notable species differences were found. In sarcolemmal fractions, while calcium binding was uniformly low (<1 nmol. x mg(-1)), oxalate stimulation was variable: low in frogs ( approximately 1.05-fold) high in mammals (120-450-fold). In sarcoplasmic reticular fractions, calcium binding was low in frogs (4-9 nmol. x mg(-1)) and much higher in mammals (322-383 nmol. x mg(-1)); oxalate stimulated calcium transport to a much greater extent in frogs (<70-fold) than in mammals (1.6-2-fold). It is concluded that thebeta(2)-adrenergic receptor appears to be strongly conserved in skeletal muscle while the use of calcium pumps evolves from reliance in Amphibia on the sarcoplasmic reticular calcium pump to the use in Mammalia of calcium pumps from both the sarcoplasmic reticulum and the plasma membrane.

Adenosine Triphosphatases↗
PubMed2001

Identification of a nonmammalian Golf subtype: functional role in olfactory signaling of airborne odorants in Xenopus laevis.

Attempts to identify the Galpha subtypes in the two compartments of the olfactory system from Xenopus, which are supposed to be specialized for detecting aquatic and volatile odorous compounds, revealed that a Galpha(o1) subtype is characteristic for the "water nose," the lateral diverticulum, whereas a novel Galpha(s) subtype predominates in the "air nose," the medial diverticulum. The newly identified Galpha(s)-type is more closely related to Galpha(olf) of rat and human than to the known Galpha(s)-isoform of Xenopus; it is therefore considered the first identified nonmammalian Galpha(olf) subtype. Sequence comparison of Galpha(olf) from amphibia and mammals revealed a particular conservation within the alpha-helical domains, which are supposed to control the GDP/GTP-exchange rate. The selective expression of different Galpha subtypes in the two anatomically separated and functionally specialized nasal compartments parallels the expression of distinct classes of olfactory receptors. Moreover, biochemical analysis revealed that stimulation with appropriate odorous compounds elicits the formation of inositol trisphosphate in the lateral diverticulum. In contrast, cyclic adenosine monophosphate signals were induced in the medial diverticulum, and this response appears to be mediated by the novel Galpha(olf) subtype. The data indicate that olfactory sensory neurons in each of the nasal cavities are equipped not only with defined sets of receptor types but also with a distinct molecular machinery for the chemo-electrical transduction process.

Amino Acid Sequence↗
PubMed1978

Golgi-staining of "primary" and "secondary" motoneurons in the developing spinal cord of an amphibian.

The Golgi technique was used to study the morphology of spinal motoneurons at various stages in the early development of swimming behaviour in embryos and larvae of the palmate newt, Triturus helveticus ((Razoumowsky). The earliest motoneurons stained appeared to be associated with the Mauthner-cell system. The overall morphology of these "primary" motoneurons seems to be similar throughout the lower vertebrates and the distinctive characteristics found in earlier descriptions of those from caudate amphibia were probably due to misinterpretation. At about the time of hatching and development of low-frequency swimming behaviour, other motoneurons were found to innervate the axial musculature, cells with a central morphology different from those of the "primary" type. It was found likely that these "secondary" motoneurons innervate a separate muscle system concerned with tonic and "slow phasic" activity, while "fast phasic" acitivity in rapid swimming is supplied by "primary" cells.

Animals↗
PubMed1980

Phylogeny of enteric serotonergic neurons.

Serotonergic neurons have previously been identified in the enteric nervous systems of humans, subhuman primates, rodents, and rabbits. The distribution of enteric serotonergic neurons in lower vertebrates was examined in order to determine if these neurons are restricted to mammals and, if they are found more generally amongst vertebrates, when they first appear in vertebrate phylogeny. Since mammalian enteric serotonergic neurons take up 3H-serotonin by a highly specific mechanism, the radioautographic demonstration of axonal uptake of 3H-serotonin was used as the primary tool in looking for these neurons. As controls, conditions known to interfere with 3H-serotonin uptake by mammalian enteric neurons were also examined. These controls included incubation with 10 microM fluoxetine, a specific antagonist, incubation in Na+-free medium, and incubation in the presence of a 100-fold excess of nonradioactive serotonin. Radio-autographic labeling had to be absent or greatly reduced under all three control conditions for labeling by 3H-serotonin to be considered specific. Labeled enteric axons were found in cyclostomes (hagfish), teleosts (goldfish), and amphibia (bull-frog) but not in tunicates (sea squirt; sea vase) or echinoderms (sea cucumber). In addition, the serotonin concentration was measured in the intestine of two vertebrates, hagfish and goldfish, that do not have serotonin-containing enterochromaffin cells. Serotonin was found in both; in hagfish, the amine concentration was highest in preparations of muscularis externa containing the myenteric plexus. It is concluded that enteric serotonergic neurons arose early in vertebrate evolution, possibly in an ancestral chordate resembling amphioxus, although probably not in more primitive prevertebrates, and that they are a general feature of the vertebrate bowel.

Animals↗
PubMed1982

Quantitative analyses of neuronal development in the lateral motor column of mouse spinal cord. III. Generation and settling patterns of large and small neurons.

The generation and settling patterns of large and small lateral motor column (LMC) neurons were compared in the spinal cords of three inbred strains of mice by means of tritiated thymidine autoradiography. No significant strain differences were observed for the number of large LMC cells (presumptive alpha motor neurons) that were heavily labeled on each injection day, although there were significant strain variations for this measure with regard to small LMC neurons (presumed gamma motor neurons and interneurons). The generation of both large and small LMC neurons began at the same time, but peak production of large cells preceded that of the small neurons. There were no strain differences observed for this relationship between the large and small cells. These findings indicate that the LMC, from the time of its initial formation, contains cells destined to become large and small neurons. The positions of large and small neurons within the adult LMC relative to their times of origin (settling patterns) were analyzed statistically. A significant ventrodorsal sequence for early-to-late generated cells was observed for both large and small LMC neurons. No significant strain differences were found in the analysis of settling patterns. A ventrodorsal settling pattern also has been described for amphibia (Prestige, '73) and, in conjunction with the proximodistal sequence of limb development described by other investigators, the ventrodorsal sequence could play a key role in the development of motor neuronal somatotopic organization.

Animals↗
PubMed1984

Changes in the topographically organized connections between the nucleus isthmi and the optic tectum after partial tectal ablation in adult goldfish.

The projection of the nucleus isthmi to the ipsilateral optic tectum was examined in normal goldfish. This was compared to the projection in animals in which the entire visual field had been induced to compress onto a rostral half tectum by caudal tectal ablation. The isthmo-tectal projection was examined by making localized injections of horseradish peroxidase into the optic tecta and observing the patterns of labeled cells within the nucleus isthmi. The teleost nucleus isthmi consists of a cell sparse medulla covered by a cellular cortex, which is thick on the rostral, medial, and dorsal surfaces of the nucleus. Almost all isthmic cells projecting to the tectum were located in the area of thick cortex. In normal fish, rostral tectal injections labeled cells in the rostroventral portion of the thick cortex; injections midway in the rostrocaudal tectal axis labeled more caudodorsally located cells, and caudal tectal injections labeled cells a little further caudally in extreme dorsal cortex. The rostroventral to caudodorsal isthmic axis was therefore seen to project rostrocaudally along the tectum. This topography contrasts somewhat with the situation seen in amphibia where the rostrocaudal tectal axis receives projections from the rostrocaudal isthmic axis. In fish with half-tectal ablations, injections near the caudal edge of the half tectum (at a site that had originally been midtectal) labeled cells that had previously projected to caudal tectum. Rostral tectal injections in fish with compression of the visual field gave a normal pattern of labeled isthmic cells. The results indicate that a topographically ordered isthmo-tectal projection exists in goldfish that may be induced to compress onto a half tectum.

Animals↗
PubMed1985

The projection of three extrathalamic cell groups to the cerebral cortex of the turtle Pseudemys.

Three extrathalamic subcortical inputs to the part of the cerebral cortex that is known to receive thalamic fibers in the turtle were examined in the present study. Direct projections from the locus coeruleus, the superior medial raphe nucleus, and a wide area of the basal telencephalon that lies ventromedial to the globus pallidus were demonstrated with the horseradish peroxidase method. Fluorescence histochemistry confirmed the presence of catecholamine-containing fibers in the rostral half of dorsal cortex and also demonstrated a dense network of serotoninergic fibers. Biochemical analysis showed the concentration of both monoamines to be relatively high; the norepinephrine concentration was 709 ng/g and the serotonin concentration was 1,750 ng/g. No evidence was found to suggest the existence of either a dopamine fiber projection to cortex comparable to that of mammalian neocortex or the presence of an epinephrine pathway to turtle cortex equivalent to the epinephrine-containing fibers in the pallium of amphibians. The coexistence of the projections from the thalamus with noradrenergic projections from the locus coeruleus, serotoninergic projections from the superior medial raphe nucleus, and presumably cholinergic projections from the basal telencephalon provide at least four distinct subcortical inputs to the reptilian dorsal cortex. Neither thalamic nor similar extrathalamic inputs have been demonstrated in the dorsal pallium of amphibia. Mammalian neocortex, in contrast, has even more elaborately differentiated inputs of both types. These results support the idea that thalamic and extrathalamic inputs to cortex appear at the same time in vertebrate evolution, and that both types of inputs are required for the normal development and function of neocortex.

Animals↗
PubMed1987

Effect of tetraploidy on dendritic branching in neurons and glial cells of the frog, Xenopus laevis.

Morphological aspects of four different groups of Golgi impregnated brain cells from a tetraploid strain of Xenopus laevis frogs were compared to analogous cells in comparably sized diploid frogs. The cells examined included neurons from the telencephalon, caudal hypothalamus, and optic tectum, and radial glial cells from the optic tectum. The brains of tetraploid frogs appeared grossly normal and were the same size and contained similar cell types as diploid brains. As observed in previous studies on polyploid amphibia, somal diameters increased significantly in tetraploid cells for each of the four groups of cells examined. Also, the total length of the dendritic arbors in tetraploid brain cells increased significantly by factors ranging from 1.4 to 2.4 times the total length of the analogous processes in diploid cells. Tetraploid neurons in the telencephalon and hypothalamus increased their arbor lengths predominantly by increasing the number of dendritic branches, while maintaining the average distance between branch points in the dendritic segments. In contrast, the tetraploid large pear-shaped neurons in the optic tectum had significantly longer terminal dendritic segments than the analogous diploid neurons, although these tetraploid neurons maintained their average number of dendritic segments per cell. Tetraploid tectal radial glial cells appeared to increase both their number of branches and the lengths of their terminal segments. Thus, the mode by which tetraploid brain cells achieved longer dendritic arbors varied from cell type to cell type. These results suggest a hypothetical basis for possible effects of genomic size on vertebrate brain structure and evolution at the cellular level.

Animals↗
PubMed1987

Development of substance P-like immunoreactivity in Xenopus embryos.

The development of substance P-like immunoreactivity (SPLI) was studied in the Xenopus embryonic nervous system in order to determine in which neuronal populations and at what developmental times SPLI is expressed. Although Rohon-Beard neurons initially were thought to be the only substance P-immunoreactive cells in the embryonic frog spinal cord, we have demonstrated that several neuronal phenotypes are immunoreactive. The earliest evidence of SPLI was seen at stage 28 (Nieuwkoop and Faber, '67), at which time only some trigeminal ganglion cells, their axons in the ophthalmic nerve, and axons in the lateral tracts of the hindbrain showed SPLI. In the embryonic brain at stages 29/30, 37/38, and 42, SPLI was seen in the hypothalamus, trigeminal ganglion cells and their peripheral axons, the sensory roots of cranial nerve IX/X, and axons in the hindbrain lateral tracts. At premetamorphic stages, SPLI was found in several populations that are immunoreactive in adult amphibia. In the embryonic spinal cord, Rohon-Beard neurons were labeled consistently with reaction product; there was a rostrocaudal time gradient of immunoreactivity with increasing development. The Rohon-Beard neurons were not immunoreactive at developmental stages in which axonal outgrowth was beginning (stage 21), but were strongly immunoreactive at stages in which target cells had been contacted (stage 29). Several types of interneurons in the spinal cord (as classified by Roberts and Clarke, '82) showed SPLI during embryonic stages. At premetamorphic stages the Rohon-Beard neurons began to disappear and the immunoreactive interneurons were distributed similarly to those reported in the adult. Dorsal root ganglia differentiated during these stages, and at this time some of the neurons belonging to these ganglia exhibited substance P-like immunoreactivity.

Animals↗
PubMed1989

Immunocytochemical reactivity of Xenopus laevis retinal rods and cones with several monoclonal antibodies to visual pigments.

Immunocytochemical reactions with several antibodies to visual pigments were used to study visual cells of the Xenopus laevis retina. Monoclonal antibodies to bovine opsin "E," 1D4, and 4B4 (reactive with the N- and C-terminus and with the loop connecting transmembrane segments 5-6, respectively) and to chicken visual pigments COS-1 and OS-2 (binding to mammalian red/green and blue cones, respectively), as well as a rabbit antifrog opsin serum 11-7, were applied to semithin and thin sections of the retina. The bound antibodies were detected with the peroxidase technique at the light microscopic level; a three-stage immunogold procedure was used for electron microscopic immunocytochemistry. The overwhelming majority of rods were labeled by monoclonal antibodies "E," 4B4, 1D4, OS-2, and serum 11-7. A small fraction (2-3%) of rods did not bind monoclonal antibodies "E" and 4B4, but this minor population of rods was strongly reactive with 1D4 and to a lesser extent with OS-2, indicating the presence of different visual pigment. These rods differ in shape from the major rod type; they are thinner, shorter, and may be comparable to the blue-sensitive ("green") rods of other amphibia. Cones were morphologically heterogeneous: double cones, large single cones, and small single cones were found, and the large single and the double cones were occasionally duplicated. Double cones and large single cones (as well as their duplicated varieties) strongly bound monoclonal antibodies COS-1 and were unlabeled by all other monoclonal antibodies, except OS-2. The small single cone was remarkably unreactive with COS-1 and "E," weakly labeled by 1D4 and 4B4, and most reactive with OS-2 and 11-7. This unique pattern of immunocytochemical reactions in the small cone type indicates the uniqueness of its visual pigment from other cone types in the Xenopus retina. The present study shows the existence of two different opsins in morphologically distinct (thick and thin) rod types and at least two cone pigments in the heterogeneous cone population.

Animals↗
PubMed1990

Organization of projections from olfactory epithelium to olfactory bulb in the frog, Rana pipiens.

One hypothesis for the coding of olfactory quality is that regions of the olfactory epithelium are differentially sensitive to particular odor qualities and that this regional sensitivity is conveyed to the olfactory bulb in a topographic manner by the olfactory nerve. A corollary to this hypothesis is that there is a sufficiently orderly connection between the epithelium and the olfactory bulb to convey this topographical coding. Thus we examined topography in the projection from epithelium to bulb in the frog, which has been the subject of numerous electrophysiological studies but has not yet been examined using modern neuroanatomical techniques. The tracer WGA-HRP was applied to the ventral or to the dorsal olfactory epithelium, or both. Anterograde transport of label to the olfactory bulb was seen after as few as 2 days; label was still present in the bulb as long as 21 days postinjection. In cases where WGA-HRP was applied to the entire epithelium, there was dense anterograde labelling of the ipsilateral olfactory bulb. In addition, a small medial portion of the contralateral bulb was labelled. Injections limited to either the ventral or dorsal epithelium produced patterns of anterograde labelling in the glomerular layer of the olfactory bulb, which varied with the size and location of the injection. With very large injections in either the dorsal or ventral epithelium, label appeared to be evenly distributed in the glomerular layer. With smaller injections in the ventral epithelium, there was heavier labelling in the lateral than in the medial portions of the glomerular layer, although light labelling was found in all regions of the glomerular layer. In contrast, injection sites restricted to the dorsal epithelium produced more anterograde labelling in the medial than lateral portions of the glomerular layer. These patterns extended throughout the dorsal-ventral extent of the bulb. Within the limits of the anterograde tracing technique used, we were unable to detect any systematic relationship between the pattern of labelling in the glomerular layer and the medial-lateral or rostral-caudal location of the injection site in either the ventral or dorsal epithelium. We conclude that in the frog, as in other amphibia, there is only a limited degree of topographic order between the epithelium and the olfactory bulb.

Animals↗
PubMed1991

Organization of the cerebellum in the pigeon (Columba livia): II. Projections of the cerebellar nuclei.

The projections of the deep cerebellar nuclei in the pigeon have been delineated using autoradiographic and histochemical (WGA-HRP) tracing techniques. A medial (CbM) and lateral (CbL) cerebellar nucleus are recognized and CbM may be further partitioned into internal, intermediate, and intercalate divisions. As in mammals, most extracerebellar projections of CbM travel in the fasciculus uncinatus (FU); the rest travel with those of CbL in the brachium conjunctivum (BC). In the pigeon, both of these pathways are bilaterally but primarily contralaterally projecting systems. FU is a predominantly descending tract, with terminations within (1) the vestibular complex, (2) a column of contiguous medial reticular nuclei from pontine to caudal medullary levels; (3) the plexus of Horsley portion of the parvicellular reticular formation, continuing through the nucleus centralis medullae oblongatae, pars dorsalis, into intermediate layer VII of the cervical spinal cord, down to cervical segment 8-9; (4) the lateral reticular nucleus and the paragigantocellular reticular nucleus; (5) the dorsal lamella of the inferior olive. Rostrally FU terminals are found in the locus ceruleus and dorsal subcerulean nucleus. Minimal FU projections are also seen to the motor trigeminal nucleus and the subnucleus oralis of the descending trigeminal system. A small projection from the intercalate division of CbM travels in BC and projects upon the midbrain central grey, the intercollicular nucleus, the lateral tectal periventricular grey, the stratum cellulare externum and, sparsely, upon the dorsolateral thalamus. The bulk of BC originates from the lateral cerebellar nucleus and consists of a massive ascending and a small descending branch. The ascending system projects upon the red nucleus and the dorsally adjacent interstitial nucleus of Cajal and midbrain central grey, the prerubral fields continuing into the stratum cellulare externum, the nucleus intercalatus thalami, the ventrolateral thalamic nucleus, the medial spiriform nucleus, the nucleus principalis precommissuralis, the nucleus of the basal optic root, the nucleus geniculatus lateralis pars ventralis, the dorsolateral thalamus, including the dorsal intermediate posterior, and the dorsolateral intermediate and anterior nuclei. BC also contains axons from the infracerebellar nucleus, which projects upon the trochlear and the oculomotor nuclei. The descending branch of BC distributes to the papilioform nucleus, the medial pontine nucleus, the gigantocellular and paramedian reticular nuclei, and, minimally, the rostral portions of the medial column and ventral lamella of the inferior olive. Taken in conjunction with data on amphibia and reptiles the present findings suggest that the fundamental ground plan of vertebrate cerebellar organization involves a medial and lateral cerebellar nucleus.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1994

Distribution and morphology of sacral spinal cord neurons innervating pelvic structures in Xenopus laevis.

Relatively little is known about the organization of neural input to pelvic viscera in amphibia. In this study, sacral spinal efferent neurons were labeled in Xenopus laevis frogs by application of horseradish peroxidase (HRP) to the tenth spinal nerve, to pelvic musculature, or to the pelvic nerve. DiI was applied to the pelvic nerve with similar results. Labeled spinal neurons were located in the intermediate gray or in the ventral horn. Neurons in the tenth dorsal root ganglion, but not in the spinal cord, were labeled after application of HRP or DiI to the pudendal nerve. The labeled neurons in the spinal cord intermediate gray were in a position comparable to that of the mammalian sacral parasympathetic nucleus (SPN). Two apparent subdivisions included 1) a medial cluster of cells with mediolaterally oriented dendrites and 2) a lateral group with dorsoventrally oriented dendrites. An intermediate group, not clearly classed with the other two, was also identifiable. In some cases, labeled tenth nerve primary afferents were seen in contact with efferent neurons of the intermediate gray. Labeled neurons in the ventral horn medial to the lateral motor column were small, with dendrites oriented mediolaterally, in a position comparable to that of the mammalian Onuf's nucleus. The peripheral targets of DiI-labeled pelvic nerve axons were the compressor cloaca muscle, cloaca, and bladder. DiI-labeled pudendal nerve axons distributed peripherally to cloacal lip and medial thigh integument. These data suggest that the pudendal nerve in amphibians is purely sensory and that both somatic and autonomic motor axons traverse the pelvic nerve.

Animals↗
PubMed1995

Substance P, bombesin, and leucine-enkephalin immunoreactivities are restored in the frog tectum after optic nerve regeneration.

Extensive regeneration of the optic nerve takes place in adult Amphibia. In this study, we have determined whether one aspect of retinotectal organisation, namely immunoreactive laminae in the retinorecipient layers of the optic tectum, is restored after optic nerve regeneration. To do so, the distributions of substance-P, bombesin, and leucine-enkephalin immunoreactivities were examined in the optic tectum of the frog Litoria (Hyla) moorei. Results of a normal series were compared with those at intervals up to 84 days and at 196 days after either unilateral deafferentation or optic nerve crush. In the normal series, distinct neuropeptide immunoreactive laminae were located within the retinorecipient tectal layers. There were two major laminae with substance-P, two with bombesin, and one with leucine-enkephalin immunoreactivities. Additional faint laminae of both substance-P and bombesin immunoreactivity were present in the tectal region that receives input from the visual streak. In addition, labelling of cell bodies and dendrites was seen elsewhere in the tectum. All except one immunoreactive lamina changed after deafferentation. The deeper of those with substance-P immunoreactivity, along with both bombesin laminae, were eventually lost; the lamina with leucine-enkephalin immunoreactivity was halved in intensity. We assume that these laminae are wholely or, in the case of the leucine-enkephalin lamina, partially associated with primary optic input. By contrast, the more superficial lamina with substance-P immunoreactivity remained unchanged and is presumably not directly related to visual input. During nerve regeneration, the intensity of all laminae associated with optic input initially fell as in the deafferentation series but, in the long term, recovered to approximately 80% of normal intensities. We conclude that ganglion cells associated with each of the immunoreactivities tested had successfully regenerated. The reduced intensity of immunoreactivities after regeneration is due presumably in part to the cell loss from the ganglion cell population. Furthermore, we discuss the findings of similar studies for Rana pipiens (Kuljis and Karten [1983] J. Comp. Neurol. 217:239-251 and [1985] 240:1-15) in light of the present findings. We argue that some of the previous observations can be reinterpreted to indicate that regeneration was not limited to ganglion cells associated with substance-P immunoreactivity as first thought.

Animals↗
PubMed1995

Embryonic development of central nervous system myelination in a reptilian species, Eumeces fasciatus.

The myelin proteolipid proteins are a vital component of the vertebrate central nervous system (CNS), contributing essential functions to the development of the myelinating cells of the CNS and to the structure of CNS myelin. Alternative splicing of the proteolipid protein (PLP) gene to produce two related isoforms occurs in Mammalia, Aves, and Reptilia, but not Amphibia. As part of a long-term investigation into the function of the different isoforms of PLP, embryonic development, myelination, and PLP gene expression in reptilian CNS were examined. PLP gene expression was already substantial by day 19 (stage 39) of the 27-day Eumeces fasciatus egg incubation period. By day 21 of incubation, also stage 39, PLP mRNA was at peak levels; there was a significant amount of CNS myelination as demonstrated by electron microscopy of the spinal cord; and the reflexive motor response was evident. Although most axons were myelinated by the time of hatching, myelin sheaths continued to increase in size and compactness after hatching. The correlation of physiological development, CNS myelination, and expression of the PLP gene in the lizard corresponded well with the developmental pattern seen in mammals.

Animals↗
PubMed1983

Monoclonal anti-VH antibodies recognize a common VH determinant expressed on immunoglobulin heavy chains from various species.

Our previous work using rabbit antibodies to the variable region of MOPC315 myeloma heavy chain (VH) has indicated the existence of framework determinant(s) common to many murine heavy chains. Here we report the characterization of anti-VH monoclonal antibodies (mAb) prepared in an attempt to elucidate the nature of the common VH determinant. We immunized AKR/J mice with a purified VH315 fragment and generated somatic cell hybrids by the fusion of the immune AKR/J splenocytes with the NS1 myeloma cells. Thirty-seven common anti-VH and 57 subgroup VHI-specific hybridomas have been established and characterized. Whereas the anti-subgroup mAb seemed to react with a determinant unique to the MOPC315 (mouse VHI) subgroup, all the anti-VH mAb reacted with myeloma heavy chains of different VH subgroups, class and allotypes. Antibody competition studies revealed that the VH subgroup determinants are distinct from the common VH determinants and that both were also recognized by the rabbit polyclonal antibodies. The common VH determinants were found to be "hidden" determinants on intact immunoglobulin molecules being exposed only on isolated heavy chains. Furthermore, they are sequential determinants since they are preserved on fully denatured heavy chains. The common VH determinants are shared by immunoglobulins of a wide range of vertebrates from amphibia to man and thus represent antigenic structures which were highly conserved throughout evolution.

Animals↗
PubMed1987

Antibody diversity in amphibians. Noninbred axolotls used the same unique heavy chain and a limited number of light chains for their anti-2,4-dinitrophenyl antibody responses.

Noninbred axolotls (Ambystoma mexicanum, amphibia, urodela) were immunized with trinitrophenylated sheep red blood cells (TNP-SRBC) and anti-2,4-dinitrophenyl (DNP)/TNP antibodies were individually purified by affinity chromatography. The isolated IgM-like antibodies were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and isoelectric focusing (IEF) under reducing conditions. The SDS-PAGE and IEF-separated heavy (H) and light (L) chains were electroblotted onto nitrocellulose, probed with mouse monoclonal antibodies specific for H or L axolotl Ig chains and stained by a rabbit anti-mouse Ig horseradish peroxidase conjugate. The specific detection of axolotl anti-DNP/TNP H chain spectrotypes shows for each of the 14 individually analyzed samples a very similar pattern of 4-5 ordered spaced bands. This suggests that all animals express the same VH chain segment representing the germinal expression of a unique VH gene. When the same analysis was performed starting from a pool of nonimmunized axolotl sera, a low background of natural anti-DNP antibodies was detected. When analyzed by IEF, the H chains of the pooled anti-DNP natural antibodies display the same pattern of restricted heterogeneity when compared to the H chain spectrotypes of the individual immune anti-DNP/TNP antibodies. The specific detection of the axolotl anti-DNP/TNP L chain spectrotypes indicates at the individual level more heterogeneous and polymorphic patterns compared with H chains, although most animals share the majority of their bands. Our experiments indicate that in axolotl, the production of antibodies to DNP results from the germinal expression of a very limited set of V genes, already expressed as naturally occurring anti-DNP antibodies before immunization. This seriously restricts the possible extension of the antibody repertoire and perhaps even the nature of antibody "specificity" in this primitive vertebrate.

Ambystoma↗