ORIGINAL PAPER Compartmentalization of the redox environment in PC-12 neuronal cells G. Maulucci Æ G. Pani Æ S. Fusco Æ M. Papi Æ G. Arcovito Æ T. Galeotti Æ M. Fraziano Æ M. De Spirito Received: 14 February 2009 / Revised: 20 April 2009 / Accepted: 29 April 2009 / Published online: 3 June 2009 Ó European Biophysical Societies’ Association 2009 Abstract Neuronal redox phenomena are involved in numerous biochemical pathways and play a key role in many pathological events and clinical situations. The oxi- dation/reduction (redox) state present in biological com- partments is a major target for possible pharmaceutical intervention and, consequently, the processes associated with its change have attracted increased attention in recent years. Here, we analyze the redox environment and its spatial compartmentalization in differentiated neuronal phenotype of PC-12 cells using a redox-sensitive protein (i.e., a mutant of the Yellow Fluorescent protein), employed ratiometrically. Redox maps of cells were gen- erated with an elevate spatial resolution, and the spatial distributions of highly oxidized and highly reduced regions have been determined. A quantitative analysis of redox maps allows the disclosure of a peculiar spatial organiza- tion of the redox environment. Keywords ROS  Neuron  Redox environment  rxYFP  Compartmentalization Introduction Reactive oxygen species (ROS), such as superoxide (O 2 - ) and hydrogen peroxide (H 2 O 2 ), are produced intracellularly as part of normal metabolic reactions (Babior 2002; Vignais 2002). ROS are very reactive oxidants (Liochev 1996; Turrens 2003) and their excessive, uncontrolled production can have detrimental effects on cellular physi- ology and function, often leading to apoptosis and to a variety of diseases (Finkel 2003). Recent studies have suggested that elevated, but sub- lethal, levels of (O 2 - ) and H 2 O 2 can act to influence intra- cellular signaling pathways in cells by modulating gene expression, cellular growth, and differentiation (Droge 2002; Finkel 1998; Hancock et al. 2001; Kamata and Hirata 1999; Klann and Thiels 1999; Rhee 1999). ROS have been shown to be essential for the NGF- induced differentiation of PC-12 cells (Katoh et al. 1997, 1999; Suzukawa et al. 2000) and, in hippocampal neurons, high levels of O 2 - (Bindokas et al. 1996) modulate neuro- nal plasticity (Hongpaisan et al. 2004). Redox state has also been shown to modulate differentiation of mesence- phalic precursors (Lee et al. 2003) and of neural crest stem cells (Morrison et al. 2000). ROS can therefore influence multiple aspects of neural differentiation and function, including the survival and the plasticity of neurons, the proliferation of neural precursors, as well as their differ- entiation into specific neuronal cell types. The determina- tion of induced variations in redox environment by ROS at high spatial resolution is therefore crucial, and could pro- mote novel therapeutic approaches aimed at protecting against oxidative stress by identifying specific redox-sen- sitive sites that could then be targeted for intervention. Whilst traditional biochemical analysis of tissue and cell extracts suffer from several technical limitations when Proceedings of the XIX Congress of the Italian Society of Pure and Applied Biophysics (SIBPA), Rome, September 2008. G. Maulucci  M. Papi  G. Arcovito  M. De Spirito (&) Istituto di Fisica, Universita ` Cattolica del Sacro Cuore, L.go Francesco Vito 1, 00168 Rome, Italy e-mail: m.despirito@rm.unicatt.it G. Pani  S. Fusco  T. Galeotti Istituto di Patologia Generale, Universita ` Cattolica del Sacro Cuore, L.go Francesco Vito 1, 00168 Rome, Italy M. Fraziano Dipartmento di Biologia, Universita ` di Roma ‘‘Tor Vergata’’, Via della Ricerca Scientifica, 00133 Rome, Italy 123 Eur Biophys J (2010) 39:993–999 DOI 10.1007/s00249-009-0470-9