| As a neurotransmitter or neuromodulator in the cochlea, ATP has it's complicated effects. In recent years, studies about the effects of ATP in the cochlea have been abundantly accumulated. It can be synthesized and released from the cells in the inner ear and thus ATP is up to certain concentration in the endolymph and perilymph. There are purinergic receptors in the cells of the cochlea and combination of ATP and purinergic receptors can cause Ca2+ concentration rise in hair cells and supporting cells. On the other hand, ATP can affect cochlear potentials such as CM, CAP and SP when perfused into perilymph compartment. At the same time, NO/cGMP pathway, as a signal-transmitting pathway, is extensively involved in the regulation of many physiologic processes. Studies have revealed that NO/cGMP pathway lies in the inner ear and takes part in the regulation of cochlear microcirculation and the physiology of supporting cells as well as function of the cochlear. Scientists have done a lot of studies on ATP and NO/cGMP pathway separately in the past. But there isn't report about the relationship between these two important factors in the inner ear. Thus the comprehension on this physiologic aspect is still not clear enough. We designed this project to elucidate this relationship so that people can wholely understand inner ear physiology.Our project includes the following two parts.Part 1. The changing of cGMP content in the guinea pig cochlea induced by whole-cochlea perfusion with DEA-NO in vitroIn order to study the relationship between ATP and NO/cGMP pathway, we chose the content of cGMP in the cochlea as indicator. Guinea pigs weredecapitated and otocysts were rapidly dissected, immersed into artificial perilymph. Holes were drilled at the apex of the cochlear, the round window and the vestibular window. Different reagents dissolved in perilymph were perfused into cochlea from the hole at apex and the outflow was released from the holes in the round and oval window. The animals were divided into three groups: control group perfused with APL basal solution; the first test group perfused with DEA-NO dissolved in APL basal solution and the second test group with ODQ, an inhibitor of sGC dissolved in APL basal solution. The modiolus with basal membrane and spiral ligament were collected and kept at -70 癈 till measuring cGMP concentration in the cochlear tissue with 125I-cGMP RIA Kit. The result showed that the level of cGMP rose markedly when perfused with ImM DEA-NO while cGMP level were markedly low in the group perfused with ODQ, the second test group but it was still considerably higher then that of the control group. We opine that the method of changing the concentration by perfusing reagents into newly dissected cochlea is feasible and reliable.Part 2. The relationship between in vitro ATP cochlea perfusing and the change of cGMP concentration as well as it's mechanism in the inner ear tissue of guinea pighi order to study the relationship between ATP and NO/cGMP pathway in the inner ear, the animals were divided into five groups perfused with APL basal solution, 330 uM ATP, A-23187, L-NNA+ATP and ODQ+ATP separately. All the reagents were dissolved in APL basal solution. The result showed that cGMP concentration could be markedly raised by perfusing A-23187, a Ca2+ ionophore, and ATP, while perfusing L-NNA and ODQ previously could inhibit the effect of ATP on the production of cGMP in the inner ear. The results indicate that NOSand sGC were involved in the process of cGMP synthesizing caused by ATP. By increasing the Ca2+ concentration in inner ear cells, ATP activated NOS and NO produced by NOS activated sGC in turn. Therefore perfusing ATP into the newly dissected cochlea could activate NO/cGMP pathway. Both ATP and NO/cGMP pathway have complicated effects in the inner ear. They are closely related rather than working independently. ATP can cause elevation of the cytoplasmic Ca2+ concentration by binding to purinergic receptors on cell membrane and NO/cGMP pathway can a...
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