Imaging Voltage Globally and in Isofrequency Lamina in Slices of Mouse Ventral Cochlear Nucleus.

The cochlear nuclei (CN) receive sensory information from the ear and perform fundamental computations before relaying this information to higher processing centers. These computations are performed by distinct types of neurons interconnected in circuits dedicated to the specialized roles of the auditory system. In the present study we explored the use of voltage imaging to investigate CN circuitry. We tested two approaches based on fundamentally different voltage sensing technologies. Using a voltage-sensitive dye we recorded glutamate receptor-independent signals arising predominantly from axons. The mean conduction velocity of these fibers of 0.27 m/sec was rapid but in range with other unmyelinated axons. We then used a genetically-encoded hybrid voltage sensor (hVOS) to image voltage from a specific population of neurons. Probe expression was controlled using Cre recombinase linked to c-fos activation. This activity-induced gene enabled targeting of neurons that are activated when a mouse hears a pure 15 kHz tone. In CN slices from these animals auditory nerve fiber stimulation elicited a glutamate receptor-dependent depolarization in hVOS probe-labeled neurons. These cells resided within a band corresponding to an isofrequency lamina, and responded with a high degree of synchrony. In contrast to the axonal origin of voltage-sensitive dye signals, hVOS signals represent predominantly post-synaptic responses. The introduction of voltage imaging to the CN creates the opportunity to investigate auditory processing circuitry in populations of neurons targeted on the basis of their genetic identity and their roles in sensory processing. Significance Statement The cochlear nucleus uses dedicated circuitry to process and interpret information from the ear. This circuitry is organized tonotopically into laminae, each containing cells with an optimal sensitivity to a specific sound frequency. By targeting a genetically-encoded hybrid voltage sensor (hVOS) to identify neurons activated during the presentation of sound, the properties and function of these neurons become accessible to study in slices of mouse ventral cochlear nucleus. Imaging hVOS signals in these slices recapitulated the tonotopic organization. Imaging with a voltage sensitive dye provided a contrasting global view of signals arising predominantly from unmyelinated axons creating a potential method for studying type II auditory nerve or DCN parallel fibers.