PATCH-CLAMP RECORDINGS FROM SUBPOPULATIONS OF AUTONOMIC AND AFFERENT NEURONS IDENTIFIED BY AXONAL TRACING TECHNIQUES

This study determined whether axonal tracing methods can be used in combination with patch-clamp techniques to examine the electrical properties of identified populations of autonomic and afferent neurons in the adult rat. Fluorescent dyes (Fast Blue, FB and Fluoro-Gold, FG) were injected into the wall of the urinary bladder or colon and into various somatic structures to label postganglionic neurons in the major pelvic ganglia (MPG) as well as visceral and somatic afferent neurons in the lumbosacral dorsal root ganglia (DRG) and trigeminal ganglia (TG). One to 3 weeks after dye injection, neurons were isolated from ganglia by enzymatic dissociation. Following dissociation, single neurons labelled with FB were identified in the three types of ganglion preparations; however FG was only identified consistently in TG neurons. FB was retained in neurons during short-term culture (1-5 days). Following 10 to 20 s exposure to UV light which was required for identification of the cells, whole-cell patch-clamp recordings revealed that the electrophysiological properties of FB-labelled cells did not differ from those of unlabelled cells. However, a more prolonged exposure (1-5 min) of the neurons to UV light produced irreversible damage to the cells which was evident as changes in the action potential, sodium current and resting membrane potential. These results indicate that patch-clamp recording in combination with axonal tracing is a useful approach for studying the electrical properties of identified populations of autonomic and afferent neurons. Preliminary data indicate that small diameter bladder and colon afferent neurons have similar electrophysiological properties and that these properties are very different from those of large diameter somatic afferent neurons.