Abstract
Calcium signals play a major role in the control of all key stages of neuronal development, and in particular in the growth and orientation of neuritic processes. These signals are characterized by high spatial compartmentalization, a property which has a strong relevance in the different roles of specific neuronal regions in information coding. In this context it is therefore important to understand the structural and functional basis of this spatial compartmentalization, and in particular whether the behavior at each compartment is merely a consequence of its specific geometry or the result of the spatial segregation of specific calcium influx/efflux mechanisms. Here we have developed a novel approach to separate geometrical from functional differences, regardless on the assumptions on the actual mechanisms involved in the generation of calcium signals. First, spatial indices are derived with a wavelet-theoretic approach which define a measure of the oscillations of cytosolic calcium concentration in specific regions of interests (ROIs) along a cell, in our case developing chick ciliary ganglion neurons. The resulting spatial profile demonstrates clearly that different ROIs along the neuron are characterized by specific patterns of calcium oscillations. Next we have investigated whether this inhomogeneity is due just to geometrical factors, namely the surface to volume ratio in the different subcompartments (e.g. soma vs. growth cone) or it depends on their specific biophysical properties. To this aim correlation functions are computed between the activity indices and the surface/volume ratio along the cell: the data thus obtained are validated by a statistical analysis on a dataset of different cells. This analysis shows that whereas in the soma calcium dynamics is highly correlated to the surface/volume ratio, correlations drop in the growth cone-neurite region, suggesting that in this latter case the key factor is the expression of specific mechanisms controlling calcium influx/efflux.
Highlights
The growth, orientation and specification of neuritic processes from developing neurons is a key event in the formation of the correct connectivity of the nervous system and is tightly regulated by a wide set of signalling mechanisms, among which complex spatiotemporal concentration, ÂpCaatt2ezrnÃsi, of changes in cytosolic free calcium play a major role [1]
We have tried to address this problem by simultaneously recording spontaneous calcium signalling activity from the soma, neurites and growth cones of E7 chick ciliary ganglion (CG) neurons in culture, and by generalizing a wavelet-based analytical approach described in a previous paper [16] to perform a spatial analysis of the oscillatory activity during a defined time interval in the different compartments of the neuron
Surface to Volume Ratio Assessment We have developed a simple approach to estimate cell thickness lx under the surface of the single regions of interests (ROIs) as a function of space x, using the same ROI parametrization already employed for the computation of Jx
Summary
The growth, orientation and specification of neuritic processes from developing neurons is a key event in the formation of the correct connectivity of the nervous system and is tightly regulated by a wide set of signalling mechanisms, among which complex spatiotemporal concentration, ÂpCaatt2ezrnÃsi , of changes in cytosolic free calcium play a major role [1]. The differences between signals at the soma and at thin peripheral compartments (such as filopodia of neurites and pre- and post-synaptic regions) have been ascribed, in most cases, to differences in the surface to volume ratio [11,12,13], but a contribution from spatial specificity in the calcium mobilizing mechanisms, based on the different distribution of channels (and/ or transporters), may be involved [14,15]. We have tried to address this problem by simultaneously recording spontaneous calcium signalling activity from the soma, neurites and growth cones of E7 chick ciliary ganglion (CG) neurons in culture, and by generalizing a wavelet-based analytical approach described in a previous paper [16] to perform a spatial analysis of the oscillatory activity during a defined time interval in the different compartments of the neuron. The aim was to correlate the differences in oscillatory activity as a function of space with the estimated surface to volume ratio, in order to uncover any specificity of the different compartments in terms of calcium mobilizing mechanisms
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