Here we demonstrate that rats can be trained to voluntarily produce head restraint in which brain motion is limited to a few microns, enabling two recently developed powerful technologies to be used together: (1) high-throughput behavioral training using computer-controlled behavior boxes, and (2) cellular resolution imaging of neural dynamics using two-photon excitation fluorescence microscopy and genetically encoded calcium sensors. The crucial technical development was a precise method
by which the brain can be returned to nearly the exact same location in space on each insertion. As we demonstrate, the spatial precision that can be obtained when the mount is engaged is a few microns, which is less than one neuron’s cell body diameter. Thus, the same field of neurons can be imaged on successive insertions and across successive days. Previous reports have described methods for acclimating rats to forced head Selleckchem PI3K Inhibitor Library restraint
by providing water reward and by gradually GDC-0199 in vitro increasing the duration of restraint (reviewed in Schwarz et al., 2010). Head-restrained rats could be trained to perform motor movements to indicate behavioral choice in sensory discrimination (Harvey et al., 2001, Stüttgen et al., 2006 and Verhagen et al., 2007) and detection tasks (Houweling and Brecht, 2008). However, the training procedures require a long acclimation period and significant experimenter involvement, precluding automation (Schwarz et al., 2010). Moreover, none of these systems allowed animals to transition between head fixation and free motion in a single session, prohibiting behavioral response modalities such as Tolmetin head movements, which are commonly used in
operant conditioning paradigms. The use of spherical treadmills has been shown to help acclimate mice to head restraint. This approach allows mice to report behavioral choice by movement of the treadmill (Dombeck et al., 2010) and can be combined with visual feedback to produce a virtual reality environment where mice can be trained to “navigate” while head-fixed (Harvey et al., 2009). One key advantage of this system is that the amount of force the animal is able to apply to the headplate can be reduced since the treadmill rotates whenever the animal tries to push with its legs. Such an approach could in principle be applied to rats, in which the neural circuitry underlying navigation has been well studied. Indeed, body-tethered rats have already been trained to operate a spherical treadmill in a virtual reality system (Hölscher et al., 2005). However, head-fixed navigation systems for rats have not yet been reported. Miniature head-mounted two-photon microscopes provide an alternative to head immobilization during in vivo imaging (Helmchen et al., 2001 and Piyawattanametha et al., 2009).