Multi-capillary column-ion mobility spectrometer (MCC-IMS) breath analysis in ventilated rats: a model with the feasibility of long-term measurements.

Rats are commonly used in medical research as they enable a high grade of standardization. The exhalome of ventilated rats has not as yet been investigated using an ion mobility spectrometer coupled with a multi-capillary column (MCC-IMS). As a first step, a rat model has to be established to measure potential biomarkers in the exhale with long-term settings, allowing constant and continuous analysis of exhaled air in time series. Therefore, eight animals were anaesthetized, prepared and ventilated for 1 h. A total of 73 peaks were directly detected with the IMS chromatogram. Thirty five of them were assigned to the ventilator system and 38 to the animals. Peak intensity varied within three measurements. The intensity of analytes of individual rats varied by a factor of up to 18. This new model will also enable continuous measurements of volatile organic compounds (VOCs) from rat's breath in long-term experiments. It is hoped that, in the future, variability and progression of VOCs can be monitored in different models of diseases using this set-up.

Neuroneuronal interconnections in the rat superior cervical ganglion; possible anatomical bases for modulatory interactions revealed by intracellular horseradish peroxidase labelling.

Electrophysiologically identified neurons of rat superior cervical ganglion were intracellularly injected with horseradish peroxidase and processed for light and electron microscopic observation. At light microscope level, neurons could be classified according to their dendritic arborization pattern in the vicinity of the soma into radiate, tufted and intermediate types. Upon electrical stimulation of the internal and external carotid nerves it was observed that radiate and intermediate neurons sent their axons into one or the other of these nerve trunks, whereas a majority of tufted neurons gave no response to stimulation of either of these postganglionic nerves. Electron microscopic exploration of horseradish peroxidase-labelled neurons revealed a surprisingly high prevalence of interconnectivity between ganglionic neurons. These contacts were both dendrosomatic and dendrodendritic, and were a universal feature of the labelled neurons explored. Twenty-two of the 23 labelled cells were found to receive direct dendritic appositions on their somata, and 13 of these 23 cells were seen each to send their dendrites into contact with at least one unlabelled neuronal soma. Dendrodendritic contacts were observed for 87% of the labelled neurons, and most of the cells (80%) were seen to form triadic contacts which included two dendrites and a preganglionic nerve ending. All these figures represent minimum incidences. None of the dendrosomatic or dendrodendritic appositions observed was overtly synaptic although several morphological features indicated the possibility of somatic and or dendritic release and uptake at sites of apposition. It is suggested that the observed appositions provide anatomical substrates for modulatory interactions between the ganglionic neurons, possibly involving slow potentials or the switching of metabolic pathways.

Fibronectin in the area opaca of the young chick embryo. Immunofluorescence and immuno-electron-microscopic study.

Distribution of fibronectin-like immunoreactivity was studied in the area opaca of the young chick embryo (stages 4-6 HH) by use of the immunofluorescence and protein A-coupled to colloidal gold techniques. Fibronectin, associated to the basement membrane, formed a fibrillar network, the pattern of which changed from the centre to the periphery of the area opaca. At the ultrastructural level, differences in fibronectin distribution were found between non-moving and moving cells. The epithelial-like cells presented fibronectin staining exclusively on their basal side. Actively migrating cells (edge and mesodermal cells) showed immunoreactive material localized around their entire surface and within the cytoplasm. The fibronectin distribution is discussed in relation to three important phenomena taking place during the early growth of the area opaca: anchorage and migration of the edge cells, modification of cell shape in relation to mechanical tension, and expansion of the area vasculosa.

Serum-free aggregate cultures of rat CNS glial cells: biochemical, immunocytochemical and morphological characterization.

Aggregate cultures of mixed glial cells, as well as of enriched astrocytes and oligodendrocytes were prepared, and maintained in serum-free medium for up to 25 days. Biochemical measurements of both neuron-specific and glia-specific enzyme activities showed that these three types of aggregate cultures were virtually devoid of neurons. Astrocyte-enriched cultures were greater than 95% pure, with oligodendrocytes as the only apparent contaminant, whereas oligodendrocyte-enriched cultures still contained a considerable proportion of astrocytes. In all these neuron-free aggregate cultures both astrocytes and oligodendrocytes attained a high degree of maturation. These findings were confirmed by morphological examinations, and by immunofluorescence studies. Furthermore, ultrastructural as well as immunocytochemical investigations using antibodies to myelin basic protein revealed that all three types of glial cell aggregate cultures contained myelin membranes, indicating that the presence of axons is not a prerequisite for the formation of myelin.

Growth and differentiation of aggregating fetal brain cells in a serum-free defined medium.

Aggregating cultures of mechanically dissociated fetal brain cells provide an excellent system for neurobiological studies of cellular growth and differentiation, but, in common with almost all culture systems, they have the disadvantage that crude serum is required in the medium. Although several cell lines have either been adapted to serum-free conditions or grown normally in serum-free media supplemented with hormones, trace elements and defined serum components, this approach has never been applied to differentiating primary cells of the central nervous system. We now describe the successful cultivation of aggregating fetal rat brain cells in a chemically defined, serum-free medium.