Study On Potassium Channels And Effect Of Cisplitin On The I_KDR Of Spiral Ganglion Neurons From Neonatal Rats

Part1Objective: To establish a cytological model of spiral ganglion neurons in noenatal rats in vitro.Methods: The primary culture of spiral ganglion neurons(SGNs) was carried out with seven days postnatal wistar rats.The process of cellular growth of SGN were observed by inverted/phase contrast microscopy.Immunocytochemical identification was performed on the cultured SGN of wistar rats by the monoclonal antibody of neurofilament protein (NFC-McAb).Results: The present work showed that the dissociated SGNs of wistar rats could survive well and had a nomal phenotypic differ entiation in vitro .The cell amount and surviving period of SGNs were able to meet the requirements of cytological exeperiments.Conclusion: This methods extends the material sources of inner ear tissue culture and provides a useful approath to the neurobiologic studies on the peripheral auditory system in vitro.Part2Objective: To observe the potassium channels characteristic of spiral ganglion neurons from postnatal rats. Methods: The potassium currents were recorded by whole-cell patch clamp technique,and studied its characteristics by pharmacology and kinetics of activation.Results: After block Na+ channels by TTX,standard whole cell voltage dependent potassium current(IK) and delayed rectifier potassium current(IKDR) were recorded who are out currents. Their activated voltage were–50mV and–40mV,respectively. 4-aminopyridine(4-AP) and tetraethylammonium(TEA) blocked components of outward potassium currents.Their half maximal activation voltage(V1/2) were (25.25±2.15)mV(n=10)and(18.65±1.88)mV(n=10).Conclusion: The spiral ganglion neurons has good shape and physiological characteristic and can be observed potassium channels,and be suitable for study on spiral ganglions neurons by patch clamp.Part3Objective: To investigate the effect of cisplatin on outward delayed rectifier potassium currents(IKDR currents)in isolated spiral ganglion neurous cells and to analyze possible intracellular mechanism of this effects by using the patch-clamp technique.Method: The IKDR was recorded and measured before and after the application of 10μmol/L cisplatin in external solution by using the whole cell patch-clamp technique.Results: Cisplatin could inhibit SGNs voltage-dependent polassium charrels,the zero current potential and reversal potential of IKDR currents were both shift to polarizing directions by 10μmol/L cisplatin.Moreover,the maxinal magnitude of IKDR currents were dicreased by 23.3% when giving the test pulse at +50 mV.Cisplatin's inhibitation on IKDR has dose-depentence with cisplatin'concerntration in extracellular fluid and the currents recoverd completely after cisplatin being washed out.Conclusion: This research explained the otoxic mechanis of cisplatin through its action on keeping from spiral ganglion neuron's IKDR from the eletrophysiological aspect and set a foundation for further research.