Invasive fungal infections: epidemiology and analysis of antifungal prescriptions in onco-haematology.

WHAT IS KNOWN AND OBJECTIVE: Invasive fungal infections (IFI) are associated with high rates of morbidity and mortality, particularly in onco-haematology patients. We aimed to study the epidemiology of IFI in neutropenic patients and estimate the economic impact of treatment of those infections. METHODS: All patients hospitalized in onco-haematology, and treated with antifungal agents, in 2005 were investigated. Four features were studied: the diagnosis for each patient, the antifungal drugs used, the thoracic densitometry reports and the sero-mycological data. Infectious episodes were stratified according to the EORTC 2008 classification criteria (10). RESULTS AND DISCUSSION: Of the 1130 patients surveyed, 192 patients received systemic antifungal agents. Of these 46% had acute leukaemia, 29% bone-marrow allografts, 7% lymphoma and 18% other malignant haemopathies. Using the EORTC 2008 criteria (10), there were 8 proved IFI (3 aspergillosis, 3 candidosis and 2 other IFI), 17 probable IFI (11 aspergillosis, 6 candidosis) and 16 possible aspergillosis. The incidence of IFI was 2·1%. Eighty patients (41·7%) had received prophylaxis: 56 with fluconazole and 24 with voriconazole. Treatment was most often empirical (n = 127, 66·1%). Combination of two antifungals was used in 17 cases. The mean duration of prophylactic, empirical, proved/probable aspergillosis-directed, candidaemia-directed and combination treatment was 19, 19, 46, 32 and 27 days, respectively. The cost of antifungal treatment in 2005 reached almost 2,000,000 €, including 427,000 € for documented infections (proved and probable), 1,246,000 € for empirical treatment and 58,300 € for prophylaxis. WHAT IS NEW AND CONCLUSION: The incidence of IFI is low but the pharmacoeconomic impact is extremely high. Improved strategies are required to reduce the frequency and duration of empirical treatment without compromising beneficial outcome.

Monoclonal antibodies to rhamnogalacturonan I backbone.

Monoclonal antibodies were raised against rhamnogalacturonan I backbone, a pectin domain, using Arabidopsis thaliana seed mucilage-derived rhamnogalacturonan I oligosaccharides--BSA conjugates. Two monoclonal antibodies, designated INRA-RU1 and INRA-RU2, selected for further characterization, were specific for the backbone of rhamnogalacturonan I, displaying no binding activity against the other pectin domains i.e. homogalacturonans, galactans or arabinans. A range of oligosaccharides was prepared by enzymatic digestion of rhamnogalacturonan I isolated from Arabidopsis thaliana seed mucilage and from sugar beet pectin, purified by low-pressure chromatography and characterized by high-performance anion-exchange chromatography and mass spectrometry. These rhamnogalacturonan I oligomers were used to characterize the binding site of the two monoclonal antibodies by competitive inhibition. Both INRA-RU1 and INRA-RU2 showed maximal binding to the [-->2)-alpha-L-rhamnosep-(1-->4)-alpha-D-galacturonic acid p-(1-->](7) structural motif but differed in their minimum binding requirement. INRA-RU2 required at least two disaccharide (rhamnose-galacturonic acid) repeats for the antibody to bind, while INRA-RU1 required a minimum of six disaccharide repeats. Furthermore, the binding capacity of INRA-RU1 decreased steeply as the number of disaccharide repeats go beyond seven. Each of these antibodies reacted with hairy regions isolated from sugar beet pectin. Immunofluorescence microscopy indicated that both antibodies can be readily used to detect rhamnogalacturonan I epitopes in various cell wall samples.

Control of glutamate clearance and synaptic efficacy by glial coverage of neurons.

Analysis of excitatory synaptic transmission in the rat hypothalamic supraoptic nucleus revealed that glutamate clearance and, as a consequence, glutamate concentration and diffusion in the extracellular space, is associated with the degree of astrocytic coverage of its neurons. Reduction in glutamate clearance, whether induced pharmacologically or associated with a relative decrease of glial coverage in the vicinity of synapses, affected transmitter release through modulation of presynaptic metabotropic glutamate receptors. Astrocytic wrapping of neurons, therefore, contributes to the regulation of synaptic efficacy in the central nervous system.

[Childhood septic temporomandibular arthritis]. / Arthrite septique temporomandibulaire de l'enfant.

OBJECTIVE: To report a clinical case of acute otitis media in a child, complicated by septic temporomandibular arthritis and to present a review of the literature. PATIENT AND METHODS: We report a case of a 7-year-old boy who presented an altered general condition, major hyperthermia, associated with a left temporozygomatic mass in a context of recurrent bilateral acute otitis media lasting for 2 months. Emergency computed tomodensitometry (CT scan) showed left temporomandibular joint arthritis. Treatment consisted of a parenteral double antibiotic therapy and prevention of temporomandibula (TM) ankylosis. RESULTS: After 20 months of follow-up, the child showed a normal ORL examination with no maxillofacial sequelae. CONCLUSION: All temporozygomatic masses presenting in a septic context should suggest the diagnosis of TM arthritis; computed tomodensitometry should be done immediately.

Comparative analysis of cell wall surface glycan expression in Candida albicans and Saccharomyces cerevisiae yeasts by flow cytometry.

The yeast Candida albicans is an opportunistic pathogen, part of the normal human microbial flora that causes infections in immunocompromised individuals with a high morbidity and mortality levels. Recognition of yeasts by host cells is based on components of the yeast cell wall, which are considered part of its virulence attributes. Cell wall glycans play an important role in the continuous interchange that regulates the balance between saprophytism and parasitism, and also between resistance and infection. Some of these molecular entities are expressed both by the pathogenic yeast C. albicans and by Saccharomyces cerevisiae, a related non-pathogenic yeast, involving similar molecular mechanisms and receptors for recognition. In this work we have exploited flow cytometry methods for probing surface glycans of the yeasts. We compared glycan expression by C. albicans and by S. cerevisiae, and studied the effect of culture conditions. Our results show that the expression levels of alpha- and beta-linked mannosides as well as beta-glucans can be successfully evaluated by flow cytometry methods using different antibodies independent of agglutination reactions. We also found that the surface expression pattern of beta-mannosides detected by monoclonal or polyclonal antibodies are differently modulated during the growth course. These data indicate that the yeast beta-mannosides exposed on mannoproteins and/or phospholipomannan are increased in stationary phase, whereas those linked to mannan are not affected by the yeast growth phase. The cytometric method described here represents a useful tool to investigate to what extent C. albicans is able to regulate its glycan surface expression and therefore modify its virulence properties.

A study of the role of neuro-glial remodeling in the oxytocin system at lactation.

Under conditions of strong secretion of neurohypophysial hormone, such as during parturition, lactation and dehydration, the hypothalamic oxytocin-system displays a remarkable morphological plasticity such that astrocytic coverage of its neurones diminishes, their surfaces become directly juxtaposed and contacted by an increased number of synapses. A growing body of evidence indicates that these anatomical changes have an impact on glutamatergic neurotransmission in the supraoptic nucleus, and may be therefore of physiological consequence. We here evaluated the consequences of the inhibition of such plasticity on the overall activity of the oxytocin system during lactation. Remodeling was prevented by performing hypothalamic microinjections in gestating rats of endoneuraminidase, an enzyme that removes polysialic acid from the neural cell adhesion molecule. Our earlier studies established that the presence of polysialic acid is a prerequisite for remodeling of the oxytocin system in the supraoptic and paraventricular nuclei. In dams in which polysialic acid was absent in all magnocellular nuclei after bilateral endoneuraminidase injections, parturition was normal and neither the frequency nor the amplitude of suckling-induced reflex milk ejections was different from vehicle-treated dams. The weight gain of pups was also normal as was water intake by the dams. We then assessed the electrical activity of antidromically identified magnocellular neurones in the polysialic acid-free supraoptic nucleus of isoflurane-anesthetized lactating rats. Basal and bursting activity characteristic of oxytocin neurones before each reflex milk ejection was not significantly different from that recorded in the supraoptic nucleus of rats with normal levels of polysialic acid. Our results indicate that neuro-glial remodeling, despite its role on fine modulation of oxytocin neuronal activity, is not essential to parturition and lactation.

The Cek1­mediated MAP kinase pathway regulates exposure of α­1,2 and ß­1,2­mannosides in the cell wall of Candida albicans modulating immune recognition.

The Cek1 MAP kinase (MAPK) mediates vegetative growth and cell wall biogenesis in the fungal pathogen Candida albicans. Alterations in the fungal cell wall caused by a defective Cek1­mediated signaling pathway leads to increased ß­1,3­glucan exposure influencing dectin­1 fungal recognition by immune cells. We show here that cek1 cells also display an increased exposure of α­1,2 and ß­1,2­mannosides (α­M and ß­M), a phenotype shared by strains defective in the activating MAPKK Hst7, suggesting a general defect in cell wall assembly. cek1 cells display walls with loosely bound material as revealed by transmission electron microscopy and are sensitive to tunicamycin, an inhibitor of N­glycosylation. Transcriptomal analysis of tunicamycin treated cells revealed a differential pattern between cek1 and wild type cells which involved mainly cell wall and stress related genes. Mapping α­M and ß­M epitopes in the mannoproteins of different cell wall fractions (CWMP) revealed an important shift in the molecular weight of the mannan derived from mutants defective in this MAPK pathway. We have also assessed the role of galectin­3, a member of a ß­galactoside­binding protein family shown to bind to and kill C. albicans through ß­M recognition, in the infection caused by cek1 mutants. Increased binding of cek1 to murine macrophages was shown to be partially blocked by lactose. Galectin-3(-/-) mice showed increased resistance to fungal infection, although galectin-3 did not account for the reduced virulence of cek1 mutants in a mouse model of systemic infection. All these data support a role for the Cek1­mediated pathway in fungal cell wall maintenance, virulence and antifungal discovery.

Role of mannose-binding lectin in intestinal homeostasis and fungal elimination.

Mannose-binding lectin (MBL) is a soluble lectin of the innate immune system that is produced by the liver and secreted into the circulation where it activates the lectin complement pathway, enhances phagocytosis of microorganisms by leukocytes, and modulates inflammation. MBL can recognize patterns on the surface of different pathogens, including Candida albicans. Our aims were to investigate whether MBL is expressed in the gut epithelium and to examine its effect on the modulation of intestinal inflammation and C. albicans elimination. Using reverse transcriptase-PCR, MBL transcripts were highly expressed in different parts of the mouse gut. MBL expression was also detected by immunoblotting and immunolocalization in response to C. albicans colonization of the gut; the highest expression of MBL was detected in the stomach. Blocking MBL by administering mannans to mice increased C. albicans colonization. MBL-deficient mice had a higher level of colonization than wild-type mice. Dextran sodium sulfate-induced colitis promoted C. albicans dissemination to the kidneys and lungs of MBL-deficient mice. MBL-deficient mice exhibited elevated expression of interleukin (IL)-17, IL-23, dectin-1, and Toll-like receptor-4. This study shows that MBL expression is induced in the gut in response to C. albicans sensing and is required for intestinal homeostasis and host defense against C. albicans.

Initiation of phospholipomannan ß-1,2 mannosylation involves Bmts with redundant activity, influences its cell wall location and regulates ß-glucans homeostasis but is dispensable for Candida albicans systemic infection.

Pathogenic and non-pathogenic fungi synthesize glycosphingolipids, which have a crucial role in growth and viability. Glycosphingolipids also contribute to fungal-associated pathogenesis. The opportunistic yeast pathogen Candida albicans synthesizes phospholipomannan (PLM), which is a glycosphingolipid of the mannosylinositol phosphorylceramide family. Through its lipid and glycan moieties, PLM contributes to the initial recognition of the yeast, causing immune system disorder and persistent fungal disease through activation of host signaling pathways. The lipid moiety of PLM activates the deregulation signaling pathway involved in yeast phagocytosis whereas its glycan moiety, composed of ß-1,2 mannosides (ß-Mans), participates to inflammatory processes through a mechanism involving Galectin-3. Biosynthesis of PLM ß-Mans involves two ß-1,2 mannosyltransferases (Bmts) that initiate (Bmt5) and elongate (Bmt6) the glycan chains. After generation of double bmtsΔ mutants, we show that Bmt5 has redundant activity with Bmt2, which can replace Bmt5 in bmt5Δ mutant. We also report that PLM is located in the inner layer of the yeast cell wall. PLM seems to be not essential for systemic infection of the yeast. However, defect of PLM ß-mannosylation increases resistance of C. albicans to inhibitors of ß-glucans and chitin synthesis, highlighting a role of PLM in cell wall homeostasis.

Bacteriome and Mycobiome Interactions Underscore Microbial Dysbiosis in Familial Crohn's Disease.

UNLABELLED: Crohn's disease (CD) results from a complex interplay between host genetic factors and endogenous microbial communities. In the current study, we used Ion Torrent sequencing to characterize the gut bacterial microbiota (bacteriome) and fungal community (mycobiome) in patients with CD and their nondiseased first-degree relatives (NCDR) in 9 familial clusters living in northern France-Belgium and in healthy individuals from 4 families living in the same area (non-CD unrelated [NCDU]). Principal component, diversity, and abundance analyses were conducted, and CD-associated inter- and intrakingdom microbial correlations were determined. Significant microbial interactions were identified and validated using single- and mixed-species biofilms. CD and NCDR groups clustered together in the mycobiome but not in the bacteriome. Microbiotas of familial (CD and NCDR) samples were distinct from those of nonfamilial (NCDU) samples. The abundance of Serratia marcescens and Escherichia coli was elevated in CD patients, while that of beneficial bacteria was decreased. The abundance of the fungus Candida tropicalis was significantly higher in CD than in NCDR (P = 0.003) samples and positively correlated with levels of anti-Saccharomyces cerevisiae antibodies (ASCA). The abundance of C. tropicalis was positively correlated with S. marcescens and E. coli, suggesting that these organisms interact in the gut. The mass and thickness of triple-species (C. tropicalis plus S. marcescens plus E. coli) biofilm were significantly greater than those of single- and double-species biofilms. C. tropicalis biofilms comprised blastospores, while double- and triple-species biofilms were enriched in hyphae. S. marcescens used fimbriae to coaggregate or attach with C. tropicalis/E. coli, while E. coli was closely apposed with C. tropicalis Specific interkingdom microbial interactions may be key determinants in CD. IMPORTANCE: Here, we characterized the gut bacterial microbiota (bacteriome) and fungal community (mycobiome) in multiplex families with CD and healthy relatives and defined the microbial interactions leading to dysbiosis in CD. We identified fungal (Candida tropicalis) and bacterial (Serratia marcescens and Escherichia coli) species that are associated with CD dysbiosis. Additionally, we found that the level of anti-Saccharomyces cerevisiae antibodies (ASCA; a known CD biomarker) was associated with the abundance of C. tropicalis We also identified positive interkingdom correlations between C. tropicalis, E. coli, and S. marcescens in CD patients and validated these correlations using in vitro biofilms. These results provide insight into the roles of bacteria and fungi in CD and may lead to the development of novel treatment approaches and diagnostic assays.

Role of extracellular signal-regulated protein kinase cascade in macrophage killing of Candida albicans.

The pathogenic yeast Candida albicans and its derived molecules stimulate a wide range of macrophage secretory functions and may adapt to escape being killed by this phagocyte. In this study, phagocytosis of C. albicans and of the nonpathogenic yeast Saccharomyces cerevisiae was shown to be associated with phosphorylation of the mitogen-activated protein kinase (MAPK)/extracellularly regulated kinase (ERK) pathway in the absence of significant activation of either p38MAPK or stress-activated protein kinase/c-Jun N-terminal kinase. However, although 80% of endocytosed C. albicans survived after 1 h, 80% of S. cerevisiae cells were killed. Considerable quantitative differences were observed between the two species in the sequential phosphorylation of MAPK/ERK kinase (MEK), extracellularly regulated kinase-1, and 90-kDa-ribosomal S6 kinases. A lower level of activation of the pathway by C. albicans was associated with a species-specific overexpression of the MEK phosphatase MAPK phosphatase (MKP)-1. Killing of both C. albicans and S. cerevisiae could be reduced using PD98059, which mimics MKP-1 and inhibits MEK phosphorylation, suggesting that specific MKP-1 activation by C. albicans could contribute to its ability to escape the yeast lytic potential of macrophages.

Beta-1,2-linked oligomannosides inhibit Candida albicans binding to murine macrophage.

Interaction of Candida albicans with cells of the macrophage lineage was examined by using heat-killed (HK) and live yeast cells. Laminarin, an analogue of the cell wall beta-glucans, strongly inhibited HK yeasts adherence to J774 cell line but had no effect on live yeast binding. Phosphopeptidomannan (PPM) from Saccharomyces cerevisiae had a limited effect on the binding of both HK and live yeasts but significant inhibition was achieved by the use of C. albicans PPM. The role of beta-1,2-oligomannosides was examined with regard to their exclusive presence within C. albicans PPM. PPM acid labile beta-1,2-oligomannosides or a synthetic beta-1,2-mannotetraose, inhibited yeasts binding in a manner comparable to the original PPM. These latter results were confirmed by using mouse peritoneal macrophages, thus suggesting a general role for beta-1,2-oligomannosides in the adherence of the yeast to the macrophage membrane.

[Candida albicans meningo-encephalo-myelo-radiculitis at an addict]. / Méningo-encéphalo-myélo-radiculite à Candida albicans chez un toxicomane.

Beside immunodepression induced by the human immunodeficiency virus, fungal infections of the central nervous system are extremely rare in heroin-addict patients. We report here a case of meningo-encephalitis with myelo-radicular lesions in a 25-year-old intravenous heroin addict but non-HIV patient, who was admitted for an acute confusion associated with gait disorders. The diagnosis of Candida albicans meningo-encephalo-myelo-radiculitis was established by magnetic resonance imagery and mycological and serological examinations of cerebrospinal fluid. The infection was cured with amphotericin B lipid complex and 5-fluorocytosine. Early diagnosis and antifungal therapy for 6 months resulted in a favorable outcome. The detection of circulating Candida mannan in cerebrospinal fluid with a more sensitive technique combined to MRI were particularly decisive to confirm Candida infection diagnosis, allowing an appropriate antifungal therapy.

Preliminary evidence for a serum disaccharide signature of invasive Candida albicans infection detected by MALDI Mass Spectrometry.

The diagnosis of systemic Candida infections is a recognized challenge. We developed a mass spectrometry strategy to detect signals from Candida molecules in patients' sera. Pre-analytical procedures were designed to extract oligosaccharides from serum. A peak m/z of at 365 was specifically revealed in sera from patients with candidaemia with regard to healthy controls. This biomarker was identified as a disaccharide, its presence did not correlate with mannanaemia or glucanaemia. Mouse models of Candida albicans colonization and infection showed that the signal was specifically associated with tissue invasion, suggesting that clinical evaluation of its usefulness in discriminating colonized and infected patients would be worthwhile.

Secretion of glycoproteins through the cell wall of Candida albicans.

A monoclonal antibody raised against the pathogenic phase of Candida albicans has been coupled to colloidal gold and used to detect the corresponding epitope in cell wall and culture medium of blastoconidia grown as germ tubes in vitro. Immunogold silver staining of Western blots of culture supernatants demonstrated release of the epitope into the culture medium. The stain revealed 3 well defined bands of 205,000, 66,000 and 30,000 Mr and a smear from the top of the gel to an Mr of 120,000. Immunoelectron microscopy of ultrathin frozen sections of the corresponding growth forms showed that epitope accumulated first in the periplasmic space, generally corresponding to plasmalemma invaginations within the cytoplasm. From these sites, it was possible to follow continuous lines of epitope distribution through the cell wall and antigenic extrusion at the cell surface. In tangential sections of intensely labeled walls, these preferential excretion ways appeared to be organized as a parallel network. Antigen emergence at the cell surface corresponded to patches of material which tended to coalesce in an easily dissociated layer, probably corresponding to the fuzzy coat. These experiments demonstrate, for the first time, preferential ways for cellular secretion through the yeast cell wall.

Localization of chitin in the cell wall of Candida albicans by means of wheat germ agglutinin. Fluorescence and ultrastructural studies.

Cytochemical localization of wheat germ agglutinin binding sites in the cell wall of Candida albicans was investigated with fluorescence and electron microscopy. Various analytical techniques were employed in order to obtain a good penetration of the cytochemical markers, glycosylated horseradish peroxidase or glycosylated ferritin. In blastospores sectioned by cryostatic methods, a weak and continuous labelling of the blastospore periphery was observed with peroxidase, whereas bud scars and inner cell wall areas were labelled with ferritin. Following enzymatic treatment with pronase whose efficiency was followed by the periodic acid-thiocarbohydrazide -- Silver proteinate technique, the inner cell wall layers of bud are strongly stained with both fluorescein and the reactions products of peroxidase. After pronase-chitinase treatment, fluorescence was observed only in the mother cell wall. Finally, ultrathin glycol methacrylate sections showed a labelling both in inner and outer layers. All these results suggest that chitin was essentially distributed in the inner wall layers near the plasmalemma and in a smaller amount in outer wall layers. On the basis of the present findings, a hypothesis of wall assembly is proposed.

Experimental study of resistance to infection by Trichophyton mentagrophytes: demonstration of memory skin cells.

The mechanisms governing the development of local immunity in experimental dermatophytosis were studied by injecting intravenously trichophytin in guinea pigs cured of a prior Trichophyton mentagrophytes infestation. Dermal cell modifications were observed which were greater in the healed zones than in those not affected during the prior dermatophyte inoculation. These modifications included lymphocyte activation and accumulation and an accumulation of basophilic leukocytes. These observations suggest that after an acute dermatophyte infection heals, immunocompetent cells remain which are more numerous at the sites of lesions and that these cells would be responsible for the increased rate of elimination of the fungus during a reinfection. This hypothesis is discussed in the framework of the relationships observed in dermatophyte infections between delayed type hypersensitivity and resistance.

Definitive chemical evidence for the constitutive ability of Candida albicans serotype A strains to synthesize beta-1,2 linked oligomannosides containing up to 14 mannose residues.

We have previously reported the presence of phosphate bound beta-1,2 linked oligomannosides with unusually high degrees of polymerization (DP > 7) in the mannan of Candida albicans strain VW32. To confirm this observation, we have prepared these oligomannosides from the mannan of C. albicans strain NIH A 207. Gel filtration chromatography and TLC analysis revealed DP up to 14. For both strains, NMR analysis confirmed the exclusive presence of beta-1,2 linkages in the pools of oligomannosides with a DP higher than 6 which presented an average DP of 10.6 (VW32) and 10.4 (NIH A 207). These results are important to consider in relation with the ability of these C. albicans derived oligomannosides to trigger TNFalpha synthesis according to their DP.

Expression of high levels of the extracellular matrix glycoprotein, tenascin-C, in the normal adult hypothalamoneurohypophysial system.

Glia and neurons of the hypothalamoneurohypophysial system (HNS) undergo reversible morphological changes, which are concomitant with the remodelling of afferents onto the neurons, under different conditions of neurohormone secretion. Here, we show that the adult rat HNS contains high levels of tenascin-C (TN-C), which is an extracellular matrix glycoprotein whose expression is usually associated with neuronal-glial interactions in the developing and lesioned central nervous system (CNS). By using light and electron microscopic immunocytochemical procedures, we visualized TN-C immunoreactivity in the hypothalamic supraoptic (SON) and paraventricular nuclei, where somata of the neurons are localized; in the median eminence, where their axons transit; and in the neurohypophysis, where they terminate. Hypothalamic areas adjacent to the magnocellular nuclei were devoid of immunoreactivity. Electron microscopy of the neurohypophysis showed immunolabelling of perivascular spaces, glial (pituicyte) and axonal surfaces, a type of labelling that also characterized the median eminence. In the hypothalamic nuclei, there was labelling of extracellular spaces and astrocytic surfaces. In normal animals, we detected no cytoplasmic reaction in glia somata, neurons, or endothelial cells. However, in animals treated with the intracellular transport blocker colchicine, there was intracytoplasmic labelling of all HNS glial cells, indicating a glial source for TN-C. Immunoblot analysis revealed TN-C isoforms of apparent high molecular weight (225, 240, and 260 kD) in the SON and median eminence, whereas lower MW forms (190/200 kD) predominated in the neurohypophysis. By using immunocytochemistry and immunoblot analysis, we found no visible differences in TN-C expression in relation to age, sex, or differing neurohypophysial secretion, which suggests that the expression of TN-C is a permanent feature of the HNS.

Evidence for structural plasticity in the supraoptic nucleus of the rat hypothalamus in relation to gestation and lactation.

Supraoptic nuclei of lactating rats present a particular anatomical organization that could serve to facilitate the synchronization of neuronal firing observed during suckling-induced reflex milk ejections. Although magnocellular neurones are usually separated by neuropil elements, particularly glial fibers, in lactating rats, numerous neurosecretory soma and dendritic profiles are in direct apposition, without glial interposition. Concomitantly, there is also a higher incidence of presynaptic terminals contacting two neurosecretory elements in the same plane of section ("double" synapses). In the present study, a quantitative ultrastructural analysis was used to trace the evolution of the structural reorganization of the nucleus at different stages of the reproductive cycle. The percentage of neurosecretory soma and dendritic profiles in direct apposition was low two weeks after the beginning of pregnancy, but the day prior to parturition, as during lactation, over 40% of all neurosecretory profiles were directly in contact and involved about 10% of the total neuronal surface membrane measured (a 5-fold increase over the corresponding frequencies recorded in virgin rats at oestrus). The contiguous neuronal membranes and associated intercellular space appeared unmodified, except for the presence of attachment plaques, that also increased in frequency at late gestation and lactation. The incidence of "double" synapses also increased by late gestation, so that at lactation, they bridged 8% of all the recorded neurosecretory somata and dendrites, (as compared to 1% in the virgin rats). Similar changes were observed during a first and second gestation and lactation. The incidence of direct appositions and "double" synapses then diminished gradually after weaning: 2 months after the end of lactation, the ultrastructure of the nucleus resembled that of virgin animals. These observations demonstrate a plasticity in the structural organization of the supraoptic nucleus that appears closely related to changing physiological states of the animal and that involves both neurone-glial relationships and the neurones' synaptic configuration.