Whole transcriptome sequencing and biomineralization gene architecture associated with cultured pearl quality traits in the pearl oyster, Pinctada margaritifera.

BACKGROUND: Cultured pearls are unique gems produced by living organisms, mainly molluscs of the Pinctada genus, through the biomineralization properties of pearl sac tissue. Improvement of P. margaritifera pearl quality is one of the biggest challenges that Polynesian research has faced to date. To achieve this goal, a better understanding of the complex mechanisms related to nacre and pearl formation is essential and can now be approached through the use of massive parallel sequencing technologies. The aim of this study was to use RNA-seq to compare whole transcriptome expression of pearl sacs that had producing pearls with high and low quality. For this purpose, a comprehensive reference transcriptome of P. margaritifera was built based on multi-tissue sampling (mantle, gonad, whole animal), including different living stages (juvenile, adults) and phenotypes (colour morphotypes, sex). RESULTS: Strikingly, few genes were found to be up-regulated for high quality pearls (n = 16) compared to the up-regulated genes in low quality pearls (n = 246). Biomineralization genes up-regulated in low quality pearls were specific to prismatic and prism-nacre layers. Alternative splicing was further identified in several key biomineralization genes based on a recent P. margaritifera draft genome. CONCLUSION: This study lifts the veil on the multi-level regulation of biomineralization genes associated with pearl quality determination.

Outcomes of endoscopic ethmoidectomy performed on a day-case basis: a prospective bi-centric study.

Evaluation of endoscopic ethmoidectomy performed as a day-case in terms of security, quality, and satisfaction of the patient. This prospective observatory bi-centric study over 1 year included 74 patients undergoing an ethmoidectomy respecting the eligibility criteria of ambulatory care. We recorded patients' demographic data, operative details, satisfaction, postoperative course, and follow-up results. Nasal symptoms were evaluated by SNOT-22 on preoperative appointment and postoperatively at D30. No non-absorbable nasal packing was used, eventually in the case of preoperative-bleeding absorbable gelatine packing. The postoperative follow-up took place at D1 by phone call and at D10 and D30 to assess complications, Visual Analogue Scale, and state of ethmoidal corridors by endoscopic exam. Patients benefited of bilateral ethmoidectomy in 82.4 % cases associated with septoplasty in 42 %. The majority (95 %) was discharged on the same day. Only one patient had bleeding at D0 and was kept in standard hospitalization, such as three other patients for medical or organizational reasons not related to surgery. At D1, 23 % described postoperative light bleeding but needed no revisit and pain was estimated at 1.3 (VAS). No readmission was observed, and no major complication was noted. SNOT-22 decreased successfully by 56 %, statistically related to postoperative treatment of corticosteroids and in the case of Samter triad. 97 % of patients were satisfied of the ambulatory care. These results suggest that within an experienced and dedicated day-case medical and paramedical team, ethmoidectomy can be safely performed on a day-case basis with high quality of taking care and satisfaction of patients.

The MERMAID study: indoor and outdoor average pollutant concentrations in 10 low-energy school buildings in France.

Indoor air quality was characterized in 10 recently built energy-efficient French schools during two periods of 4.5 days. Carbon dioxide time-resolved measurements during occupancy clearly highlight the key role of the ventilation rate (scheduled or occupancy indexed), especially in this type of building, which was tightly sealed and equipped with a dual-flow ventilation system to provide air refreshment. Volatile organic compounds (VOCs) and inorganic gases (ozone and NO2 ) were measured indoors and outdoors by passive techniques during the occupied and the unoccupied periods. Over 150 VOC species were identified. Among them, 27 species were selected for quantification, based on their occurrence. High concentrations were found for acetone, 2-butanone, formaldehyde, toluene, and hexaldehyde. However, these concentrations are lower than those previously observed in conventional school buildings. The indoor/outdoor and unoccupied/occupied ratios are informative regarding emission sources. Except for benzene, ozone, and NO2 , all the pollutants in these buildings have an indoor source. Occupancy is associated with increased levels of acetone, 2-butanone, pentanal, butyl acetate, and alkanes.

Prognostic and predictive values of oncogenic BRAF, NRAS, c-KIT and MITF in cutaneous and mucous melanoma.

BACKGROUND: Mutations of BRAF, NRAS and c-KIT oncogenes are preferentially described in certain histological subtypes of melanoma and linked to specific histopathological features. BRAF-, MEK- and KIT-inhibitors led to improvement in overall survival of patients harbouring mutated metastatic melanoma. OBJECTIVES: To assess the prevalence and types of BRAF, NRAS, c-KIT and MITF mutations in cutaneous and mucous melanoma and to correlate mutation status with clinicopathological features and outcome. METHODS: Clinicopathological features and mutation status of 108 samples and of 98 consecutive patients were, respectively, assessed in one retrospective and one prospective study. Clinicopathological features were correlated with mutation status and the predictive value of these mutations was studied. RESULTS: This work identified significant correlations between BRAF mutations and melanoma occurring on non-chronic sun-damaged skin and superficial spreading melanoma (P < 0.05) on one hand, and between NRAS mutations and nodular melanoma (P < 0.05) on the other hand. Younger age (P < 0.05), microscopic (P < 0.05) and macroscopic (P < 0.05) lymphatic involvement at diagnosis of primary melanoma were significantly linked to BRAF mutations. A mutated status was a positive predictive factor of a response to BRAF inhibitors (OR = 3.44). Mutated melanoma showed a significantly (P = 0.038) higher objective response rate to cytotoxic chemotherapy (26.3%) than wild-type tumours (6.7%). CONCLUSION: Clinical and pathological characteristics of the primary melanoma differed between wild-type and BRAF- or NRAS-mutated tumours. Patients with BRAF-mutated tumours were younger at diagnosis of primary melanoma. Patients carrying mutations showed better responses better to specific kinase inhibitors and interestingly also to systemic cytotoxic chemotherapy.

Microtubule-associated protein 2c reorganizes both microtubules and microfilaments into distinct cytological structures in an actin-binding protein-280-deficient melanoma cell line.

The emergence of processes from cells often involves interactions between microtubules and microfilaments. Interactions between these two cytoskeletal systems are particularly apparent in neuronal growth cones. The juvenile isoform of the neuronal microtubule-associated protein 2 (MAP2c) is present in growth cones, where we hypothesize it mediates interactions between microfilaments and microtubules. To approach this problem in vivo, we used the human melanoma cell, M2, which lacks actin-binding protein-280 (ABP-280) and forms membrane blebs, which are not seen in wild-type or ABP-transfected cells. The microinjection of tau or mature MAP2 rescued the blebbing phenotype; MAP2c not only caused cessation of blebbing but also induced the formation of two distinct cellular structures. These were actin-rich lamellae, which often included membrane ruffles, and microtubule-bearing processes. The lamellae collapsed after treatment with cytochalasin D, and the processes retracted after treatment with colchicine. MAP2c was immunocytochemically visualized in zones of the cell that were devoid of tubulin, such as regions within the lamellae and in association with membrane ruffles. In vitro rheometry confirmed that MAP2c is an efficient actin gelation protein capable of organizing actin filaments into an isotropic array at very low concentrations; tau and mature MAP2 do not share this rheologic property. These results suggest that MAP2c engages in functionally specific interactions not only with microtubules but also with microfilaments.

Infratemporal fossa tumors: When to suspect a malignant tumor? A retrospective cohort study of 62 cases.

OBJECTIVES: Infratemporal fossa (ITF) tumors are rare and little is known about their general epidemiology, making it sometimes difficult for clinicians, who seldom encounter them, to distinguish between benign and malignant forms on the basis of the initial clinical and radiological work-up alone. The objectives of this retrospective study were: (i) to determine the respective prevalences of the various histologic types of ITF tumor, and (ii) to assess associations between certain clinical and radiological features and malignancy. METHODS: A single-center observational study in a university hospital included all new consecutive cases of ITF tumor treated from January 2000 to December 2016. Histologic type, demographics, clinical presentation and imaging findings were analyzed. RESULTS: In total, 62 patients were included. 74% of tumors were benign (n=46) and 26% malignant. Juvenile nasopharyngeal angiofibroma, adenoid cystic carcinoma and schwannoma were the most frequent histologic types, accounting for 47%, 16% and 10% of cases, respectively. The only clinical or imaging signs significantly associated with malignancy were trismus, facial pain, facial hypoesthesia and neural invasion on magnetic resonance imaging (all P-values<0.05). CONCLUSION: This study provides general epidemiological data on ITF tumors, and identified several clinical and radiologic signs to help clinicians suspect malignancy.

I kappaB alpha physically interacts with a cytoskeleton-associated protein through its signal response domain.

The I kappaB alpha protein is a key molecular target involved in the control of NF-kappaB/Rel transcription factors during viral infection or inflammatory reactions. This NF-kappaB-inhibitory factor is regulated by posttranslational phosphorylation and ubiquitination of its amino-terminal signal response domain that targets I kappaB alpha for rapid proteolysis by the 26S proteasome. In an attempt to identify regulators of the I kappaB alpha inhibitory activity, we undertook a yeast two-hybrid genetic screen, using the amino-terminal end of I kappaB alpha as bait, and identified 12 independent interacting clones. Sequence analysis identified some of these cDNA clones as Dlc-1, a sequence encoding a small, 9-kDa human homolog of the outer-arm dynein light-chain protein. In the two-hybrid assay, Dlc-1 also interacted with full-length I kappaB alpha protein but not with N-terminal-deletion-containing versions of I kappaB alpha. I kappaB alpha interacted in vitro with a glutathione S-transferase-Dlc-1 fusion protein, and RelA(p65) did not displace this association, demonstrating that p65 and Dlc-1 contact different protein motifs of I kappaB alpha. Importantly, in HeLa and 293 cells, endogenous and transfected I kappaB alpha coimmunoprecipitated with Myc-tagged or endogenous Dlc-1. Indirect immunofluorescence analyzed by confocal microscopy indicated that Dlc-1 and I kappaB alpha colocalized with both nuclear and cytoplasmic distribution. Furthermore, Dlc-1 and I kappaB alpha were found to associate with the microtubule organizing center, a perinuclear region from which microtubules radiate. Likewise, I kappaB alpha colocalized with alpha-tubulin filaments. Taken together, these results highlight an intriguing interaction between the I kappaB alpha protein and the human homolog of a member of the dynein family of motor proteins and provide a potential link between cytoskeleton dynamics and gene regulation.

Juvenile and mature MAP2 isoforms induce distinct patterns of process outgrowth.

Microtubule-associated protein-2 (MAP2) is the most abundant MAP in neurons, where its distribution is restricted to the somatodendritic compartment. This molecule undergoes developmentally regulated alternative splicing, resulting in at least two isoforms, a juvenile isoform (termed MAP2c) and a mature isoform (MAP2), with greatly different molecular masses. Spodoptera frugiperda (Sf9) cell expression of the juvenile versus the mature MAP2 isoform generates two distinct patterns of process outgrowth. The smaller juvenile isoform induces multiple short thin processes. Mature MAP2 tends to induce single processes that are considerably thicker than those processes induced by juvenile MAP2. We found important differences in the variability of spacing between microtubules and the number of microtubules along the processes induced by MAP2c and mature MAP2. MAP2c showed variability with most microtubules spaced as closely as with tau, but some spaced as far apart as with mature MAP2. Over their length, the mature MAP2 processes demonstrate proximo-distal taper, which corresponds to a narrowing of the spacing between microtubules from 90 nm to 40 nm. Moreover, there is a decreased number of microtubules in mature MAP2-induced processes whereas in tau and MAP2-induced processes, the number of microtubules is constant along the length. Based on these observations, we conclude that MAP2 isoforms can serve as architectural elements by establishing specific morphological features of processes and specific arrangements of their microtubules.

Increased beta-actin expression in an invasive moloney sarcoma virus-transformed MDCK cell variant concentrates to the tips of multiple pseudopodia.

An invasive variant of Moloney sarcoma virus-transformed MDCK cells (MSV-MDCK-INV), which was isolated by the repeated selection of cells that successfully traversed a Matrigel-coated filter, exhibits increased motile ability and presents an elongated cell shape and numerous pseudopodia. Although stress fibers are present in both MDCK and MSV-MDCK cells, MSV-MDCK-INV cells contain no stress fibers and exhibit a dense concentration of actin at the tips of pseudopodia. Relative to both MDCK and MSV-MDCK cells, the MSV-MDCK-INV cells exhibit increased expression of beta-actin and redistribution of beta-actin to the tips of pseudopodia. These actin concentrations are enriched in both F- and G-actin and, thus, represent dynamic regions of actin cytoskeleton remodeling. The acquisition of invasive properties by epithelial transformants is, therefore, associated with the increased expression of beta-actin and its concentration in actin-rich domains, which may drive pseudopodial extension and facilitate tumor cell invasion.

Taxol selectively blocks microtubule dependent NF-kappaB activation by phorbol ester via inhibition of IkappaBalpha phosphorylation and degradation.

Activation of the NF-kappa-B transcription factors has been shown to be directly influenced by changes in the microtubule cytoskeleton network. To better understand cytoskeletal regulation of NF-kappaB, experiments were performed to determine whether the microtubule (MT) stabilizing agent taxol could modulate NF-kappaB activation in the presence of different NF-kappa-B inducers. Pretreatment of murine NIH3T3 and human 293 cells with 5 microM taxol resulted in complete inhibition of phorbol, 12-myristate, 13-acetate (PMA) mediated NF-kappaB activation, detected as the loss of DNA binding and reduced NF-kappaB dependent reporter gene activity. Furthermore, in COS-7 and NIH3T3 cells, PMA-induced Ikappa-Balpha turnover was dramatically reduced in taxol treated cells, mediated via the inhibition of IkappaBalpha phosphorylation. However, taxol did not prevent TNF-alpha induced Ikappa-Balpha phosphorylation, degradation, or NF-kappaB activation, indicating that TNF-alpha acts through a microtubule-independent pathway. In vitro kinase assays with PMA stimulated cell extracts demonstrated that taxol reduced protein kinase C activity by 30%, thus implicating the loss of PKC activity as a possible regulatory target of taxol-mediated suppression of NF-kappa-B. Since PMA causes modulation of cytoarchitecture through PKC activation, microtubule integrity and cell morphology was analysed by indirect immunofluorescence. Both PMA and nocodazole, a MT depolymerizing agent, caused microtubule depolymerization, whereas TNF-alpha did not alter MT integrity; concomitant taxol treatment blocked both nocodazole and PMA induced depolymerization of MTs, as well as NF-kappaB induction, thus demonstrating a link between microtubule depolymerization and NF-kappaB activation. These observations illustrate a novel biological activity of taxol as a selective inhibitor of NF-kappa-B activity, suggesting a link between the state of microtubule integrity and gene regulation.

Inactivation of Rho signaling pathway promotes CNS axon regeneration.

Regeneration in the CNS is blocked by many different growth inhibitory proteins. To foster regeneration, we have investigated a strategy to block the neuronal response to growth inhibitory signals. Here, we report that injured axons regrow directly on complex inhibitory substrates when Rho GTPase is inactivated. Treatment of PC12 cells with C3 enzyme to inactivate Rho and transfection with dominant negative Rho allowed neurite growth on inhibitory substrates. Primary retinal neurons treated with C3 extended neurites on myelin-associated glycoprotein and myelin substrates. To explore regeneration in vivo, we crushed optic nerves of adult rat. After C3 treatment, numerous cut axons traversed the lesion to regrow in the distal white matter of the optic nerve. These results indicate that targeting signaling mechanisms converging to Rho stimulates axon regeneration on inhibitory CNS substrates.

Dystonin Is Essential for Maintaining Neuronal Cytoskeleton Organization.

The mouse neurological mutant dystonia musculorum (dt) suffers from a hereditary sensory neuropathy. We have previously described the cloning and characterization of the dt gene, which we named dystonin (Dst). We had shown that dystonin is a neural isoform of bullous pemphigoid antigen 1 (Bpag1) with an N-terminal actin-binding domain. It has been shown previously that dystonin is a cytoskeletal linker protein, forming a bridge between F-actin and intermediate filaments. Here, we have used two different antibody preparations against dystonin and detected a high-molecular-weight protein in immunoblot analysis of spinal cord extracts. We also show that this high-molecular-weight protein was not detectable in the nervous system of all dt alleles tested. Immunohistochemical analysis revealed that dystonin was present in different compartments of neurons-cell bodies, dendrites, and axons, regions which are rich in the three elements of the cytoskeleton (F-actin, neurofilaments, and microtubules). Ultrastructural analysis of dt dorsal root axons revealed disorganization of the neurofilament network and surprisingly also of the microtubule network. In this context it is of interest that we observed altered levels of the microtubule-associated proteins MAP2 and tau in spinal cord neurons of different dt alleles. Finally, dt dorsal root ganglion neurons formed neurites in culture, but the cytoskeleton was disorganized within these neurites. Our results demonstrate that dystonin is essential for maintaining neuronal cytoskeleton integrity but is not required for establishing neuronal morphology. Copyright 1998 Academic Press.

Thienyl-BOPHY dyes as promising templates for bulk heterojunction solar cells.

The synthesis and characterization of bis(difluoroboryl)-1,2-bis((1H-pyrrol-2-yl)methylene)hydrazone functionalized with two lateral vinyl-thienyl modules and exhibiting strong absorption in the 400-800 nm window in thin films are reported. Bulk heterojunction solar cells assembled with these dyes and a fullerene derivative (PC71BM), using very small quantities of the additive diiodooctane, give a power conversion efficiency as high as 4.3% with short-circuit current values of 10.9 mA cm(-2), an open-circuit voltage of 0.7 V and external quantum efficiencies higher than 70% over a broad range of wavelengths (580 to 720 nm).

High conductivity organic thin films for spintronics: the interface resistance bottleneck.

Highly electrochemically doped poly(2,5-bis(3-dodecyl-2-yl)-thieno[3,2-b]thiophene (pBTTT) thin films exhibiting remarkably high conductivities values reaching 3000-5000 Ω(-1) cm(-1) are investigated. Experimental evidence of delocalized transport properties of this material at the onset of metallicity makes it an ideal candidate for spin valve device integration. Nevertheless, the interface resistance between the polymer and metallic electrodes is orders of magnitudes larger than the expected spin resistance of the active channel. This prevents the collection of a spin current. This finding can explain the lack of success in making lateral organic spin valves reported in the literature, especially the related absence of spin signals in non-local spin valve and Hanle current measurements in organic thin films.

Antigenic map of the rat cerebellar cortex: the distribution of parasagittal bands as revealed by monoclonal anti-Purkinje cell antibody mabQ113.

Both anatomical and physiological mapping methods have revealed that the mammalian cerebellar cortex consists of a family of parasagittal bands of cells, each band with its own pattern of afferent and efferent axons. Monoclonal antibody mabQ113 recognizes an unknown polypeptide antigen that is confined to a subset of rat cerebellar Purkinje cells. Immunoreactive cells are arranged into parasagittal bands extending throughout the vermis and hemispheres. Expression of the Q113 epitope by individual Purkinje cells may not be all-or-nothing, since the bands tend to be more strongly stained in the vermis than the hemispheres. The band display is symmetrical about the midline and reproducible from individual to individual. Whole-mount immunocytochemistry and serial reconstruction reveal a median band of mabQ113+ Purkinje cells adjacent to the midline (P1+) and six other positive bands disposed symmetrically at either side (P2+ to P7+). Bands are distinct throughout most of the cortex but tend to fuse ventrally and caudally. There are two sources of interindividual differences. Firstly, most animals express supernumerary "satellite" bands in the vermis. Satellite bands are usually only one cell wide, are not bilaterally symmetrical, and differ in position and number from individual to individual. Secondly, the precise position of an individual band can differ, perhaps according to the variable cortical lobulation, for example, the position of P4+ in lobules VIII/IX and P6+ in lobule VII. While a scheme of parasagittal bands is a good description of the vermian organization, the distribution of mabQ113+ and mabQ113- Purkinje cells in the hemispheres may be better described as a checkerboard of antigenic patches.

Immunocytochemical demonstration of topographic ordering of Purkinje cell axon terminals in the fastigial nuclei of the rat.

We have used a monoclonal antibody, mabQ113, which selectively stains a subset of cerebellar Purkinje cells, to study the topography of the corticonuclear projection of the median vermis to the fastigial nuclei in the rat. The fastigial projection zone contains both mabQ113+ and mabQ113- Purkinje cells grouped into parasagittal bands. The immunoreactivity extends throughout the Purkinje cell including the axon terminals and thus it is possible to investigate the topographic distribution of mabQ113+ terminals. In the fastigial nuclei the target cells receiving mabQ113+ axon terminals are concentrated in the caudal pole. In the rostral pole, cells receive anti-GAD+ terminals but not mabQ113+ terminals. There is no gradient in anti-GAD staining. Double-labelling experiments with mabQ113, anti-GAD, and an antisynaptic antibody mabQ155 suggest that there is little or no mixing of mabQ113+ and mabQ113- Purkinje cell terminals on the same target neuron.

Development of parasagittal zonation in the rat cerebellar cortex: MabQ113 antigenic bands are created postnatally by the suppression of antigen expression in a subset of Purkinje cells.

Monoclonal antibody mabQ113 recognizes a polypeptide antigen that, in the adult cerebellum, is confined to a subset of Purkinje cells that are clustered together to form parasagittal bands interposed by similar nonimmunoreactive bands. The Purkinje cell compartments are congruent with bands of climbing fibers projecting from subregions of the inferior olivary complex (IOC). The array of mabQ113 parasagittal bands appears late in the development of the cortex. Weak mabQ113 immunoreactivity is first seen at postnatal day 6 (P6) in the Purkinje cells of the posterior lobe of the vermis. From the earliest stages there are signs of differential expression of the mabQ113 antigen in clusters of Purkinje cells: four mabQ113+ clusters are clearly present in the posterior lobe of the vermis at P6-P7. Their relation to the adult band display remains uncertain. During the next few days immunoreactivity spreads rostrally throughout the rest of the vermis and laterally to include the Purkinje cells in the hemispheres, until by P12 all the Purkinje cells in the cerebellum are mabQ113+. Nevertheless, signs of the adult band display are seen already in the vermis where the cells destined to become the vermal mabQ113+ bands (P1+, P2+ and P3+) stain more intensely than their neighbours. Following the stage of global mab113 epitope expression, bands are created by the selective suppression of immunoreactivity by Purkinje cells in the P- regions. By P15 the mabQ113+ and mabQ113- bands are clearly differentiated in the vermis and selective staining has begun to appear in the hemispheres also. The band pattern matures gradually during the third and fourth postnatal weeks until the adult appearance is attained by P30. The cerebellar afferent projections were lesioned to explore the interplay of cerebellar input and mabQ113 expression. The olivocerebellar projection was lesioned bilaterally by using 3-acetylpyridine in the adult and unilaterally in the newborn by electrolytic lesion and unilateral inferior cerebellar pedunculectomy. Mossy fibers from the dorsal and ventral spinocerebellar tracts were lesioned surgically both in adults and in newborn and trigeminal projections to the cerebellum were removed in the newborn by unilateral ablation of the spinal trigeminal nucleus. The consequences of total blockage of vibrissal and hindlimb inputs were also explored in both adults and neonates. None of these treatments led to a modification in the pattern of mabQ113 epitope expression.(ABSTRACT TRUNCATED AT 400 WORDS)

Expression of compartmentation antigen zebrin I in cerebellar transplants.

The mammalian cerebellum is divided into multiple parasagittal compartments as defined by the organization of afferent and efferent projections and by the pattern of expression of several biochemical markers. One such marker is the antigen zebrin I, a 120 kD polypeptide of unknown function that is expressed differentially by a subset of Purkinje cells. Zebrin I+ Purkinje cells are grouped into an array of 14 parasagittal bands interposed by zebrin I- compartments. This Purkinje cell compartmentation corresponds to compartments in the olivocerebellar projection. The afferent axon compartments are present prior to the expression of the mature zebrin I phenotype, thus raising the possibility that differential afferent input regulates the zebrin I phenotype of the target of that input. Lesion studies in the neonate preclude a role for afferent inputs in the regulation of zebrin I expression postnatally, but a prenatal role in commitment still remains open. To explore this possibility, cerebellar anlagen were dissected from embryos at embryonic days 12-15, that is, prior to any contact with afferents, and transplanted ectopically into adult hosts. In the first series of experiments, the grafts were placed into the anterior chamber of the eye, and in the second series, into cavities prepared in the neocortex. Grafts were allowed to mature and then were immunoperoxidase or immunofluorescence stained for zebrin I immunoreactivity. Zebrin I was expressed by grafted Purkinje cells in cortico and in oculo. Double-labelling experiments confirmed that both the zebrin I+ and the zebrin I- phenotypes were present. The zebrin I immunoreactivity revealed that the zebrin I+ Purkinje cells resemble those in situ with an extensive dendritic arborization that extends through the molecular layer perpendicular to the long axes of the folia. In conclusion, the present data suggest that afferent input does not play a role in the determination of the zebrin I phenotype of Purkinje cells.

Synaptophysin expression during synaptogenesis in the rat cerebellar cortex.

In order to study the mechanisms of synaptogenesis in the rat cerebellar cortex, a library of monoclonal antibodies has been generated against proteins of the isolated synapse. One recognizes a glycosylated 38 kDa protein that is concentrated in the synaptic vesicle fraction and resembles synaptophysin biochemically in its molecular weight, charge, and pattern of glycosylation. In the adult cerebellar cortex, the antisynaptophysin(mabQ155) immunoreactivity is codistributed with synapses. Immunoreactivity is strongest in the molecular layer where punctate deposits of reaction product outline the Purkinje cell dendrites. Discrete small profiles, consistent with the distribution of basket cell axon terminals, surround the Purkinje cells, and in the granular layer the synaptic glomeruli are intensely stained. There is no immunoreactivity in the white matter axon tracts. Electron microscope immunocytochemistry confirms the synaptic location of the antigen and suggests that the reaction product is associated with synaptic vesicles. Both round and flat vesicle populations are immunoreactive. Antisynaptophysin(mabQ155) has been used to follow synaptogenesis in the developing rat cerebellum. In the newborn rat (P0), despite the paucity of synapses, there is some specific immunoreactivity, especially in the subcortical white matter. Electron microscopy shows that the antigenicity is associated with vesicles within growth cones, filopodia, and immature axon profiles. During development, antisynaptophysin immunoreactivity increases progressively, along with the maturing cell populations, for both the granule cell-Purkinje cell and the mossy fiber-granule cell synapses. Quantitative biochemical analysis confirms the cytochemical results. These data suggest that neuronal growth cones express a synapse-specific antigen before complete morphological synapses are present.