One of the major obstacles hindering the progress of the study on cochlear neurotransmission is the lack of experimental preparations adequate for studying the process in situ, a technique that allows direct observations of both the pre- and post-synpatic responses in situ by either electrophysiological or imaging methods. For the purpose of studying the membrane biophysics of hair cells (HCs) and spiral ganglion (SG) neurons in situ, we propose to develop a cochlear slice preparation in which tissue slices are cut from the cochlea of early postnatal mice by a vibratome along the modiolus axis. Our preliminary data have shown that viable cochlear slices could be made from early postnatal cochleae (younger that P2), and normal neural circuits connecting the HCs and SG neurons were intact in slices and exposed to external experimental manipulations needed for either electrophysiological or optical recordings. HCs and SG neurons showed normal resting membrane potentials in cultures. SG neurons fired action potentials in responses to applications of glutamate receptor agonists.
The specific aims i n our proposal will further investigate the feasibility of recording cochlear neurotransmission in situ by: (1) Further optimize the protocol for making cochlear slices from embryonic and postnatal mice and the culture methods for maintaining functions of cochlear slices in long-term cultures. (2) Study the feasibility of using the electrophysiological and imaging approaches to investigate the electrophysiological properties of HCs and SG neurons in the cochlear slice and hemicochlear preparations, and optimize the protocols for experiments. The proposal represents the first attempt to culture cochlear slices for long-term and perform electrophysiological recordings from cochlear slices/hemicochleae. Although long-term cultures and recordings of brain slices are mature techniques now (Jaeger, et al., 1989; Augestine, 1994) and 2-dimensional vibration patterns in the organ of Corti have been successfully measured using the hemicochlear preparation (Richter et al., 1998), no preliminary data exist as for the feasibility of current proposal. The successful completion of this project, very much like what the brain slice technique has brought to the studies of neuronal circuits in the brain, potentially could lead to breakthroughs in the studies of cochlear synaptic physiology, tonotopic organizations of the intrinsic electrical properties of the membrane of HCs and SG neurons, and the issues regarding the role of type II SG neurons in cochlear neurotransmission.
| Peng, B G; Li, Q X; Ren, T Y et al. (2004) Group I metabotropic glutamate receptors in spiral ganglion neurons contribute to excitatory neurotransmissions in the cochlea. Neuroscience 123:221-30 |
| Peng, Ben-Gang; Ahmad, Shoab; Chen, Shanping et al. (2004) Acid-sensing ion channel 2 contributes a major component to acid-evoked excitatory responses in spiral ganglion neurons and plays a role in noise susceptibility of mice. J Neurosci 24:10167-75 |
| Lin, Xi; Webster, Paul; Li, Qingxia et al. (2003) Optical recordings of Ca2+ signaling activities from identified inner ear cells in cochlear slices and hemicochleae. Brain Res Brain Res Protoc 11:92-100 |
