A FRET Analysis of Rho-A Activity During the Establishment of Polarity in Cultured Hippocampal Neurons

WOJNACKI JOSE; VON BILDERLING CATALINA; QUASSOLLO GONZALO; GASTALDI LAURA; PIETRASANTA LIA ; CACERES ALFREDO

Huerta Grande

Congreso; Reunión Anual de la Sociedad Argentina de Neurociencia; 2011

Sociedad Argentina de Neurociencia

Studies over the last decade on the establishment of neuronal polarity have implicated the small GTPase Rho-A as a key regulator of this process. In particular it has been shown that over-expression of Rho-A halts neuronal polarization on hippocampal neurons whereas expression of dominant-negative Rho-A results in multiple axon formation (Da Silva et al., 2003; Chuang et al., 2005; Conde and Caceres, 2009). Stressing the importance of this protein, our laboratory has recently shown that the Rho-A activator Lfc has a negative impact on the development of polarity (Conde et al, 2010). These observations clearly suggest that inhibition of Rho-A activity is required for axon formation and neuronal polarization. All these studies, and related ones, have assumed that Rho-A inhibition should take placeat the growth cone of the future axon, but this has never been proved so far. Using a Rho-A biosensor and a FRET approach we have now obtained evidence about the spatial regulation of Rho-A activity during polarization. The results obtained provide evidence suggesting that a decreased Rho-A activity spatially localized to the axon parallels neuronal polarization. This event appears to be related with increased microtubule stabilization since low doses of taxol, that induce multiple axon formation are also paralleled by decreased Rho-activity along axons shafts and tips.