Developmental aspect of the gonadotropin-releasing hormone system.

Gonadotropin-releasing hormone (GnRH) regulates the hypothalamo-pituitary-gonadal (HPG) axis in all vertebrates studied. GnRH neurons that regulate the HPG axis are primarily derived from progenitor cells in the nasal compartment (NC) and migrate along olfactory system derived fibers across the cribriform plate to destinations in the forebrain. Across their long and uncommon migratory route many factors are likely important for their successful development. Several classes of molecules are being studied for their potential influences on migration, including those related to cell surface interactions (membrane receptors, adhesion molecules, extracellular matrix (ECM) molecules, etc.) and those related to communication across distances (neurotransmitters, peptides, chemoattractant or repellent molecules). Of the classes of molecules associated with cell surface interactions, glycoconjugates with terminal galactose, are temporally and spatially expressed on olfactory fibers that guide GnRH neurons and may play role(s) in migration. Of the molecules associated with communication across distances, the neurotransmitter gamma-aminobutyric acid (GABA) is associated with the GnRH migration pathway and influences the position and organization of GnRH neurons in vitro and in vivo. Furthermore, galactose-containing glycoconjugates and GABA are associated with GnRH neurons in species ranging from humans to lamprey. In mice and rats, GABA is found transiently within a subpopulation of GnRH neurons as they migrate through the NC. One of the key elements in considering regulators of GnRH neuron migration is the diversity of GnRH synthesizing cells. For example, only subpopulations of GnRH neurons also contain GABA, specific GABA receptors, or select glycoconjugates. Similarly, treatments that influence GnRH neuronal migration may only affect specific subsets and not the entire population. It is likely that we will not be able to characterize the migration of all GnRH neurons by a single factor. By combining molecular inquiries with genetic models, single cell analyses, and an in vitro migration model, we are beginning to decipher one of the most critical events in the establishment of the reproductive axis.

Deleted in colorectal cancer (DCC) regulates the migration of luteinizing hormone-releasing hormone neurons to the basal forebrain.

Luteinizing hormone-releasing hormone (LHRH) neurons migrate from the vomeronasal organ (VNO) to the forebrain in all mammals studied. In mice, most LHRH neuron migration is dependent on axons that originate in the VNO but bypass the olfactory bulb and project into the basal forebrain. Thus, cues that regulate the trajectories of these vomeronasal axons are candidates for determining the destination of LHRH neurons. Using in situ hybridization techniques, we examined the expression of Deleted in colorectal cancer (DCC), a vertebrate receptor for the guidance molecule netrin-1, during development of the olfactory system. DCC is expressed by cells in the olfactory epithelium (OE) and VNO, and in cells migrating from the OE and VNO from embryonic day 11 (E11) to E14. Some DCC(+) cells on vomeronasal axons in the nose also express LHRH. However, DCC expression is downregulated beginning at E12, so few if any LHRH neurons in the forebrain also express DCC. In rat, DCC is expressed on TAG-1(+) axons that guide migrating LHRH neurons. We therefore examined LHRH neuron migration in DCC(-/-) mice and found that trajectories of the caudal vomeronasal nerve and positions of LHRH neurons are abnormal. Fewer than the normal number of LHRH neurons are found in the basal forebrain, and many LHRH neurons are displaced into the cerebral cortex of DCC(-/-) mice. These results are consistent with the idea that DCC regulates the trajectories of a subset of vomeronasal axons that guide the migration of LHRH neurons. Loss of DCC function results in the migration of many LHRH neurons to inappropriate destinations.

Patterning of olfactory sensory connections is mediated by extracellular matrix proteins in the nerve layer of the olfactory bulb.

In early rat embryos when axons from sensory neurons first contact the olfactory bulb primordium, lactosamine-containing glycans (LCG) are detected on neurons that are broadly distributed within the olfactory epithelium, but that project axons to a very restricted region of the ventromedial olfactory bulb. LCG(+) axons extend through channels defined by the coexpression of galectin-1 and beta2-laminin. These two extracellular matrix molecules are differentially expressed, along with semaphorin 3A, by subsets of ensheathing cells in the ventral nerve layer of the olfactory bulb. The overlapping expression of these molecules creates an axon-sorting domain that is capable of promoting and repelling subsets of olfactory axons. Specifically, LCG(+) axons preferentially grow into the region of the nerve layer that expresses high amounts of galectin-1, beta2-laminin, and semaphorin 3A, whereas neuropilin-1(+) axons grow in a complementary pattern, avoiding the ventral nerve layer and projecting medially and laterally. These studies suggest that initial patterning of olfactory epithelium to olfactory bulb connections is, in part, dependent on extracellular components of the embryonic nerve layer that mediate convergence and divergence of specific axon subsets.

Semaphorin 3A is required for guidance of olfactory axons in mice.

Semaphorin 3A (Sema3A) is a membrane-associated secreted protein that has chemorepulsive properties for neuropilin-1 (npn-1)- expressing axons. Although mice lacking the Sema3A protein display skeletal abnormalities and heart defects, most axonal projections in the CNS develop normally. We show here that Sema3A is expressed in the lamina propria surrounding the olfactory epithelium (OE) and by ensheathing cells in the nerve layer of the ventral olfactory bulb (OB) throughout development. Subsets of sensory neurons expressing npn-1 are distributed throughout the OE and extend fibers to the developing OB. In wild-type mice, npn-1-positive (npn-1(+)) axons extend to lateral targets in the rostral OB and medial targets in the caudal OB, avoiding regions expressing Sema3A. In Sema3A homozygous mutant mice, many npn-1(+) axons are misrouted into and through the ventral nerve layer, beginning as early as embryonic day 13 and continuing at least until birth. At postnatal day 0, npn-1(+) glomeruli are atypically located in the ventral OB of Sema3A(-/-) mice, indicating that aberrant axon trajectories are not corrected during development and that connections are made in inappropriate target regions. In addition, subsets of OCAM(+) axons that normally project to the ventrolateral OB and some lactosamine-containing glycan(+) axons that normally target the ventral OB are also misrouted in Sema3A mutants. These observations indicate that Sema3A expression by ensheathing cells plays an important role in guiding olfactory axons into specific compartments of the OB.

Effects of gamma-aminobutyric acid(A) receptor manipulation on migrating gonadotropin-releasing hormone neurons through the entire migratory route in vivo and in vitro.

GnRH neurons originate in the nasal compartment and migrate along vomeronasal fibers over the cribiform plate to the forebrain. Previously, we found gamma-aminobutyric acid (GABA) present in GnRH neurons during development. To clarify the influence of GABA across the entire GnRH migration route, we examined the effects of muscimol and bicuculline (GABA(A) agonist and antagonist) in vivo and in vitro, maintaining the integrity of the nasal-forebrain connection. For in vivo experiments, mice were administered muscimol, bicuculline, or vehicle on days 10-15 of pregnancy and were killed on embryonic day 15 (E15). For in vitro experiments, 250-microm parasagittal slices of whole heads of E13 mice were incubated with muscimol, bicuculline, or vehicle for 2 days. Muscimol inhibited GnRH cell migration and decreased extension of GnRH fibers. Bicuculline treatment led to a disorganized distribution of GnRH cells in the forebrain and a concomitant dissociation of GnRH cells from fibers of guidance. These results suggest that GABA's influence on GnRH development changes as the cells move out of the nasal compartment and extend processes toward the median eminence.

Polysialic acid facilitates migration of luteinizing hormone-releasing hormone neurons on vomeronasal axons.

Luteinizing hormone-releasing hormone (LHRH) neurons migrate from the olfactory placode to the forebrain in association with vomeronasal nerves (VNN) that express the polysialic acid-rich form of the neural cell adhesion molecule (PSA-NCAM). Two approaches were used to investigate the role of PSA-NCAM: injection of mouse embryos with endoneuraminidase N, followed by the analysis of LHRH cell positions, and examination of LHRH cell positions in mutant mice deficient in the expression of NCAM or the NCAM-180 isoform, which carries nearly all PSA in the brain. The enzymatic removal of PSA at embryonic day 12 significantly inhibited the migration of nearly half of the LHRH neuron population, without affecting the VNN tract itself. Surprisingly, the absence of NCAM or NCAM-180 did not produce this effect. However, a shift in the route of migration, resulting in an excess number of LHRH cells in the accessory olfactory bulb, was observed in the NCAM-180 mutant. Furthermore, it was found that PSA expressed by the proximal VNN and its distal branch leading to the accessory bulb, but not the branch leading to the forebrain, was associated with the NCAM-140 isoform and thus was retained in the NCAM-180 mutant. These results provide two types of evidence that PSA-NCAM plays a role in LHRH cell migration: promotion of cell movement along the VNN tract that is sensitive to acute (enzymatic), but not chronic (genetic), removal of PSA-NCAM, and a preference of a subset of migrating LHRH cells for a PSA-positive axon branch over a PSA-negative branch in the NCAM-180 mutant.

Differential laminin isoform expression in the developing rat olfactory system.

Members of the laminin family influence mammalian cells in a variety of ways, mediating adhesion, proliferation, migration, and growth of neuronal processes. Specific laminin domains act through a number of cellular interaction sites to mediate these activities. In the developing olfactory system, axons grow from the olfactory epithelium to synaptic sites in the olfactory bulb a matrix rich in laminins and known mediators of laminin-axon interactions include integrins and a galectin-1/glycoconjugate adhesion system. Using biochemistry, immunocytochemistry, and in situ hybridization, we identified alpha 2, alpha 3, beta 1, beta 2 and gamma 1 laminin isoforms in the late embryonic and neonatal rat olfactory system. However, alpha 1-containing laminin could not be detected in association with olfactory neurons. Immunocytochemistry revealed that beta 2 laminin is preferentially expressed in the ventral and lateral nerve layer of the olfactory bulb and in the main olfactory axon tracks, but is undetectable in the accessory system during embryonic and early postnatal development. In contrast, beta 1 and gamma 1 laminins are evenly distributed throughout the olfactory bulb and in both the main and accessory olfactory axon tracks. The differential localization of laminin chains in vivo is likely to have functional significance for the development and maintenance of the olfactory system.

Gonadotropin-releasing hormone containing neurons and olfactory fibers during development: from lamprey to mammals.

Gonadotropin releasing-hormone (GnRH) regulates the hypothalamo-pituitary-gonadal axis in all vertebrates. The vast majority of GnRH neurons are thought to be derived from progenitor cells in medial olfactory placodes. Several antibodies and lectins that recognize cell surface carbohydrates have been useful for delineating the migratory pathway from the olfactory placodes and vomeronasal organ, through the nasal compartment, and across the cribriform plate into the brain. In rats, alpha-galactosyl-linked glycoconjugates (immunoreactive with the CC2 monoclonal antibody) are expressed on fibers along the GnRH migration pathway and approximately 10% of the GnRH neuronal population. In lamprey, the alpha-galactosyl binding lectin, Grifonia simplicifolia-I (GS-1), identifies cells and fibers of the developing olfactory system. In contrast to the CC2 immunoreactive GnRH neurons in rats, the GS-1 does not label a subpopulation of presumptive GnRH neurons in lamprey. Results from these and other experiments suggest that GnRH neurons in developing lamprey do not originate within the olfactory placode, but rather within proliferative zones of the diencephalon. However, the overlap of olfactory- and GnRH-containing fibers from prolarval stages to metamorphosis, suggest that olfactory stimuli may play a major role in the regulation of GnRH secretion in lamprey throughout life. By contrast, olfactory fibers are directly relevant to the migration of GnRH neurons from the olfactory placodes in mammalian species. Primary interactions between olfactory fibers and GnRH neurons are likely transient in mammals, and so in later life olfactory modulation of GnRH secretion is likely to be indirect.

Expression of gamma-aminobutyric acid and gonadotropin-releasing hormone during neuronal migration through the olfactory system.

Neurons containing the decapeptide GnRH originate in the olfactory placodes and migrate into the central nervous system during fetal development. The neurotransmitter gamma-aminobutyric acid (GABA) has been proposed as a trophic factor and may also influence neuronal migration. Immunocytochemical analyses were conducted in fetal rats, mice, and humans to identify potential developmental relationships between cells containing GABA, and GnRH neurons. Cells containing GABA were found along the nasal portion of the GnRH migration pathway in rats, mice, and humans during development. A peak number of cells containing immunoreactive GABA was observed in the nasal compartment of rats at embryonic day 15. At this time (E15), a majority of GnRH neurons were clustered in the region of the cribriform plate. By postnatal day 1, all GnRH neurons had migrated into the CNS and GABA cells were virtually absent from the nasal compartment. Double-label and confocal analyses of GABA and GnRH in mice and rats demonstrated that some olfactory GABAergic neurons coexpress GnRH. This implies that neurons that transiently express GABA originate in olfactory placodes and migrate into the forebrain. Based on the transient dual-label and adjacent relationships between GABA and GnRH containing cells in the nasal compartment, and other data showing migrational and trophic roles for GABA in development, we suggest that GABA may directly influence GnRH neuronal migration and development.

Migration of neurons containing gonadotropin releasing hormone (GnRH) in slices from embryonic nasal compartment and forebrain.

During development, neurons containing gonadotropin-releasing hormone traverse fiber bundles in the nose, cross into the brain, and move through a maze of glial and axonal fibers. To test whether GnRH neurons utilize cues intrinsic to their migration route to traverse the nasal/brain boundary, tissue slices that maintain connections between the forebrain and nasal compartment were prepared from mouse embryos. Cell migration between the nasal and brain compartments was evident based on changes in cell positions after successive days in vitro.

The migration of luteinizing hormone-releasing hormone neurons in the developing rat is associated with a transient, caudal projection of the vomeronasal nerve.

Luteinizing hormone-releasing hormone (LHRH) neurons originate in the olfactory placode and vomeronasal organ and migrate to the brain from embryonic day 14 (E14) in the rat. We investigated the development of the vomeronasal nerve and its role as a guide for migrating LHRH neurons. Using fluorescent, lipophilic dye tracing methods, we observed axons that emerge from the vomeronasal organ and cross the nasal septum as several large fascicles. At E14-15, these fascicles converge as they enter the region of the cribriform plate and subsequently disperse, projecting dorsally and caudally across the olfactory bulb and rostral forebrain. At E16, the dorsal branch of the vomeronasal nerve forms a more tightly fasciculated projection; the caudal fibers remain dispersed, extending along the medial forebrain. The number of caudally directed axons decreases during development, leaving four or five present at postnatal day 4 (P4). Immunohistochemical studies indicate that the vomeronasal nerve can be divided into four spatially distinct subpopulations of fibers. One subset, composed of caudal fibers that terminate in the lamina terminalis, selectively expresses TAG-1, a transient axonal surface glycoprotein and PSA-N-CAM, a highly polysialated form of neural cell adhesion molecule. The extension and subsequent retraction of this branch of the vomeronasal nerve corresponds spatially and temporally with the migration of LHRH neurons from the nasal cavity to the brain. Our studies show that between E14 and E18, LHRH neurons migrate in contact with the TAG-1+, PSA-N-CAM+ caudal branch of the vomeronasal nerve.

Can gonadal steroids influence cell position in the developing brain?

The preoptic area/anterior hypothalamus (POA/AH) is a site where hormones dramatically influence development. The POA/AH is comprised of multiple subgroups, but little is known about the derivation of these subgroups during development. Results from several laboratories suggest that some cells in the POA/AH originate from progenitor cells in other regions of the developing nervous system. We are exploring pathways for migration in the developing POA/AH in two ways. First, we are examining the distribution of radial glial processes as potential migratory guides using immunocytochemistry. We have identified a transient pattern of radial glial processes from the lateral ventricles to the pial surface at the base of the POA/AH. Additionally, the expression of a molecule in radial glial processes originating in the third ventricle was decreased by prenatal treatment with testosterone. Second, we are utilizing time-lapse video microscopy in vitro to assess the extent and direction of movements of fluorescent dye-labeled cells at different ages in brain slice preparations from the POA/AH of developing rats. Data from these studies indicate that cell migration in the POA/AH includes movements along dorsal-ventral routes and from lateral to medial positions, in addition to the predicted medial to lateral pathway away from the third ventricle. Several researchers have examined effects of gonadal steroids on neurite outgrowth, cell differentiation, cell death, and synaptogenesis. The determination of cell position, however, may be a key event influenced by gonadal steroids earlier in development. The characterization of migratory pathways that contribute to permanent changes in brain structure and ultimately function is essential for unraveling the process of sexual differentiation.

Altered 9-O acetylation of disialogangliosides in cerebellar Purkinje cells of the nervous mutant mouse.

Some gangliosides in the nervous system are developmentally down-regulated, but many other gangliosides continue to be expressed in the adult nervous system. We have previously demonstrated that the 9-O-acetylated gangliosides recognized by a monoclonal antibody, P-path, confer unique compartmentation among Purkinje cell groups in the normal adult cerebellum. We have continued to explore the role of this group of gangliosides in cerebellar organization by investigating the biochemical and cellular expression of this unique epitope in the cerebellum of the mutant mouse, nervous, where postnatally, most Purkinje cells degenerate. Overall ganglioside composition of nervous cerebellum is similar to wild type cerebellum. However, quantitative analysis of gangliosides by TLC-immunostaining shows that the relative concentration of 9-O-acetylated gangliosides varies considerably. In nervous cerebellum, there is more than a three-fold increase in the concentration of 9-O-acetyl disialolactosyl ceramide (GD3), and 9-O-acetyl disialolactoneotetraosyl ceramide (LD1) is decreased to 25% of wild type. In addition, GD3 ganglioside, the immediate precursor of 9-O-acetyl GD3, is detected at 1/3 of the level of wild type cerebellum, and LD1 ganglioside, the precursor of 9-O-acetyl LD1, is virtually absent from nervous cerebellum. Thus, in nervous cerebellum the ratio of 9-O-acetyl GD3 to its disialoganglioside precursor is dramatically increased compared to wild type cerebellum. In accord with the altered expression of 9-O-acetyl gangliosides, immunoelectron microscopy demonstrates a change in the subcellular distribution in mutant Purkinje cells. Instead of being associated with the somatic and dendritic membranes, P-path immunoreactivity is located internally, in the cytoplasm of Purkinje cell bodies and their dendrites. In addition to the changes in the cerebellum, the other regions of the brain decreased in size by about 15% in the nervous mutant. In the ganglioside composition of these regions of nervous brain, 9-O-acetyl GD3 nearly doubled, but 9-O-acetyl LD1 and other gangliosides did not differ. Our findings of significant changes in 9-O-acetylated gangliosides, accompanied by the overall decrease in brain size, suggest that carbohydrate or glycolipid metabolism is abnormal in the nervous mutant mouse brain.

Rat olfactory neurons can utilize the endogenous lectin, L-14, in a novel adhesion mechanism.

L-14 is a divalent, lactosamine-binding lectin expressed in many vertebrate tissues. In the rat nervous system, L-14 expression has been observed previously in restricted neuronal subsets within the dorsal root ganglia and spinal cord. In this study we report that L-14 is expressed by nonneuronal cells in the rat olfactory nerve. We demonstrate that L-14 binds and co-localizes with two ligands in the rat olfactory system: a beta-lactosamine-containing glycolipid, and a putative member of the laminin family. The former is expressed on the surfaces of nascent olfactory axons originating from neuron cell bodies in the olfactory epithelium. The latter is present in the extracellular matrix of the axonal path leading to synaptic targets in the olfactory bulb. In vitro, we find that recombinant L-14 promotes primary olfactory neuron adhesion to two laminin family members, and promotes intercellular adhesion. Both activities are dose-dependent, and are independent of integrin-mediated mechanisms. We have thus found that L-14 can serve two distinct adhesive functions in vitro, and propose that L-14 in vivo can promote olfactory axon fasciculation by crosslinking adjacent axons and promote axonal adhesion to the extracellular matrix.

A unique neuronal glycolipid defines rostrocaudal compartmentalization in the accessory olfactory system of rats.

A monoclonal antibody, CC6, reacts with a complex glycolipid whose expression in the rat is restricted to the olfactory system. Structural analysis reveals that the glycolipid contains two alpha-fucose branches; one each at internal and external beta-galactose residues. Immunocytochemistry demonstrates in rat embryos that CC6 glycolipid expression is restricted to a subset of neurons in the vomeronasal organ (VNO) and their corresponding axon projections in the caudal accessory olfactory bulb (AOB). This pattern of expression in the accessory olfactory system is the converse of the pattern revealed by a previously characterized antiglycolipid antibody that reacts with VNO neurons projecting to the rostral AOB. The CC6-reactive glycolipid is also expressed on a subset of neurons in the main olfactory epithelium. Postnatally, axons from these CC6 positive sensory neurons converge to form a limited number of axon bundles running longitudinally through the nerve layer of the main olfactory bulb. These data provide further evidence that groups of vomeronasal and olfactory neurons expressing unique surface molecules project axons that terminate in selected targets in the AOB and OB, respectively.

The expression of H-like blood group glycolipids in small cell carcinoma of the lung.

Monoclonal antibody SM1 has been shown to be preferentially reactive with small cell carcinoma of the lung (SCCL) cell lines by fluorescent and radioimmunoassay membrane staining (1). Using solid phase indirect radioimmunoassay, the antigen is not detected in non-SCCL lung carcinomas histologically classified as squamous carcinoma, adenocarcinoma or large cell carcinoma, and other tumors, viz; pheochromocytoma, a mesoderm derived lymphoblastic leukemia cell line or in normal human brain, heart, liver, colon, endothelial tissues of the aorta and blood vessels, skin, omentum, muscle, lung parenchyma and is weakly reactive with bronchial mucosa, pancreas, and kidney. The membrane antigens detected by SM1 were isolated from small cell carcinoma of the lung (SCCL) cell line, SW2, using anion exchange chromatography and thin layer chromatography, and were further analysed by exoglycosidase and endoglycosidase treatments followed by chemical staining and immunostaining with SM1 and other antibodies. We show here that SM1 antibody reacts with a group of fucose-containing neutral glycolipids and gangliosides many of which are cross-reactive with antibodies to H antigens.

Peptide growth factor control of olfactory neurogenesis and neuron survival in vitro: roles of EGF and TGF-beta s.

The olfactory epithelium (OE) is unique in the mammalian nervous system as a site of continual neurogenesis. Though many studies have described this process in vivo and olfactory neurogenesis can be demonstrated in vitro, the specific factors that modulate this process have not been defined. Noting the common ectodermal origin and structural similarity between the OE and epidermis, peptide factors known to modulate epidermal differentiation were tested in OE cultures. Our results demonstrate that EGF acts as a mitogen for the basal cells that give rise to olfactory neurons and that transforming growth factor-beta s (TGF-beta s) promote neurogenesis. Using a neutralizing antibody, we show that it is likely that the endogenous neurogenic factor is TGF-beta 2, or a very closely related factor.

Relationship of migrating luteinizing hormone-releasing hormone neurons to unique olfactory system glycoconjugates in embryonic rats.

Following their birth in olfactory placode, luteinizing hormone-releasing hormone (LHRH)-containing neurons migrate across the developing cribriform plate and form a dispersed population in the mammalian basal forebrain. The present study reveals the colocalization of unique glycoconjugate antigens (detected with monoclonal antibody CC2) on a subset of LHRH-immunoreactive (LHRHir) cell bodies and growth cones in the rostral forebrain during embryonic development in rats. In addition, LHRHir neurons were found along CC2-immunoreactive (CC2ir) fibers in the nasal cavity, across the cribriform plate, and in the rostral forebrain. At embryonic Day 16 (E16) approximately 20% of the LHRHir neuronal population in the forebrain had the CC2 epitope on surfaces of cell bodies. This percentage fell as the number of LHRHir neurons in the forebrain increased. Prior to the detection of LHRH-containing neurons, beginning on E14, CC2ir glycoconjugates were observed on vomeronasal cells and axons and also on a dorsomedial subset of olfactory neurons and axons. As early as E14 CC2ir fibers extended into the rostral forebrain. LHRHir neurons were seen in close apposition to CC2ir fibers in both the nasal cavity and rostral forebrain. These studies raise the possibility that CC2ir glycoconjugates provide a specific chemical guide for a subset of LHRH neurons along a part of their migratory pathways. The small percentage of LHRHir neurons which have CC2ir on their surfaces prenatally may constitute a selective homogenous functional subgroup within the population of LHRH neurons.

A monoclonal anti-glycoconjugate antibody defines a stage and position-dependent gradient in the developing sympathoadrenal system.

The expression of complex carbohydrate antigens was analysed in developing sympathoadrenal cells of the rat using monoclonal antibodies that react with unique carbohydrate structures. CC1 and CC4 are monoclonal antibodies that react specifically with beta-N-acetylgalactosamine and alpha-galactose/alpha-fucose moieties, respectively. CC1-reactive glycoconjugates are expressed in embryonic superior cervical ganglion (SCG) cells as early as embryonic day 15 (E15). CC4 is expressed in the SCG only for a brief period starting at E18 and then disappearing at P5. During their transient period of expression, CC1 antigens are expressed uniformly throughout the SCG at E15-17, but are then restricted to the rostral portion of the SCG from E18 to P4. CC4 is also concentrated in the rostral portion of the SCG between E21 and P4. In the adrenal medulla, CC1 and CC4 antigens display a post-natal onset of expression commencing approximately at P14 and continue to be expressed on a subset of cells which contain tyrosine hydroxylase (TH). The expression of CC1, however, is restricted to phenylethanolamine-N-methyltransferase-(PNMT)-negative chromaffin cells, whereas CC4 is not. CC1 and CC4-expressing cells appear to be scattered throughout the adrenal medulla without any particular topographic orientation. These findings suggest that the CC1 monoclonal antibody defines a stage-specific differentiation antigen in the sympathoadrenal lineage. Additionally, the CC1 antigen may confer important positional information in the embryonic SCG by distinguishing rostral from caudal neuronal cell bodies.

Dysmyelinogenesis in animal model of GM1 gangliosidosis.

Magnetic resonance imaging (MRI), pathologic examinations, and biochemical analyses were performed on 2 different canine mutants with GM1 gangliosidosis (i.e., English Springer Spaniel and Portuguese Water Dog) and on age- and sex-matched controls. Serial MRI studies were also performed on a child with infantile-onset GM1 gangliosidosis. The affected dogs had abnormalities on MRI, including a relative increase in gray matter and an abnormal signal intensity of cerebral and cerebellar white matter observed on T2-weighted MRI. White matter changes on MRI were similar to white matter abnormalities observed in a 15-month-old boy with GM1 gangliosidosis. The weight ratio of white to gray matter from the frontal lobe was markedly reduced. Microscopic examination revealed characteristic ballooned neurons which stained lightly with Luxol-fast blue. The central cerebral and cerebellar folia white matter exhibited pallor and gliosis, while the corpus callosum and fornix stained normally with Luxol-fast blue. Axons appeared intact on Bodian staining. Ultrastructural studies revealed fewer myelinated axons in affected puppies. Total gangliosides in gray matter were elevated. Thin-layer chromatography demonstrated GM1 ganglioside as the predominant ganglioside. The amount of cerebrosides and sulfatides was reduced in the gray and white matter when compared to controls but the ratio in gray and white matter remained unchanged. Immunostaining of neutral glycolipids disclosed increased amounts of stage-specific embryonic antigen-1 glycolipid in gray matter. These findings suggest that canine models for GM1 gangliosidosis are associated with abnormal myelin development which may be similar to the human disease.