Analysis of optical properties of the mouse cranium--implications for in vivo multi photon laser scanning microscopy.

Trans-cranial imaging is the least invasive method for optical in vivo studies of structures in the mouse brain and has found wide application over the last few years. An important issue is how and to what extent the cranium and the tissue between the cranium and the focal point detract from the quality of the recorded images. Here we address this issue by recording transmission images in wild type mice at five wavelengths in the visible and near-infrared spectrum. The recorded laser scanning microscopic images were analyzed pixel by pixel in order to quantify the light attenuation and shading as function of the location of the focal point relative to the cranium. Additional images demonstrate the effects of the mouse crania on the images of fluorescent microspheres in the low micrometer range. The results of this study demonstrate that light attenuation by the cranium, though with typical losses of less than 20% of the incident light, induces shading effects during the imaging process. Geometrical shapes and sizes in the images of the recorded objects may differ substantially depending on whether they have been recorded trans-cranially or not. This is true even for comparatively large structures such as cell somata. Our results call for a more realistic appraisal of the potential of the trans-cranial imaging approach, particularly when it comes to absolute measurements of sizes and shapes of small objects. As trans-cranial imaging has found wide use in contemporary research it is important that the results be interpreted with due caution.

Water entry into astrocytes during brain edema formation.

The process of brain edema formation has been studied extensively at the macroscopic level. In contrast, little is known about water fluxes and volume changes at the cellular level in the initial phase of brain edema. Insight in these "microscopic" events could pave the way for more efficient prevention and therapy. Here, we report measurements of brain cell volume responses recorded in vivo in a model of systemic hyponatremia. Transgenic mice expressing fluorescent proteins in astrocytes were subjected to hypo-osmotic stress and two photon laser scanning microscopy. Volume measurements of glial cells in the cerebellum and the visual cortex indicate that individual astrocytes undergo a position-dependent increase in cell volume by a factor of two or more during edema formation. Our data are the first to show that volume changes can be monitored at the cellular level in vivo and demonstrate that astrocytes are sites of water entry in the initial phase of brain edema formation. The uptake of water in astrocytes is likely to reflect the strong expression of aquaporin-4 in these cells.

Features of neuronal synchrony in mouse visual cortex.

Synchronization of neuronal discharges has been hypothesized to play a role in defining cell assemblies representing particular constellations of stimulus features. In many systems and species, synchronization is accompanied by an oscillatory response modulation at frequencies in the gamma-band. The cellular mechanisms underlying these phenomena of synchronization and oscillatory patterning have been studied mainly in vitro due to the difficulty in designing a direct in vivo assay. With the prospect of using conditional genetic manipulations of cortical network components, our objective was to test whether the mouse would meet the criteria to provide a model system for the study of gamma-band synchrony. Multi-unit and local field potential recordings were made from the primary visual cortex of anesthetized C57BL/6J mice. Neuronal responses evoked by moving gratings, bars, and random dot patterns were analyzed with respect to neuronal synchrony and temporal patterning. Oscillations at gamma-frequencies were readily evoked with all types of stimuli used. Oscillation and synchronization strength were largest for gratings and decreased when the noise level was increased in random-dot patterns. The center peak width of cross-correlograms was smallest for bars and increased with noise, yielding a significant difference between coherent random dot patterns versus patterns with 70% noise. Field potential analysis typically revealed increases of power in the gamma-band during response periods. Our findings are compatible with a role for neuronal synchrony in mediating perceptual binding and suggest the usefulness of the mouse model for testing hypotheses concerning both the mechanisms and the functional role of temporal patterning.

Perceptual learning of highly demanding visual search tasks.

Inefficient visual search can become efficient with practice [Vision Research 35 (1995) 2037; 40 (2000) 2925]. In this study, we wondered whether this improvement depends on unique visual features associated with the target, on differences in item-specific brightness distribution between target and distractors, or only on a change in the allocation of attention and thus global search strategy. We found that both, unique visual features and differences in brightness distribution lead to parallelisation with practice of originally inefficient search. Prolonged practice of inefficient search tasks lacking both unique visual features and differences in brightness distribution (conjunctions) does not lead to improved performance, thus indicating that perceptual learning in visual search does not solely reflect an unspecific global improvement in search strategy. Changing the brightness polarity of the stimuli leads to instantaneous, complete transfer to the new task. There is no transfer but rather trade-off between the learning based on unique visual features or on differences in brightness distribution between target and distractors.