Manganese-enhanced MRI of layer-specific activity in the visual cortex from awake and free-moving rats.

Visual processing, an important function of the central nervous system, starts in the retina and continues in several sub-cortical regions and the visual cortex. In the rat brain, visual information is first processed at the level of the lateral geniculate nucleus, which then provides thalamic input to visual cortical layers IV and deep layer III, (Paxinos, 1985). Pathways to other brain regions implicated in vision also originate in the primary visual cortex, including those to the superior colliculus (from layer V) and the accessory visual cortex (from layers II through VI). Brain activity from awake and free-moving animals is assessed using either immunocytochemical staining techniques (Montero and Jian, 1995) or encoding activity based on the accumulation of an injected metabolic marker like 2-14C deoxyglucose (Cooper and Thurlow, 1991; Toga and Collins, 1981). In either case, readout is performed from histological slices and so requires sacrificing the animal. Non-invasive techniques are available, such as blood oxygenation-level dependent (BOLD) MRI or perfusion imaging that allow for multiple measurements from the same animal over time, although such methods have relatively lower spatial resolution and provide only indirect (i.e., hemodynamic) mapping of the location of brain activity. In addition, assessing animal brain activity inside a scanner requires anesthesia, which can modulate the results (Masamoto et al., 2007). There is a need for a high resolution imaging method that more directly measures encoded brain activity from awake and free-moving subjects. Manganese-enhanced MRI (MEMRI) with systemically administered MnCl2 has been used to examine neuronal activity in vivo (Berkowitz et al., 2006; Yu et al., 2005). Also, uptake of manganese following direct injection into neuronal structures been used for in vivo tract-tracing studies (Lindsey et al., 2007; Watanabe et al., 2001). The paramagnetic Mn2+ ion can enter active neurons through voltage-gated calcium channels, among others channels (Lee et al., 2005). Since Mn2+ efflux from cells is slow, its stimulation-dependent accumulation in active brain regions can be measured as a decrease in tissue T1 at high spatial resolution hours later (Aoki et al., 2004; Alvestad et al. 2007; Sun et al., 2006). In this manner, MEMRI has measured sound-evoked activity in the midbrain from awake and free moving rodents, with spatial resolution surpassing that available in BOLD (Yu et al., 2005). However, in that study stimulation-dependent differences in whole cortical manganese accumulation were not found. It is not yet known if more spatially selective analysis of manganese accumulation in the cortex would be more sensitive to activity differences. In this study, we used MEMRI to examine manganese accumulation from awake and free moving rats exposed to visually stimulating environment. We reasoned that preservation of layer-specific information from the cortex might be a more sensitive approach to analysis of activity-dependent changes, since some layers of the primary visual cortex (e.g., layer IV) are particularly responsive to stimulation (Cooper and Thurlow, 1991; Toga and Collins, 1981), and therefore would demonstrate activity-dependent changes in manganese accumulation.