Neural activity manifests itself as complex spatiotemporal activation patterns in cell populations. Even for local neural circuits, a comprehensive description of network activity has been impossible so far. Here we demonstrate that two-photon calcium imaging of bulk-labeled tissue permits dissection of local input and output activities in rat neocortex in vivo. Besides astroglial and neuronal calcium transients, we found spontaneous calcium signals in the neuropil that were tightly correlated to the electrocorticogram. This optical encephalogram (OEG) is shown to represent bulk calcium signals in axonal structures, thus providing a measure of local input activity. Simultaneously, output activity in local neuronal populations could be derived from action potential-evoked calcium transients with single-spike resolution. By using these OEG and spike activity measures, we characterized spontaneous activity during cortical Up states. We found that (i) spiking activity is sparse (<0.1 Hz); (ii) on average, only Ϸ10% of neurons are active during each Up state; (iii) this active subpopulation constantly changes with time; and (iv) spiking activity across the population is evenly distributed throughout the Up-state duration. Furthermore, the number of active neurons directly depended on the amplitude of the OEG, thus optically revealing an input-output function for the local network. We conclude that spontaneous activity in the neocortex is sparse and heterogeneously distributed in space and time across the neuronal population. The dissection of the various signal components in bulk-loaded tissue as demonstrated here will enable further studies of signal flow through cortical networks.bulk loading ͉ population imaging ͉ presynaptic ͉ sparse coding U nderstanding how information is represented and processed in the mammalian neocortex requires measurement not only of single-cell dynamics but also of spatiotemporal activity patterns in identified networks of neurons in vivo. So far, optical imaging of intrinsic or voltage-sensitive dye signals has revealed spatiotemporal dynamics on the scale of cortical columns but has lacked cellular resolution (1). Extracellular recording methods have enabled simultaneous measurements from multiple cells but suffer from poorly defined cell identities, lack of spatial resolution, and are incapable of resolving nonactive neurons (2). These techniques thus fall short on providing a comprehensive description of cortical microcircuits. In particular, these methods cannot monitor the activation of afferent axons that represent the input into a particular local region. Of key importance to a further understanding of information processing in the neocortex will be a method that is capable of simultaneously resolving both input and output of cortical microcircuits with single-cell and single-spike resolution.Here we apply recently developed techniques for two-photon calcium imaging of neocortical cell populations in vivo (3-5) to characterize neocortical activity during Up-and Down-state fluctua...

It is unclear how the complex spatiotemporal organization of ongoing cortical neuronal activity recorded in anesthetized animals relates to the awake animal. We therefore used two-photon population calcium imaging in awake and subsequently anesthetized rats to follow action potential firing in populations of neurons across brain states, and examined how single neurons contributed to population activity. Firing rates and spike bursting in awake rats were higher, and pair-wise correlations were lower, compared with anesthetized rats. Anesthesia modulated population-wide synchronization and the relationship between firing rate and correlation. Overall, brain activity during wakefulness cannot be inferred using anesthesia.

Fusing left and right eye images into a single view is dependent on precise ocular alignment, which relies on coordinated eye movements. During movements of the head this alignment is maintained by numerous reflexes. Although rodents share with other mammals the key components of eye movement control, the coordination of eye movements in freely moving rodents is unknown. Here we show that movements of the two eyes in freely moving rats differ fundamentally from the precisely controlled eye movements used by other mammals to maintain continuous binocular fusion. The observed eye movements serve to keep the visual fields of the two eyes continuously overlapping above the animal during free movement, but not continuously aligned. Overhead visual stimuli presented to rats freely exploring an open arena evoke an immediate shelter-seeking behaviour, but are ineffective when presented beside the arena. We suggest that continuously overlapping visual fields overhead would be of evolutionary benefit for predator detection by minimizing blind spots.