Fig.1.

Schematic representation of the two-photon microscopy setup. The excitation beam is produced by a femtosecond pulsed infrared tunable (720–1020 nm) laser. The laser power is modulated by an Acousto-Optic Modulator (AOM). The beam is scanned in the xy direction by galvanometric mirrors present in the scan head of a Zeiss LMS 7 MP two-photon microscope. The beam then passes through a LP690 dichroic mirror and is focused in the brain of the anaesthetized animal by a 20X-1.0 NA water immersion objective. The emitted epifluorescence is collected and reflected by the LP690 mirror in a non-descanned mode. The fluorescence is finally splitted and filtered using a set of dichroic mirrors and filters and collected by a set of 5 non-descanned detectors mounted in cascade (NDD). The characteristics of the dichroic mirrors and filters are depicted on the scheme.