A theory of rate coding control by intrinsic plasticity effects.

Intrinsic plasticity (IP) is a ubiquitous activity-dependent process regulating neuronal excitability and a cellular correlate of behavioral learning and neuronal homeostasis. Because IP is induced rapidly and maintained long-term, it likely represents a major determinant of adaptive collective neuronal dynamics. However, assessing the exact impact of IP has remained elusive. Indeed, it is extremely difficult disentangling the complex non-linear interaction between IP effects, by which conductance changes alter neuronal activity, and IP rules, whereby activity modifies conductance via signaling pathways. Moreover, the two major IP effects on firing rate, threshold and gain modulation, remain unknown in their very mechanisms. Here, using extensive simulations and sensitivity analysis of Hodgkin-Huxley models, we show that threshold and gain modulation are accounted for by maximal conductance plasticity of conductance that situate in two separate domains of the parameter space corresponding to sub- and supra-threshold conductance (i.e. activating below or above the spike onset threshold potential). Analyzing equivalent integrate-and-fire models, we provide formal expressions of sensitivities relating to conductance parameters, unraveling unprecedented mechanisms governing IP effects. Our results generalize to the IP of other conductance parameters and allow strong inference for calcium-gated conductance, yielding a general picture that accounts for a large repertoire of experimental observations. The expressions we provide can be combined with IP rules in rate or spiking models, offering a general framework to systematically assess the computational consequences of IP of pharmacologically identified conductance with both fine grain description and mathematical tractability. We provide an example of such IP loop model addressing the important issue of the homeostatic regulation of spontaneous discharge. Because we do not formulate any assumptions on modification rules, the present theory is also relevant to other neural processes involving excitability changes, such as neuromodulation, development, aging and neural disorders.

[Acute superior vena cava syndrome on port catheter]. / Syndrome cave supérieur aigu sur chambre à cathéter implantable.

We are reporting the case of a 78-year-old patient, presenting an acute superior vena cava syndrome, caused by a port catheter, implanted 4 years ago. The outcome was unfavourable, despite of a fibrinolytic treatment.

Specific adsorption of functionalized colloids at the surface of living cells: a quantitative kinetic analysis of the receptor-mediated binding.

This paper presents a statistical experimental study of the adsorption of colloids onto the plasma membrane of living cells mediated by specific ligand-receptor interactions. The colloids consist of lipid multilamellar liposomes (spherulites) functionalized by Shiga toxin B-subunit (STxB), while cells are cervix carcinoma epithelial cells expressing the Shiga toxin receptor, the glycolipid globotriaosyl ceramide (Gb3). The specificity of the colloid adsorption is demonstrated using both confocal microscopy and flow cytometry, while a thorough cytometry study on living cells allows characterizing the kinetics of this specific adsorption. The final number of bound colloids and the characteristic adsorption time are shown to depend on bulk concentration, as expected for a thermodynamic equilibrium. However, the colloids appear to be irreversibly attached to the membrane. We interpret this apparent irreversibility as the result of a progressive recruitment of receptors. The methodology used here, whereby microscopic mechanisms are deduced from direct quantitative measurements on living cells, might allow the optimization of drug delivery systems or the quantification of virus infectivity.

Key role of receptor density in colloid/cell specific interaction: a quantitative biomimetic study on giant vesicles.

This paper presents an experimental study of the adsorption of colloids on model membranes mediated by specific ligand-receptor interactions. The colloids consist of lipid multilamellar liposomes (spherulites) functionalized with the B-subunit of Shiga Toxin (STxB), while the membranes are lipid Giant Unilamellar Vesicles (GUV) containing STxB lipid receptor, Globotriaosylceramide (Gb3). Through confocal microscopy and flow cytometry, we show the specificity of the adsorption. Moreover, we show that flow cytometry can be used to efficiently quantify the kinetics of colloid adsorption on GUVs with very good statistics. By varying the bulk colloid concentration and receptor density in the membrane, we point out the existence of an optimum Gb3 density for adsorption. We propose that this optimum corresponds to a transition between reversible colloid adsorption at low Gb3 density and irreversible adsorption, and likely spherulite fusion, at high density. We compare our results both to STxB-colloids adhering on living cells and to free STxB proteins interacting with GUVs containing Gb3. This biomimetic system could be used for a quantitative evaluation of the early stage of virus infection or drug delivery.

RGD-functionalized spherulites as targeted vectors captured by adherent cultured cells.

Spherulites are multilamellar vesicles consisting of concentric shells that can encapsulate small organic molecules or macromolecules. We investigate the possibility of targeting neutral spherulites to adherent culture cells by functionalizing their surface with RGD-containing ligands. The strength and specificity of association of RGD spherulites with several cell lines (EAhy 926 endothelial cell line, human umbilical vein endothelial cell (HUVEC) and human osteoprogenitor (HOP) primary cells) was studied, and the molecular interaction of RGD spherulites with the EAhy 926 cell surface was investigated. We show that, after binding to cells, spherulites are internalized.

Role of a striatal slowly inactivating potassium current in short-term facilitation of corticostriatal inputs: a computer simulation study.

Striatal output neurons (SONs) integrate glutamatergic synaptic inputs originating from the cerebral cortex. In vivo electrophysiological data have shown that a prior depolarization of SONs induced a short-term (

The spherulites(TM): a promising carrier for oligonucleotide delivery.

Concentric multilamellar microvesicles, named spherulites(TM), were evaluated as an oligonucleotide carrier. Up to 80% oligonucleotide was encapsulated in these vesicles. The study was carried out on two different spherulite(TM) formulations. The spherulite(TM) size and stability characteristics are presented. Delivery of encapsulated oligonucleotide was performed on a rat hepatocarcinoma and on a lymphoblastoid T cell line, both expressing the luciferase gene. We showed that spherulites(TM) were able to transfect both adherent and suspension cell lines and deliver the oligonucleotide to the nucleus. Moreover, 48-62% luciferase inhibition was obtained in the rat hepatocarcinoma cell line when the antisense oligonucleotide targeted to the luciferase coding region was encapsulated at 500 nM concentration in spherulites(TM) of different compositions.

A model study of cellular short-term memory produced by slowly inactivating potassium conductances.

We analyzed the cellular short-term memory effects induced by a slowly inactivating potassium (Ks) conductance using a biophysical model of a neuron. We first described latency-to-first-spike and temporal changes in firing frequency as a function of parameters of the model, injected current and prior history of the neuron (deinactivation level) under current clamp. This provided a complete set of properties describing the Ks conductance in a neuron. We then showed that the action of the Ks conductance is not generally appropriate for controlling latency-to-first-spike under random synaptic stimulation. However, reliable latencies were found when neuronal population computation was used. Ks inactivation was found to control the rate of convergence to steady-state discharge behavior and to allow frequency to increase at variable rates in sets of synaptically connected neurons. These results suggest that inactivation of the Ks conductance can have a reliable influence on the behavior of neuronal populations under real physiological conditions.

Diversity of firing frequency control by A-currents in vertebrate neurons: a modeling study.

Experimental and modeling studies have accumulated strong evidence suggesting that A-currents control firing rates in invertebrate neurons. However, the direct demonstration of a similar role remains to be established in vertebrate neurons. We tested this possibility in a simulated neuron embedded with a generic model of vertebrate A-currents. Under simulated current-clamp protocols, the generic A-current produced a modest frequency reduction (15 Hz) that was constant within all firing frequencies. Modifications in steady-state properties of the A-current model within known physiological ranges annihilated or dramatically increased firing frequency reduction. These results suggest that the influence of A-currents on firing frequency should differ strongly among vertebrate neurons, and that modulations influencing A-currents provide a powerful control over the excitability of vertebrate neurons.

Attractors and pathological aspects in excitable cells.

In this article, physiological and pathological forms of excitability are studied in a two-dimensional electrical model of excitable cell endowed with a generic inward persistent conductance. Bifurcation analysis of the model is performed as a function of the maximal inward persistent conductance, the input current, or the voltage dependency of the activation function. Several discharge modes are exhibited, including: (1) a basic mode that corresponds to a resting potential and production of action potential; (2) bistability between resting potential and self-sustained spiking; (3) a pacemaker mode of discharge; and (4) bistability between resting potential and plateau potential. These behaviours can be compared to experimentally described physiological and pathological forms of excitability that depend upon inward persistent conductances. In the results obtained, attractors allow for a qualitative description of physiological and pathological states. However, it is not possible to obtain an unambiguous identification of particular 'physiological attractors' or 'pathological attractors'. In the perspective of the theory of dynamical systems, we suggest that pathological states can be modelled in two different ways, i.e. by bifurcation (as in the present model) or by perturbation. We also highlight some other theoretical concepts that may be relevant to a theoretical description of pathology.

Phosphorothioate oligonucleotides derived from human immunodeficiency virus type 1 (HIV-1) primer tRNALys3 are strong inhibitors of HIV-1 reverse transcriptase and arrest viral replication in infected cells.

Retroviral reverse transcriptase (RT) is involved in the selection of a specific tRNA primer which initiates proviral DNA minus-strand synthesis. Studies of the interactions between human immunodeficiency virus type 1 (HIV-1) RT and primer tRNALys3 have shown that the dihydrouridine (diHU), anticodon, and pseudouridine regions of tRNA are highly protected in the RT-tRNA complex. The CCA 3' end of tRNA is also in close contact with the enzyme during the cDNA initiation step. Using synthetic oligoribonucleotides corresponding to the anticodon and diHU regions, we have previously shown a low but significant inhibition of HIV-1 RT activity. We extend this observation and show that primer tRNA-derived oligodeoxynucleotides (ODNs) carrying a phosphorothioate (PS) modification are strong inhibitors of HIV-1 RT. The affinity of PS-ODNs for the enzyme was monitored by gel mobility shift electrophoresis. Experiments with HIV-1-infected human cells (MT-2 cells) were performed with the latter ODNs. A PS-ODN corresponding to the 3' end of tRNALys3 (acceptor stem [AS]) was able to inhibit HIV-1 replication. No effect of the other modified ODNs was observed in infected cells. The analysis of HIV-1 RNase H activity in a cell-free system strongly suggests that the inhibitory effect of the PS-AS may be mediated via both a sense and an antisense mechanism.

Bistable behaviour in a neocortical neurone model.

Intracellular recordings have shown that neocortical pyramidal neurones have an intrinsic capacity for regenerative firing. The cellular mechanism of this firing was investigated by computer simulations of a model neurone endowed with standard action potential and persistent sodium (gNaP) conductances. The firing mode of the neurone was determined as a function of leakage and NaP maximal conductances (gl and gNaP). The neurone had two stable states of activity (bistable) over wide range of gl and gNaP, one at the resting potential and the other in a regenerative firing mode, that could be triggered by a transient input. This model points to a cellular mechanism that may contribute to the generation and maintenance of long-lasting sustained neuronal discharges in the cerebral cortex.

Detection of t(2;5)(p23;q35) translocation by reverse transcriptase polymerase chain reaction and in situ hybridization in CD30-positive primary cutaneous lymphoma and lymphomatoid papulosis.

The t(2;5) generates a chimeric NPM-ALK transcript encoded by the nucleophosmin NPM gene fused to the anaplastic lymphoma kinase gene ALK. Using a reverse transcriptase nested polymerase chain reaction assay we have detected NPM-ALK transcripts within CD30+ primary cutaneous lymphoma and lymphomatoid papulosis (LP). The t(2;5) was identified in 4 out of 9 CD30+ anaplastic lymphomas and in 1 out of 4 CD30+ pleomorphic lymphomas. Moreover, the t(2;5) was detected in 3 out of 10 LPs. All NPM-ALK-positive lymphomas and 1 NPM-ALK-positive LP exhibited a clonal rearrangement of the T cell receptor gamma-chain gene. The t(2;5) was detected in 2 cases of LP without other evidence for a clonal lymphoid population. To identify cells carrying the t(2;5) translocation, we used immunohistochemistry to detect the ALK-encoded p80 protein and in situ hybridization for the specific detection of NPM-ALK transcripts. Both p80 protein and NPM-ALK transcripts were expressed by anaplastic or large CD30+ lymphoma cells with positive NPM-ALK amplification. The presence of t(2;5) in a subset of CD30+ cutaneous lymphoma and LP may indicate a common pathogenesis with a subset of anaplastic nodal lymphoma.

Human immunodeficiency virus type 1 replication within cystic lymphoepithelial lesion of the salivary gland.

Cystic lymphoepithelial lesions of salivary glands (CLLSG) are nodular or diffuse salivary gland enlargements that are observed in patients who tested positive for human immunodeficiency virus type 1 (HIV-1). Two cases of CLLSG are reported. Particular emphasis is placed on the presence of HIV-1 major-core protein (P24), HIV-1 RNA sequences, Epstein-Barr virus (EBV) DNA sequences, and lymphocyte receptor gene rearrangement. Lymphoid alterations consisted of explosive hyperplasia with a prominent follicular reticular dendritic cell (DRC) network and numerous intrafollicular CD8+ lymphocytes. Intrafollicular DRC strongly expressed HIV-1 major-core protein and HIV-1 RNA, indicating that most DRCs actively replicated the HIV-1 virus. The presence of active HIV-1 replication within DRC and the absence of clonal EBV infected lymphoid population strongly suggest that CLLSG pathogenesis is primarily induced by HIV-1. The presence of oligoclonal immunoglobulin gene rearrangements in our cases, however, suggest the need of long-term follow-up of such patients to determine whether CLLSG could be a benign prelymphomatous disease.

In vitro infection of human macrophages with human T-cell leukemia virus type 1.

The tropism of the human T-cell leukemia virus type 1 (HTLV-1) for the cells of monocyte-macrophage lineage was evaluated by the coculture of blood monocyte-derived macrophages, with irradiated cells of HTLV-1 producing cell lines MT2 or C91/PL. The susceptibility to HTLV-1 was assessed by the detection of viral DNA using the polymerase chain reaction method. HTLV-1 gene expression in the cells was detected using in situ hybridization and by immunofluorescent staining of viral antigen. The presence of type C virus-like particles detected by electron microscopy and the ability to infect normal cord blood lymphocytes demonstrated that the infected macrophages produced infectious virus. These results indicate that human macrophages are susceptible in vitro to productive HTLV-1 infection, and thus might be involved in the pathogenesis of HTLV-1-related diseases.

Analysis of HIV-1 expression in vivo with in situ hybridization and the polymerase chain reaction.

The objective of the present study was to compare the data of in situ hybridization (ISH), RNA polymerase chain reaction (PCR/RNA) and p24 core antigen (p24 Ag) enzyme immunoassay (EIA) for the detection of HIV-1 expression in peripheral blood mononuclear cells (PBMCs) and in plasma of infected patients at various CDC stages. PBMCs of 24 patients mostly of CDC stage II were obtained from heparinized blood samples, cytocentrifuged and hybridized with a (35S) labelled single-stranded RNA probe specific for gag-pol of LAVBru HIV-1 allowing the detection of genomic and/or messenger RNA. The corresponding plasma samples were used for the determination of p24 Ag by EIA and detection of HIV-1 genomic RNA by RT-PCR using specific primers in the LTR, gag and env regions. Whereas p24 was detected in only six out of 24 patients, both ISH and PCR/RNA enabled the detection of viral RNAs in more than 60% of the patients; cumulation of positive results of ISH and RT-PCR showed that 100% of patients at stage IV and 83% of patients at stages II/III have molecular signs of HIV expression therefore indicating that transcription of the provirus is a highly frequent event, even in the early stages of the disease, and, pleading for undertaking a very early antiviral chemotherapy.

In vitro assays show a dissociation of reverse transcriptase activity and core antigen (p24) production in two HIV-1 isolates from a patient receiving long-term treatment with zidovudine (ZDV).

Two HIV-1 isolates were obtained from a patient receiving long-term treatment with zidovudine (ZDV). The in vitro sensitivity to ZDV triphosphate of the reverse transcriptase (RT) from both isolates appeared to be unchanged compared to that of the LAV-Bru HIV-1 reference strain. When isolates were grown in CEM cells (a T-lymphoblastoid tumor cell line) and their RT activity and core antigen (p24) production were determined, the level of p24 production compared to RT activity was high; in infected CEM cells treated with ZDV, RT activity was at background level while the p24 production was still significant, thus indicating a dissociation of RT activity and core antigen production.

Quantitative in situ hybridization using strand specific RNA probes: expression of the bunyavirus Germiston S segment in mosquito cells.

Infection of Vero (monkey) cells by Germiston bunyavirus is highly cytopathic with cell lysis and virus production at a high titre, whereas infection of Aedes albopictus C6/36 (mosquito) cells leads, after an acute primary phase, to a persistent non-cytopathic infection with a loss in virus production. In this report we demonstrate that single-stranded RNA probes can be successfully used in an in situ hybridization assay to quantify viral expression during this persistent infection. The steady-state levels of viral S-RNA segment (genomic and messenger sense) during the acute phase were similar to those observed in lytically infected Vero cells, but appeared delayed. Both senses of S-RNA were detected throughout persistent infection but in lower amounts, in less than 10% of the cells and always in the cytoplasm of infected cells. The number of copies per cell of messenger sense S-RNAs remained low during persistent infection whereas a higher fluctuation was observed for genomic S-RNAs. In situ hybridization with specific stranded RNA probes provides both qualitative and quantitative informations, that can lead to a better understanding of virus-cell interactions.

Detection of the bunyavirus Germiston in VERO and Aedes albopictus C6/36 cells by in situ hybridization using cDNA and asymmetric RNA probes.

Using Germiston virus infected vertebrate (VERO) and invertebrate (Aedes albopictus C6/36) cells, paraformaldehyde-glutaraldehyde fixative allowed the best preservation of cellular morphology and the highest hybridization signals with cDNA and asymmetric RNA probes against the viral S segment. Asymmetric RNA probes always gave higher sensitivity and better specificity of in situ hybridization than the nick-translated symmetric DNA probe in both vertebrate and invertebrate cells. The study of Aedes albopictus C6/36 cells persistently infected with Germiston virus showed that only a small number of cells contained the S segment, and that the replication and transcription of the S segment took place in the cytoplasm of acutely and persistently infected cells.