<b>Whole cell patch-clamp recordings of skeletal muscular </b><b>DRG neurons in anesthetized whole </b><b>animal preparations </b>

Abstract

Dorsal root ganglion (DRG) neurons that innervate the skeletal muscle (SK-DRG neurons) are differentiated into various subpopulations and each exhibits different receptive properties expressing different ion channels in different combinations. In this study, we introduce a novel method for in vivo patch-clamp recording of the SK-DRG neurons in rats to allow integrated analysis of their diverse properties. To identify the SK-DRG neurons, the fluorescent retrograde tracer DiI was injected into the left anterior tibial muscle of 6-7-week-old rats anesthetized with pentobarbital sodium. Three to 6 weeks after the injection, the rats were deeply anesthetized again, and each animal was mounted on the stage of an upright microscope. After isolating the left L4-DRG along with the sciatic nerve, maintaining the peripheral connection, it was placed in a chamber on the stage. The individual neurons on the ganglionic surface were then visualized.Whole-cell patch-clamp recording was established from the DiI-positive DRG neurons. Small- and medium-sized DRG neurons, which are mainly associated with nociception, could be classified into subpopulations based on axonal conduction velocity, receptive properties (mechanical and thermal stimuli), and current expression profiles (hyperpolarization-activated current and instantaneous current). Some of them responded to experimental ischemic muscle contraction. Therefore, we discuss some components that could cause muscle pain during ischemic contraction. Our novel method is useful for analyzing the diverse properties of SK-DRG neurons and for ascertaining the factors that cause pain in pathological conditions such as ischemia.