Form and function: combined anatomical and functional imaging of behavioral circuits in the medicinal leech
Maria Ashaber1, Yusuke Tomina1, Eric Bushong2, Pegah Kassraian Fard1, Mark Ellisman2, Tom Deerinck2, Daniel A. Wagenaar1
Behaviors in invertebrate as well as vertebrate animals occur as the result of a tight interplay between multiple neurons, enabled by highly specific neuronal connectivity. However, the relationship between activity (function) and connectivity (form) in most neuronal circuits remains poorly understood. To bridge this gap, combining functional and anatomical imaging is needed. Here we combine voltage-sensitive dye (VSD) imaging with serial blockface electron microscopy (SBEM) to construct a linked database of the activity and connectivity patterns within the segmental ganglion of the medicinal leech. We simultaneously recorded from most neurons in the ganglion during several motor behaviors with a VSD sufficiently sensitive to record subthreshold neuronal activity. We then anatomically imaged the entire ganglion with SBEM. As a proof of concept, we have manually traced a critical motor neuron (dorsal excitatory of longitudinal muscles, DE-3) and identified all of its presynaptic partners, finding a surprising number of them. The number of synapses from a given partner onto the target was found to be strongly correlated to the functional coherence of those cells in behavior. We also verified the anatomical presence of many synaptic connections that had previously been described in the physiological literature. Since DE-3 is a common output of the pattern generators for several behaviors that are included in our VSD recordings, our data set promises new insight into how nervous systems are able to produce diverging behavioral outputs using a single common circuit.