We aim to identify the mechanisms whereby the developing vertebrate nervous system controls visually guided behavior and rapid perceptual decision making (behavioral choice) at the level of synapses, cells and circuits. A main goal is to find neural circuits that perform certain computational steps such as stimulus detection, object classification and action selection, and to understand how circuit function emerges from synaptic connectivity. To achieve this, we use multiphoton calcium imaging and single-cell recordings targeted at genetically defined cell types in the zebrafish nervous system in combination with 3D electron microscopy.
Virtual prey capture sequences in a closed-loop visual environment
Larval zebrafish perform rapid prey capture sequences, in which discrete swim bouts are chained into a sequence, predominantly controlled by the visual system. To better understand the relation between vision and action during this important behavior, we set out to obtain a detailed picture of what the fish sees when freely swimming and hunting small ciliates, its natural prey. From high-speed recordings of freely swimming larvae, body kinematics are automatically parameterized and spectrally analyzed. This way we observed that prey capture sequences are composed of versions of a basic motor pattern, which are superimposed with a graded turning component that allows the larva to adjust its heading direction on a fine, continuous scale towards the target (Trivedi and Bollmann, 2013). Furthermore, we developed closed-loop visual stimulation methods, which allow us to elicit multi-step prey capture sequences in larvae embedded in a virtual environment. These methodological advances allow us to test the role of visual feedback during prey capture. For example, we observed that small spatio-temporal shifts in stimulus updates result in significantly longer reaction times. We hypothesize that visual feedback-driven activity interacts with residual activity from previous motor steps, which could lead to accelerated programming of subsequent motor commands. We now combine these methods with two-photon Ca2+ imaging and patch-clamp recordings in order to investigate the spatio-temporal patterns of activity underlying these visually driven motor sequences.
Layer-specific processing of stimulus motion in tectal microcircuits
Neurons sensitive to the direction of a moving stimulus have been observed at all levels of the visual system, but the underlying mechanisms remain controversial, especially in visual centers beyond the retina. A critical question is whether direction selectivity is coded in retinal ganglion cell activity and relayed to downstream targets, or whether direction selectivity emerges de novo in higher visual circuits from basic, topographically organized input maps. We could address this question for two morphologically distinct cell types in the zebrafish optic tectum (Gabriel et al, 2012). Using the Gal4-UAS-expression system, distinct neuronal cell types in the tectum were labeled with the genetically encoded Ca2+ indicator GCaMP-3. Multi-photon Ca2+ imaging showed that cell types with directional preference for opposing directions were morphologically distinct and exhibited dendritic arborizations in different layers of the tectal neuropil. The dendritic profiles colocalized with presynaptic retinal ganglion cell terminals in sublayers with corresponding directional tuning. Furthermore, multi-photon targeted patch clamp recordings show that these cell-types receive directionally tuned excitatory inputs in the preferred direction and weaker, tuned inhibitory inputs. These findings demonstrate a remarkable correspondence between the functional and structural profile of morphologically distinct cell types, which indicates that the central principle of lamina-specific feature extraction applies not only to the retina but also to higher visual centers.
Classification of object size during visually guided prey capture
Zebrafish larvae perform orienting swims towards a prey-like target or away from a threatening object depending on the size of a visual stimulus. Using high-speed kinematic analysis of swimming behavior, multi-photon calcium imaging, in vivo electrophysiology and closed-loop visual stimulation methods, we find that object size is classified in the activity of different neural populations throughout the retinotectal system (Preuss et al., 2014). First, we devised visual stimuli that closely mimic salient visual features of a prey-like object. In response to these ethologically relevant stimuli, we observed that object size is encoded in the activity of different retinal ganglion cell types (RGC) projecting to the optic tectum. Then, using multi-photon targeted patch clamp recordings, we find that a class of horizontal cells called superficial interneurons (SINs) exhibit distinct, size-dependent synaptic inputs, depending on the tectal layer in which their dendrites arborize. SINs fall into different response types, with a distinct class of SINs preferentially tuned to small, prey-like stimuli. These SINs arborize in the most superficial neuropil, where RGCs are predominantly tuned to small moving objects. At the next processing stage, tectal cells in the periventricular zone exhibit differential size selectivities, with approximately equal fractions of cells tuned either to small or large moving objects. We conclude that behaviorally relevant size classification begins in the retina, and that parallel, size-selective channels activate distinct, size-processing subnetworks in the tectum. These distinct subnetworks may represent a bifurcation point for behavioral decision making during visually controlled motor behavior.
The wiring diagram of the zebrafish spinal cord
A wiring diagram of the central nervous system (CNS) would represent a valuable structural basis for understanding information processing in this vertebrate model system. In collaboration with W. Denk, we aim to analyze the CNS of the larval zebrafish at the connectomic level, using serial block-face electron microscopy (SBEM). We have acquired a high-contrast SBEM data set comprising several segments of the zebrafish spinal cord, which yields information on how the reticulo-spinal system connects to spinal motor circuits. This allows us to map the synaptic contacts between descending hindbrain neurons, spinal interneurons and different pools of motoneurons in order to address the question whether recruitment patterns of motor neurons may be implemented in specific connectivity rules between these cell types.