可视膜片钳法记录初级传入纤维介导脊髓背角神经元突触后电流.pdfVIP

550 Acta Physiologica Sinica, August 25, 2004, 56 (4): 550-557 Experimental Technique Visually guided patch-clamp recording of spinal dorsal horn neuron’s postsynaptic current evoked by primary afferent fiber WAN Ye-Hong, WANG Yu-Ying, DAI Fei, HU San-Jue* Institute of Neuroscience, The Fourth Military Medical University, Xi’an, Shaanxi 710033, China Abstract: The authors describe here the procedures for using the gelatin half-embedding method to obtain thin spinal cord slices with attached dorsal roots and performing visually guided whole-cell patch-clamp recording of postsynaptic currents evoked by primary afferent fibers in rat spinal dorsal horn. A segment of spinal cord with attached dorsal roots was prepared and half-embedded in an agar block with 20% (w/v) gelatin. Thin spinal cord slices with attached dorsal roots were obtained with a vibratome and whole-cell patch-clamp configuration was established under the infrared observation. At the holding potential of 70 mV, spontaneous excitatory postsynaptic currents (EPSCs) and dorsal root stimulation-evoked EPSCs were recorded as inward currents. According to the conduction velocity of afferent fibers and stimulus threshold, evoked EPSCs that are mediated by A-like or C-like fibers were distinguished. At the holding potential of 0 mV, spontaneous inhibitory postsynaptic currents (IPSCs) and dorsal root stimulation-evoked IPSCs were recorded as outward currents. Using 5 µmol/L strychnine or 20 µmol/L bicuculline, GABAergic or glycinergic evoked IPSCs could be isolated. Using visual patch-clamp method synaptic transmission can be accurately assessed by measuring postsynaptic currents of the dorsal horn neurons. More importantly, with the aid of infrared observation,