Calcium signals and FGF-2 induced neurite growth in cultured parasympathetic neurons: spatial localization and mechanisms of activation.

The growth of neuritic processes in developing neurons is tightly controlled by a wide set of extracellular cues that act by initiating downstream signaling cascades, where calcium signals play a major role. Here we analyze the calcium dependence of the neurite growth promoted by basic fibroblast growth factor (bFGF or FGF-2) in chick embryonic ciliary ganglion neurons, taking advantage of dissociated, organotypic, and compartmentalized cultures. We report that signals at both the growth cone and the soma are involved in the promotion of neurite growth by the factor. Blocking calcium influx through L- and N-type voltage-dependent calcium channels and transient receptor potential canonical (TRPC) channels reduces, while release from intracellular stores does not significantly affect, the growth of neuritic processes. Simultaneous recordings of calcium signals elicited by FGF-2 at the soma and at the growth cone show that the factor activates different patterns of responses in the two compartments: steady and sustained responses at the former, oscillations at the latter. At the soma, both voltage-dependent channel and TRPC blockers strongly affect steady-state levels. At the growth cone, the changes in the oscillatory pattern are more complex; therefore, we used a tool based on wavelet analysis to obtain a quantitative evaluation of the effects of the two classes of blockers. We report that the oscillatory behavior at the growth cone is dramatically affected by all the blockers, pointing to a role for calcium influx through the two classes of channels in the generation of signals at the leading edge of the elongating neurites.

SiO2 nanoparticles modulate the electrical activity of neuroendocrine cells without exerting genomic effects.

Engineered silica nanoparticles (NPs) have attracted increasing interest in several applications, and particularly in the field of nanomedicine, thanks to the high biocompatibility of this material. For their optimal and controlled use, the understanding of the mechanisms elicited by their interaction with the biological target is a prerequisite, especially when dealing with cells particularly vulnerable to environmental stimuli like neurons. Here we have combined different electrophysiological approaches (both at the single cell and at the population level) with a genomic screening in order to analyze, in GT1-7 neuroendocrine cells, the impact of SiO2 NPs (50 ± 3 nm in diameter) on electrical activity and gene expression, providing a detailed analysis of the impact of a nanoparticle on neuronal excitability. We find that 20 µg mL-1 NPs induce depolarization of the membrane potential, with a modulation of the firing of action potentials. Recordings of electrical activity with multielectrode arrays provide further evidence that the NPs evoke a temporary increase in firing frequency, without affecting the functional behavior on a time scale of hours. Finally, NPs incubation up to 24 hours does not induce any change in gene expression.

A simple method to study cellular migration.

We describe here a simple and fast method for the characterisation of cell motion. By projecting on a single plane different positions of the cell a ribbon is generated, whose characteristics can be related to the type of motion. The proposed method allows both to determine, very quickly, the motility of a population of cells and to investigate and characterise properties of a single cell's motion. The methodology presented here can be applied to a large range of cell movement and also adapted and extended to other problems involving biological motion.

In vitro identification of dividing neuronal precursors from chick embryonic ciliary ganglion.

In chick parasympathetic ciliary ganglion the neuronal birthdate is well defined, between 2.5 and 5.5 days of embryonic development, and neuronal precursor cells that are able to differentiate into neurons in vitro can be isolated from E4.5 ganglia. In this report, using bromodeoxyuridine incorporation and Maplb immunostaining, we demonstrate that these cells can be isolated from E7-E8 chick embryos as well, suggesting that neuronal precursor cells are still present in the ciliary ganglion after the end of the in vivo neurogenesis. These precursor cells retain the ability to divide and generate newly differentiated neurons in vitro when cultured in a chemically defined medium. Such a capacity is highly stimulated by bFGF but not by CNTF.