, 2010 and Rothschild et al., 2010) as well as from the mouse olfactory bulb (Wachowiak et al., 2004) and rat cerebellum (Sullivan et al., 2005). Various approaches can be used for extracting the action potential activity underlying such somatic calcium transients (Holekamp et al., 2008, Kerr et al., 2005, Sasaki et al., 2008, Vogelstein et al., 2010, Vogelstein et al., 2009 and Yaksi and Friedrich, 2006). For example, an effective approach is the “peeling algorithm” (Grewe et al., 2010), which is based on subtracting single action-potential-evoked calcium transients from the fluorescent trace until no additional event is present in the residual trace. Again, GECIs can be adapted as well for such studies of neuronal

network function in different animal models (see also Table 1). Meanwhile, they have been used in rodents, Drosophila, this website C. elegans, zebrafish, and even primates ( Díez-García et al., 2005, Heider et al., 2010, Higashijima Nintedanib et al., 2003, Horikawa et al., 2010, Li et al., 2005, Lütcke et al., 2010,

Tian et al., 2009, Wallace et al., 2008 and Wang et al., 2003). A promising application of in vivo two-photon calcium imaging is the investigation of neuronal network plasticity. For example, experimental paradigms of visual deprivation (e.g., stripe rearing to influence orientation selectivity or unilateral eyelid closure to influence ocular dominance plasticity) have been shown to impact significantly the functional properties of mouse visual

cortex neurons (Kreile et al., 2011 and Mrsic-Flogel et al., 2007). Similarly, calcium imaging has been used to study the plasticity of neuronal networks in mouse models of disease, for example after ischemic only damage of the somatosensory cortex (Winship and Murphy, 2008). There is a wide interest to examine brain circuits in relation to defined behaviors in awake animals. To achieve this, there are at present two major strategies involving calcium imaging as the central method for cellular functional analysis. One approach involves the use of head-mounted portable minimicroscopes (see section on imaging devices); the other concentrates on the study of head-fixed animals involving the use of standard two-photon microscopes. Figure 8A illustrates an experiment that was performed in the motor cortex of head-fixed mice that were engaged in an olfactory discrimination test (Komiyama et al., 2010). The animals were trained to lick in response to odor A and to stop licking in response to odor B (Figure 8Aa). The somatic calcium transients that were recorded in motor cortical neurons of the behaving mice had an excellent signal-to-noise ratio (Figures 8Ab–8Ac). Such experiments involving head fixation are possible because the mice have been gradually adapted to the experimental set-up, which includes the training in a tube-like construction which provides protection to the animal (in that particular study training lasted for 5 days on average).