Carotenoid pigmentation in salmon: variation in expression at BCO2-l locus controls a key fitness trait affecting red coloration.

Carotenoids are primarily responsible for the characteristic red flesh coloration of salmon. Flesh coloration is an economically and evolutionarily significant trait that varies inter- and intra-specifically, yet the underlying genetic mechanism is unknown. Chinook salmon (Oncorhynchus tshawytscha) represents an ideal system to study carotenoid variation as, unlike other salmonids, they exhibit extreme differences in carotenoid utilization due to genetic polymorphisms. Here, we crossed populations of Chinook salmon with fixed differences in flesh coloration (red versus white) for a genome-wide association study to identify loci associated with pigmentation. Here, the beta-carotene oxygenase 2-like (BCO2-l) gene was significantly associated with flesh colour, with the most significant single nucleotide polymorphism explaining 66% of the variation in colour. BCO2 gene disruption is linked to carotenoid accumulation in other taxa, therefore we hypothesize that an ancestral mutation partially disrupting BCO2-l activity (i.e. hypomorphic mutation) allowed the deposition and accumulation of carotenoids within Salmonidae. Indeed, we found elevated transcript levels of BCO2-l in white Chinook salmon relative to red. The long-standing mystery of why salmon are red, while no other fishes are, is thus probably explained by a hypomorphic mutation in the proto-salmonid at the time of divergence of red-fleshed salmonid genera (approx. 30 Ma).

Sperm competition, but not major histocompatibility divergence, drives differential fertilization success between alternative reproductive tactics in Chinook salmon.

Post-copulatory sexual selection processes, including sperm competition and cryptic female choice (CFC), can operate based on major histocompatibility (MH) genes. We investigated sperm competition between male alternative reproductive tactics [jack (sneaker) and hooknose (guard)] of Chinook salmon (Oncorhynchus tshawytscha). Using a full factorial design, we examined in vitro competitive fertilization success of paired jack and hooknose males at three time points after sperm activation (0, 15 and 60 s) to test for male competition, CFC and time effects on male fertilization success. We also examined egg-mediated CFC at two MH genes by examining both the relationship between competitive fertilization success and MH divergence as well as inheritance patterns of MH alleles in resulting offspring. We found that jacks sired more offspring than hooknose males at 0 s post-activation; however, jack fertilization success declined over time post-activation, suggesting a trade-off between sperm speed and longevity. Enhanced fertilization success of jacks (presumably via higher sperm quality) may serve to increase sneaker tactic competitiveness relative to dominant hooknose males. We also found evidence of egg-mediated CFC (i.e. female × male interaction) influencing competitive fertilization success; however, CFC was not acting on the MH genes as we found no relationship between fertilization success and MH II ß1 or MH I α1 divergence and we found no deviations from Mendelian inheritance of MH alleles in the offspring. Our study provides insight into evolutionary mechanisms influencing variation in male mating success within alternative reproductive tactics, thus underscoring different strategies that males can adopt to attain success.

Flavored e-cigarette use: Characterizing youth, young adult, and adult users.

The purpose of this study is to investigate how the use of flavored e-cigarettes varies between youth (12-17 years old), young adults (18-29 years old), and older adults (30 + years old). Cross-sectional surveys of school-going youth (n = 3907) and young adult college students (n = 5482) in Texas, and young adults and older adults (n = 6051) nationwide were administered in 2014-2015. Proportions and 95% confidence intervals were used to describe the percentage of e-cigarette use at initiation and in the past 30 days that was flavored, among current e-cigarette users. Chi-square tests were applied to examine differences by combustible tobacco product use and demographic factors. Most e-cigarette users said their first and "usual" e-cigarettes were flavored. At initiation, the majority of Texas school-going youth (98%), Texas young adult college students (95%), and young adults (71.2%) nationwide said their first e-cigarettes were flavored to taste like something other than tobacco, compared to 44.1% of older adults nationwide. Fruit and candy flavors predominated for all groups; and, for youth, flavors were an especially salient reason to use e-cigarettes. Among adults, the use of tobacco flavor at initiation was common among dual users (e-cigarettes + combustible tobacco), while other flavors were more common among former cigarette smokers (P = 0.03). Restricting the range of e-cigarette flavors (e.g., eliminating sweet flavors, like fruit and candy) may benefit youth and young adult prevention efforts. However, it is unclear what impact this change would have on adult smoking cessation.

Effect of parental mate choice and semi-natural early rearing environment on the growth performance and seawater tolerance of Chinook salmon Oncorhynchus tshawytscha.

To assess whether parental mate choice and early rearing in a semi-natural spawning channel may benefit the culture of Chinook salmon Oncorhynchus tshawytscha, 90 day growth trials were conducted using hatchery O. tshawytscha (hatchery), mate choice O. tshawytscha (i.e. the offspring of parents allowed to choose their own mate) that spent 6 months in a spawning channel prior to hatchery rearing (channel) and mate choice O. tshawytscha transferred to the hatchery as fertilized eggs (transfer). During the growth trials, all O. tshawytscha stocks were reared separately or in either mixed channel and hatchery or transfer and hatchery groups for comparison of performance to traditional practices. After 60 days in fresh water, all O. tshawytscha were transferred to seawater for an additional 30 days. Reared separately, all stocks grew c. 4.5 fold over 90 days but specific growth rate (G) and food conversion efficiency were higher in fresh water than after seawater transfer on day 60. In contrast, hatchery O. tshawytscha from mixed hatchery and channel and hatchery and transfer growth trials had a larger mass and length gain than their counterparts on day 60, but reduced G in seawater. In general, plasma levels of growth hormone, insulin-like growth factor I and cortisol did not differ among any O. tshawytscha groups in either the separate or mixed growth trials. Despite some differences in gill Na(+),K(+)-ATPase activity, all O. tshawytscha had a high degree of seawater tolerance and experienced virtually no perturbation in plasma chloride following seawater transfer. Overall, all O. tshawytscha exhibited similar growth and seawater performance under traditional hatchery conditions and any benefit derived from either parental mate choice or semi-natural early rearing environment was only observed in the presence of mutual competition with hatchery O. tshawytscha.

Transcriptional differences between triploid and diploid Chinook salmon (Oncorhynchus tshawytscha) during live Vibrio anguillarum challenge.

Understanding how organisms function at the level of gene expression is becoming increasingly important for both ecological and evolutionary studies. It is evident that the diversity and complexity of organisms are not dependent solely on their number of genes, but also the variability in gene expression and gene interactions. Furthermore, slight differences in transcription control can fundamentally affect the fitness of the organism in a variable environment or during development. In this study, triploid and diploid Chinook salmon (Oncorhynchus tshawytscha) were used to examine the effects of polyploidy on specific and genome-wide gene expression response using quantitative real-time PCR (qRT-PCR) and microarray technology after an immune challenge with the pathogen Vibrio anguillarum. Although triploid and diploid fish had significant differences in mortality, qRT-PCR revealed no differences in cytokine gene expression response (interleukin-8, interleukin-1, interleukin-8 receptor and tumor necrosis factor), whereas differences were observed in constitutively expressed genes, (immunoglobulin (Ig) M, major histocompatibility complex (MHC) -II and beta-actin) upon live Vibrio anguillarum exposure. Genome-wide microarray analysis revealed that, overall, triploid gene expression is similar to diploids, consistent with their similar phenotypes. This pattern, however, can subtly be altered under stress (for example, handling, V. anguillarum challenge) as we have observed at some housekeeping genes. Our results are the first report of dosage effect on gene transcription in a vertebrate, and they support the observation that diploid and triploid salmon are generally phenotypically indistinguishable, except under stress, when triploids show reduced performance.

The MHC and non-random mating in a captive population of Chinook salmon.

Detailed analysis of variation in reproductive success can provide an understanding of the selective pressures that drive the evolution of adaptations. Here, we use experimental spawning channels to assess phenotypic and genotypic correlates of reproductive success in Chinook salmon (Oncorhynchus tshawytscha). Groups of 36 fish in three different sex ratios (1:2, 1:1 and 2:1) were allowed to spawn and the offspring were collected after emergence from the gravel. Microsatellite genetic markers were used to assign parentage of each offspring, and the parents were also typed at the major histocompatibility class IIB locus (MHC). We found that large males, and males with brighter coloration and a more green/blue hue on their lateral integument sired more offspring, albeit only body size and brightness had independent effects. There was no similar relationship between these variables and female reproductive success. Furthermore, there was no effect of sex ratio on the strength or significance of any of the correlations. Females mated non-randomly at the MHC, appearing to select mates that produced offspring with greater genetic diversity as measured by amino-acid divergence. Females mated randomly with respect to male genetic relatedness and males mated randomly with respect to both MHC and genetic relatedness. These results indicate that sexual selection favours increased body size and perhaps integument coloration in males as well as increases genetic diversity at the MHC by female mate choice.

Heritability and Y-chromosome influence in the jack male life history of chinook salmon (Oncorhynchus tshawytscha).

Jacking in chinook salmon (Oncorhynchus tshawytscha) is an alternative reproductive strategy in which males sexually mature at least 1 year before other members of their year class. We characterize the genetic component of this reproductive strategy using two approaches; hormonal phenotypic sex manipulation, and a half-sib breeding experiment. We 'masculinized' chinook salmon larvae with testosterone, reared them to first maturation, identified jacks and immature males based on phenotype, and genotyped all fish as male ('XY') or female ('XX') using PCR-based Y-chromosome markers. The XY males had a much higher incidence of jacking than the XX males (30.8% vs 9.9%). There was no difference in body weight, gonad weight, and plasma concentrations of testosterone and 17beta-estradiol between the two jack genotypes, although XY jacks did have a higher gonadosomatic index (GSI) than XX jacks. In the second experiment, we bred chinook salmon in two modified half-sib mating designs, and scored the number of jacks and immature fish at first maturation. Heritability of jacking was estimated using two ANOVA models: dams nested within sires, and sires nested within dams with one-half of the half-sib families common to the two models. The sire component of the additive genetic variance yielded a high heritability estimate and was significantly higher than the dam component (h(2)(sire) = 0.62 +/- 0.21; h(2)(dam) = -0.14 +/- 0.12). Our experiments both indicated a strong sex-linked component (Y-chromosome) to jacking in chinook salmon, although evidence for at least some autosomal contribution was also observed.

Abstinence education for urban youth.

Teen sexual problems in the U.S. are reaching enormous proportions. Attempts to prevent common problems, pregnancy and sexually transmitted diseases, are underway through the persistent efforts of community, health, academic and government organizations. Abstinence education is one of the current attempts. However, the lack of well defined theoretical frameworks and analyses of outcomes have limited progress in the study of abstinence education. This article describes a pilot program in abstinence-only education provided to six groups of young teens within an urban middle school. The framework for the program, cognitive social learning theory, is described and operationalized. Student pretest-posttest attitudes, open-ended written comments about the program and the researchers' anecdotes about behavioral changes in the students are the outcome measures. Positive attitudes about premarital abstinence increased for all six groups; for four of the six groups the increase was statistically significant.

Alpha and beta subunits of CaM-kinase II are localized in different neurons in chick ciliary ganglion.

The ciliary ganglion of the chicken contains only two types of neurons. Using monoclonal antibodies against the alpha and the beta subunits of Ca2+/calmodulin-stimulated protein kinase II (CaMPK-II) we found that the alpha-subunit was localized to the choroid neurons while beta subunit was associated with the ciliary neurons. As both neurons receive their inputs from the oculomotor nerve, while their postganglionic axons leave via different nerves, the ciliary ganglion of the chicken is a neuronal system in which the functional differences between alpha and beta CaMPK-II homopolymers in the regulation of synaptic transmission can be investigated.

Patterns of morphological variation within myelin internodes of normal peripheral nerve: quantitative analysis by confocal microscopy.

Knowledge of variations in the morphology of normal myelinated peripheral nerve fibres is fundamental to subsequent interpretation of neuropathology. It would be advantageous for structural analysis of normal variations to be based on entire myelin internodes, but acquisition of such data via the classic approach of nerve fibre teasing has been hindered by limitations in optical resolution and specimen preparation. This study addressed these limitations through a new confocal imaging method which permits detailed visualisation of individual myelinated fibres in intact peripheral nerve trunks, and quantitated previously unrecognised patterns of morphological variation within normal internodes. The study focused particularly on Schmidt-Lanterman incisures, the narrow cytoplasmic channels which traverse normal compact myelin and provide foci for disruption of the compact sheath in a number of peripheral neuropathies. Analysis was based on confocal fluorescence images of multiple sequential internodes, traced within posterior tibial nerve trunks of adult male Sprague-Dawley rats. The strength of relationships between internodal size variables (length, fibre diameter, myelin sheath thickness) and total number of incisures per internode were documented. Each internode was divided into 4 regions of equivalent length (regions 1-4), and variations in the distribution of incisures and Schwann cell nuclear location were evaluated. Regional variations were consistent, irrespective of differences in fibre diameter, myelin sheath thickness, and internodal length. Expressed in terms of proximodistal orientation, there was a unimodal distribution of incisures within internodes of this fibre population (diameter range 5-9 microns), with region 3 containing the highest number of incisures and region 4 the lowest (P < 0.05). The Schwann cell nucleus was located more frequently in region 3 than in region 2 (P < 0.01). Contrary to previous reports, an incisure was found in close association with the nucleus in at least 50% of internodes. Documentation of frequent incisure-nuclear association and consistent patterns of variation within internodes extends knowledge of the microanatomy of normal peripheral nerve, and may provide insight into the functional role of incisures. Demonstration of such patterns in normal nerve may contribute to the understanding of pathological change, for example progression of ovoid formation from midinternodal regions during wallerian degeneration.

Cryo-immunogold ultrastructural localization of laminin in adult rat peripheral nerve.

Elucidation of the functional roles of the extracellular matrix component laminin in adult peripheral nerve has been hindered by differing accounts of its ultrastructural localization. This is the first report applying the advantages of the cryo-immunogold technique to laminin localization in peripheral nerve. Laminin labelling was found over the basal lamina and possibly over the immediately subjacent Schwann cell plasma membrane, but specific labelling appeared to be absent from other membranes (including those of non-myelinated axon/Schwann cell clusters) and from endoneurial collagen fibrils. It would appear that the functional roles played by laminin in normal adult peripheral nerve are likely to be mediated via its localization in the basal lamina, rather than through a more widespread distribution within the endoneurium.

Imaging myelinated nerve fibres by confocal fluorescence microscopy: individual fibres in whole nerve trunks traced through multiple consecutive internodes.

Current methods of morphological analysis do not permit detailed imaging of individual myelinated fibres over substantial lengths without disruption of neighbouring, potentially significant, cellular and extracellular relationships. We report a new method which overcomes this limitation by combining aldehyde-induced fluorescence with confocal microscopy. Myelin fluorescence was intense relative to that from other tissue components, enabling individual myelinated nerve fibres to be traced for distances of many millimeters in whole PNS nerve trunks. Image obtained with a Bio-Rad MRC-600 confocal laser scanning microscope clearly displayed features of PNS and CNS myelinated fibres including nodes of Ranvier; fibre diameter; sheath thickness and contour; branch points at nodes; as well as (in the PNS) Schmidt-Lanterman incisures and the position of Schwann cell nuclei. Direct comparisons using the same specimens (whole nerve trunks; also teased fibres) showed confocal imaging to be markedly superior to conventional fluorescence microscopy in terms of contrast, apparent resolution and resistance to photobleaching. Development of the fluorophore was examined systemically in sciatic nerves of young adult rats. In separate experiments, animals were perfused systemically using (1) 5% glutaraldehyde; (2) Karnovsky's solution; (3) 4% paraformaldehyde; buffered with either 0.1 M sodium phosphate or sodium cacodylate (pH 7.4). The concentration of glutaraldehyde in the fixative solution was the principal determinant of fluorescence intensity. Confocal imaging was achieved immediately following perfusion with 5% glutaraldehyde or Karnovsky's. Fluorescence intensity increased markedly during overnight storage in these fixatives and continued to increase during subsequent storage in buffer alone. The fluorophore was stable and resistant to fading during storage (15 months at least), enabling data collection over extended periods. To demonstrate application of the method in neuropathology, individual fibres in transected sciatic nerve trunks were traced through multiple successive internodes: Classical features of Wallerian degeneration (axonal swelling and debris; ovoid formation and incisure changes; variation among fibres in the extent of degeneration) were displayed. The method is compatible with subsequent ultrastructural examination and will complement existing methods of investigation of myelinated fibre anatomy and pathology, particularly where preservation of 3-dimensional relationships or elucidation of spatial gradients are required.

Morphometric analysis of axons myelinated during adult life in the mouse superior cervical ganglion.

In experimental studies addressing the regulation of myelin formation and maintenance by Schwann cells, the sympathetic nervous system of young adult rodents has served a key role as an essentially nonmyelinated yet modifiable control tissue. Nevertheless there is clear evidence of substantial myelination in the superior cervical ganglion (SCG) of normal mice and rats of more advanced age. Against this background, interpretation of experimental outcomes in particular sympathetic tissues will require detailed quantitative control data taking account of animal age. To provide a baseline for future investigations on myelin remodelling, an ultrastructural morphometric study of myelinated fibres in the SCG was undertaken in 4 strains (QS, Balb/C, C57 and CBA) of adult male mice aged 32-72 wk. Numbers of myelinated fibres in SCG cross-sections varied substantially between individual animals, and the mean numbers for QS (132), Balb/C (165) and CBA (254) were significantly higher than that for C57 (32). Both axonal and fibre diameter were distributed unimodally (means for the 4 strains ranged from 2.3-2.4 microns and 3.2-3.6 microns respectively). Myelin spiral length was distributed unimodally and skewed to the right (range of means = 227-357 microns) and was significantly greater in QS mice as compared with the other 3 strains. While the mean g ratio (axonal diameter/fibre diameter) was significantly lower in QS mice than in the other 3 strains, the range for mean g ratio was 0.64-0.73, indicating that myelination had proceeded appropriately even though late in onset in this tissue. The index of circularity was high in all strains, both for axons (range of means = 0.80-0.88) and fibres (range of means = 0.84-0.89). The small axonal and fibre diameter and unimodal distribution are consistent with the characteristics of autonomic myelinated fibres and it is probable that most are postganglionic sympathetic fibres arising within the SCG. In terms of providing a sufficient population of myelinated fibres for future experimental studies, the QS, Balb/C and CBA strains would be preferable to C57 mice.

Structural maturation of synapses in the rat superior cervical ganglion continues beyond four weeks of age.

We have examined the morphology of preganglionic synapses in the rat superior cervical ganglion (SCG) at 10 days, 4 weeks and 1 year. Between 10 days and 4 weeks the mean thickness of the postsynaptic density (PSD) increased from 45.9 +/- 0.1 nm to 52.1 +/- 1.7 nm (P = 0.017), the mean length of the PSD (0.41 +/- 0.02 microns) did not change, and the distribution of synapses on the neuronal surface changed with a decrease in the proportion of somatic and an increase in the proportion of dendritic spine synapses. Since both synapse elimination and synapse formation are occurring during this period several mechanisms may contribute to these changes. However, between 4 weeks and 1 year, when there is no net change in the number of synapses, the mean length of the PSD increased to 0.53 +/- 0.02 microns (P = 0.001), there was no change in either the mean thickness of the PSD or the distribution of the synapses but the proportion of concave ('smile') synapses increased. A comparison with previous developmental studies of synapses in cerebral cortex of rat and chicken indicate that both the nature and the rate of synapse maturation can vary between different populations of synapses.

Myelin sheath survival after guanethidine-induced axonal degeneration.

Membrane-membrane interactions between axons and Schwann cells are required for initial myelin formation in the peripheral nervous system. However, recent studies of double myelination in sympathetic nerve have indicated that myelin sheaths continue to exist after complete loss of axonal contact (Kidd, G. J., and J. W. Heath. 1988. J. Neurocytol. 17:245-261). This suggests that myelin maintenance may be regulated either by diffusible axonal factors or by nonaxonal mechanisms. To test these hypotheses, axons involved in double myelination in the rat superior cervical ganglion were destroyed by chronic guanethidine treatment. Guanethidine-induced sympathectomy resulted in a Wallerian-like pattern of myelin degeneration within 10 d. In doubly myelinated configurations the axon, inner myelin sheath (which lies in contact with the axon), and approximately 75% of outer myelin sheaths broke down by this time. Degenerating outer sheaths were not found at later periods. It is probably that outer sheaths that degenerated were only partially displaced from the axon at the commencement of guanethidine treatment. In contrast, analysis of serial sections showed that completely displaced outer internodes remained ultrastructurally intact. These internodes survived degeneration of the axon and inner sheath, and during the later time points (2-6 wk) they enclosed only connective tissue elements and reorganized Schwann cells/processes. Axonal regeneration was not observed within surviving outer internodes. We therefore conclude that myelin maintenance in the superior cervical ganglion is not dependent on direct axonal contact or diffusible axonal factors. In addition, physical association of Schwann cells with the degenerating axon may be an important factor in precipitating myelin breakdown during Wallerian degeneration.

Myelin sheath survival following axonal degeneration in doubly myelinated nerve fibers.

Axonal contact plays a critical role in initiating myelin formation by Schwann cells. However, recent studies of "double myelination" have indicated that myelin maintenance continues in Schwann cells completely displaced from physical contact with the axon. This raises the possibility either that diffusible trophic factors are produced by the axon, or that the axon is not required for myelin maintenance by these displaced Schwann cells. To test these hypotheses, the axons involved in double myelination in the mouse superior cervical ganglion (SCG) were transected surgically by a transganglionic lesion. The inferior pole of the SCG was resected to limit axonal regeneration. This method produced a typical Wallerian pattern of degeneration in the superior pole, without compromising the blood supply or introducing nonspecific trauma. EM analysis at 1 and 5 d postoperatively showed that initially the axon degenerated, followed by breakdown of the inner myelin sheath. In those configurations where the outer Schwann cell was only partly displaced from the axon, the outer myelin sheath degenerated simultaneously. However, in completely displaced internodes the outer sheath survived degeneration of the axon and inner sheath. Outer internodes remained intact for at least 5 weeks after transection (the longest time point in this study), at which time they enclosed reorganized processes of the inner Schwann cells, their basal lamina, and numerous collagen fibrils. Axonal regeneration within surviving outer internodes was rare and was characterized by the development of typical Remak ensheathment by the inner Schwann cells. We conclude that in the mouse SCG, myelin maintenance does not depend on the continued presence of the axon. These data suggest further that myelin breakdown in Wallerian degeneration may be initiated by mechanisms other than absence of a viable axon.

Myelin maintenance by Schwann cells in the absence of axons.

Formation and maintenance of myelin sheaths in the peripheral nervous system are regulated by unknown molecular interactions that are thought to depend upon physical contact between Schwann cells and axons. However, recent studies describing axons surrounded by two concentric myelin internodes in the superior cervical ganglion (SCG) of normal rodents have demonstrated that the outer myelin internodes are maintained without physical contact with the axon. To determine whether the centrally enclosed axon has a trophic effect in maintaining these remote outer internodes, we have produced axonal degeneration by surgical or chemical means. The results indicate that maintenance of myelin internodes totally displaced from axonal contact depends neither upon the presence of the axon nor on diffusible axonal factors. A further implication of these studies is that myelin breakdown during Wallerian degeneration is regulated by a positive signal which originates in degenerating nerves, rather than solely by loss of axonal trophic substances.

A human colon cancer cell line established from collagen matrix cultures transplanted into nude mice.

A new human colon cancer cell line (020588) has been derived by means of a combined in vitro matrix-in vivo xenograft technique. The tumour cell line is carcino-embryonic antigen positive, displays a marker chromosome and proliferates in chemically-defined serum-free culture medium. The chemosensitivity pattern for the tumour cell line was similar to that observed for the parent tumour cells. The novel method used to establish this continuous human tumour cell line may have several advantages over standard techniques.

Distribution of P0 protein and the myelin-associated glycoprotein in peripheral nerves from Trembler mice.

The Trembler mouse has a dysymelination of peripheral nerves that includes hypomyelination, failure of myelin compaction, and demyelination/remyelination. We have localized the myelin proteins P0 and myelin associated glycoprotein in Trembler peripheral nerve and correlated their distributions with the ultrastructure of myelin internodes. Immunocytochemically, myelin-associated glycoprotein was localized in Schwann cell periaxonal membranes, Schmidt-Lanterman incisures, paranodal loops, and internal and external mesaxons. P0 staining was located over compact myelin and regions of Schwann cell cytoplasm rich in Golgi membranes. An unusual abundance of small, P0-stained, Golgi-related vesicles was found in some Schwann cells. P0 protein was also detected in multiple spiral wraps of myelin-associated glycoprotein-positive mesaxon membranes. At some sites the periodicity of the myelin membranes was intermediate to that found in mesaxon membranes and compact myelin. The distance between apposing extracellular leaflets was similar to that found in mesaxon membranes, while the cytoplasmic leaflets were fused but twice as thick as normal major dense lines. These intermediate membranes were stained by P0 and myelin-associated glycoprotein antiserum. These studies suggest that altered transport and/or translocation of P0 and myelin-associated glycoprotein results in defective myelin compaction in Trembler peripheral nerve.

Mechanisms of synaptic plasticity. Changes in postsynaptic densities and glutamate receptors in chicken forebrain during maturation.

We have shown that the synapse maturation phase of synaptogenesis is a model for synaptic plasticity that can be particularly well-studied in chicken forebrain because for most forebrain synapses, the maturation changes occur slowly and are temporally well-separated from the synapse formation phase. We have used the synapse maturation phase of neuronal development in chicken forebrain to investigate the possible link between changes in the morphology and biochemical composition of the postsynaptic density (PSD) and the functional properties of glutamate receptors overlying the PSD. Morphometric studies of PSDs in forebrains and superior cervical ganglia of chickens and rats have shown that the morphological features of synapse maturation are characteristic of a synaptic type, but that the rate at which these changes occur can vary between types of synapses within one animal and between synapses of the same type in different species. We have investigated, during maturation in the chicken forebrain, the properties of the N-methyl-D-aspartate (NMDA) subtype of the glutamate receptors, which are concentrated in the junctional membranes overlying thick PSDs in the adult. There was no change in the number of NMDA receptors during maturation, but there was an increase in the rate of NMDA-stimulated uptake of 45Ca2+ into brain prisms. This functional change was not seen with the other ionotropic subtypes of the glutamate receptor and was NMDA receptor-mediated. The functional change also correlated with the increase in thickness of the PSD during maturation that has previously been shown to be due to an increase in the amount of PSD associated Ca(2+)-calmodulin stimulated protein kinase II (CaM-PK II). Our results provide strong circumstantial evidence for the regulation of NMDA receptors by the PSD and implicate changing local concentrations of CaM-PK II in this process. The results also indicate some of the ways in which properties of existing synapses can be modified by changes at the molecular level.