Immersive virtual environment for visuo-vestibular therapy: preliminary results.

The sense of equilibrium aggregates several interacting cues. On vestibular areflexic patients, vision plays a major role. We developed an immersive therapeutic platform, based on 3D opto-kinetic stimulation that enables to tune the difficulty of the balance task by managing the type of optic flow and its speed. The balance adjustments are recorded by a force plate, quantified by the length of the center of pressure trajectory and detection of disequilibrium corrections (leans, compensation step). Preliminary analysis shows that (i) patients report a strong immersion feeling in the motion flow, triggering intense motor response to "fight against fall"; (ii) the ANOVA factorial design shows a significant effect of flow speed, session number and gaze anchor impact. In conclusion, this study shows that 3D immersive stimulation removes essential limits of traditional opto-kinetic stimulators (limited 2D motions and remaining fixed background cues). Moreover, the immersive optic flow stimulation is an efficient tool to induce balance adaptive reactions in vestibular patients. Hence, such a platform appears to be a powerful therapeutic tool for training and relearning of balance control processes.

Statistical analysis and parsimonious modelling of dendrograms of in vitro neurones.

The processes whereby developing neurones acquire morphological features that are common to entire populations (thereby allowing the definition of neuronal types) are still poorly understood. A mathematical model of neuronal arborizations may be useful to extract basic parameters or organization rules, hence helping to achieve a better understanding of the underlying growth processes. We present a parsimonious statistical model, intended to describe the topological organization of neuritic arborizations with a minimal number of parameters. It is based on a probability of splitting which depends only on the centrifugal order of segments. We compare the predictions made by the model of several topological properties of neurones with the corresponding actual values measured on a sample of honeybee (olfactory) antennal lobe neurones grown in primary culture, described in a previous study. The comparison is performed for three populations of segments corresponding to three neuronal morphological types previously identified and described in this sample. We show that simple assumptions together with the knowledge of a very small number of parameters allow the topological reconstruction of representative (bi-dimensional) biological neurones. We discuss the biological significance (in terms of possible factors involved in the determinism of neuronal types) of both common properties and cell-type specific features, observed on the neurones and predicted by the model.

Two novel monoclonal antibodies (1.9.E and 4.11.C) against olfactory bulb ensheathing glia.

We produced and characterized two monoclonal antibodies, termed 1.9.E and 4.11.C, that specifically recognize olfactory bulb ensheathing glia. Both antibodies were generated using the olfactory nerve layer (ONL) of newborn rat olfactory bulbs (P0, P1) as immunogens. The specificity of these antibodies was tested by immunofluorescence techniques on tissue sections and cultures of adult and neonatal rat olfactory bulbs, and by Western blot analysis. 1.9.E labeled the ONL and glomerular layer of the olfactory bulb (OB) of adult rats. In newborn rats, 1.9.E immunostained ensheathing cells from the ONL and peripheral olfactory fascicles. Furthermore, 1.9.E reacted with some processes of the radial glia in the periventricular germinal layer of the newborn rat. Although 4.11.C also specifically labeled ensheathing cells in the adult OB, it did not stain any cell type in the ONL of newborn rats. The lack of double labeling with either 1.9.E or 4.11.C and anti-olfactory marker protein (OMP) antibody, a specific marker for olfactory axons, indicated that none of the monoclonals recognized olfactory axons. Double immunostaining of adult OB cultures with 1.9.E or 4.11.C and anti-p75-nerve growth factor receptor revealed that both antibodies specifically recognized ensheathing glia in those cultures. Filaments were strongly labeled throughout the entire cytoplasm of ensheathing cells, suggesting that 1.9.E and 4.11.C immunoreacted with ensheathing glia cytoskeleton. 4.11.C stained a few Schwann cells in adult sciatic nerve sections. Moreover, 4.11.C immunostained cortical astrocyte cultures from newborn rats (P1). In Western blot analysis both antibodies recognized a major component, migrating with an apparent molecular weight of 60 kDa, from olfactory nerve and glomerular layer (ONGL) extracts of adult and neonatal rats. The pattern of immunoreactivity of 1.9.E and 4.11.C antibodies suggest that both antibodies are specific markers for olfactory ensheathing glia in the adult rat central nervous system (CNS).

Expression of the surface antigen A2B7 in adult and developing honeybee olfactory pathway.

In order to identify molecules involved in the development of the honeybee olfactory pathway, hybridoma technology has been used. Among different cell lines, A2B7 has been selected. It produces a specific antibody for a surface glycoprotein of 91 kDa. This protein is mainly expressed by both the antennal receptor cells and mushroom body neurons. Based on (i) the spatio-temporal pattern of expression during pupal development; (ii) the cell surface location of the antigen; and (iii) the partial molecular characterization of the antigen, a putative role for this protein in axonal fasciculation and guidance is discussed.

Brain FOS-like expression in developing and adult honeybees.

This study was undertaken to examine the expression of the protooncogene c-fos in an invertebrate, the honeybee. We investigated first the expression of Fos-like proteins, testing different methodologies (immunohistochemistry, immunocytochemistry, immunoblotting). These studies were carried out at the embryonic, nymphal and adult stages. We found immunoreactive bands at approximately 30 kD and 57 kD. The first ones are revealed at the embryonic, nymphal and adult stages. The approximately 57 kD proteins are present at the adult stage. Beside this qualitative variation, intensive modification occurs with a burst of protein expression a few days after the insect emergence. The results suggest that Fos-like proteins are found in the developing and maturing nervous system. Basal level of expression is found in the olfactory pathway, convincing to explore the protooncogene c-fos expression in the cellular and molecular analyses of olfactory plasticity.

A morphometric classification of pupal honeybee antennal lobe neurones in culture.

Both intrinsic (programmed) and extrinsic (non-programmed) factors are thought to play a role in the morphogenesis of neurones in the honeybee antennal lobe (the first relay station in the olfactory pathway) during development. We present here a morphometric and statistical analysis of a large population of pupal honeybee antennal lobe neurones grown in primary culture. Quantitative parameters were used to characterize neuronal shapes. On the basis of such morphometric measurements, an automatic classification allowed three morphological types to be distinguished. Their main characteristics are presented and the role of intrinsic and extrinsic factors in their determination is discussed.

Morphological analysis of honeybee antennal cells growing in primary cultures.

A culture technique for the in vitro growth of antennal cells from honeybee is described. On the basis of morphological and immunocytochemical criteria, the cultured cells could be classified into neural and non-neural cells. Neural cells (type D) exhibited the main morphological features of insect olfactory receptor neurones (ORNs). Non-neural cells were large, flat cells that could be divided into three main types: Type A, B and C cells. Type A cells were spindle-like cells and resembled insect myocytes in culture. Type B cells were large cells with a veil-like cytoplasm. These cells tended to group and vacuolate towards the center of the cellular aggregate. Type C cells were either bipolar (Type C1) or multipolar (Type C2) flat cells which closely resembled insect glial cells in cultures.

Specification and implementation of a digital Hopfield-type associative memory with on-chip training.

The definition of the requirements for the design of a neural network associative memory, with on-chip training, in standard digital CMOS technology is addressed. Various learning rules that can be integrated in silicon and the associative memory properties of the resulting networks are investigated. The relationships between the architecture of the circuit and the learning rule are studied in order to minimize the extra circuitry required for the implementation of training. A 64-neuron associative memory with on-chip training has been manufactured, and its future extensions are outlined. Beyond the application to the specific circuit described, the general methodology for determining the accuracy requirements can be applied to other circuits and to other autoassociative memory architectures.

Developmental study of afferented and deafferented bee antennal lobes.

The role of antennal sensory projections on the ontogeny of the bee antennal lobe was analyzed using both light and transmission electron microscopy. Normal and deafferented developing antennal lobes were examined. The results obtained show that (1) initiation of synaptogenesis in the antennal lobe is independent of the arrival of sensory inputs; (2) sensory inputs are necessary for setting up the glomerular antennal lobe organization; (3) regressive events, such as the reduction of synapse density, occur during the development of the antennal lobe; and (4) glomeruli formation appears as related to glia development.

A quantitative ultrastructural study of the honeybee antennal lobe.

This paper describes the ultrastructural organization of the honeybee antennal lobe, including the distribution of synapses within the antennal lobe neuropile and the distribution of the afferent fibres in the antennal nerve and its afferent tracts. We show that: 1) The antennal nerve and tracts T3-T6 are composed of a heterogeneous population of fibres, with respect to their diameters, whereas two afferent tracts (T1 and T2) are composed of fibres of almost homogeneous diameter. 2) Synapses are mainly localized in the glomeruli with a higher frequency in the cortical layer than in the core of the glomerulus. Nevertheless a few synapses are found in the coarse neuropile. 3) Reciprocal synapses have been identified in the cortical layer. At the ultrastructural level, the organization of the bee antennal lobe was largely unknown and these results bring the anatomical background needed in order to carry out a developmental study related to the bee antennal lobe structures.

The morphology and ultrastructure of antennal lobe cells from pupal honeybees (Apis mellifera) growing in culture.

A culture technique for the in vitro growth and differentiation of antennal lobe cells from the honeybee, Apis mellifera, is described and the ultrastructure of the growing cells is analysed. Two types of cell are present in the cultures and from their morphology and ultrastructure they can be identified as glial cells and neurones. The neurones have a granular cytoplasm, abundant endoplasmic reticulum and a small, densely stained nucleus. They produce long processes with varicosities that contain dense-core and clear vesicles. In contrast the glial cells have clear cytoplasm, little endoplasmic reticulum and a distinct cytoskeletal organisation. These cells produce short, flat processes that spread over the surface of the culture dish. Although a number of cell contacts have been identified in the cultures no synapses have yet been seen. These cultures provide a good in vitro model for an analysis of the interactions between cells derived from the antennal lobe of the honey bee.