An in vitro spinal cord slice preparation for recording from lumbar motoneurons of the adult mouse.
The development of central nervous system slice preparations for electrophysiological studies has led to an explosion of knowledge of neuronal properties in health and disease. Studies of spinal motoneurons in these preparations, however, have been largely limited to the early postnatal period, as adult motoneurons are vulnerable to the insults sustained by the preparation. We therefore sought to develop an adult spinal cord slice preparation that permits recording from lumbar motoneurons. To accomplish this, we empirically optimized the composition of solutions used during preparation in order to limit energy failure, reduce harmful ionic fluxes, mitigate oxidative stress, and prevent excitotoxic cell death. In addition to other additives, this involved the use of ethyl pyruvate, which serves as an effective nutrient and antioxidant. We also optimized and incorporated a host of previously published modifications used for other in vitro preparations, such as the use of polyethylene glycol. We provide an in-depth description of the preparation protocol and discuss the rationale underlying each modification. By using this protocol, we obtained stable whole cell patch-clamp recordings from identified fluorescent protein-labeled motoneurons in adult slices; here, we describe the firing properties of these adult motoneurons. We propose that this preparation will allow further studies of how motoneurons integrate activity to produce adult motor behaviors and how pathological processes such as amyotrophic lateral sclerosis affect these neurons.
Department of Anatomy and Neurobiology, Dalhousie University, Halifax, Nova Scotia, Canada.
Pratip
P
Mitra
Robert M
RM
Brownstone
D000200:Action Potentials / Q000187:drug effects / Q000502:physiology; D000367:Age Factors; D000818:Animals; D001703:Biophysics; D002795:Choline O-Acetyltransferase / Q000235:genetics / Q000378:metabolism; D004558:Electric Stimulation; D005260:Female; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D066298:In Vitro Techniques; D008161:Lumbosacral Region; D008297:Male; D051379:Mice; D008822:Mice, Transgenic; D009046:Motor Neurons / Q000502:physiology*; D018408:Patch-Clamp Techniques; D013116:Spinal Cord / Q000166:cytology*
D049452:Green Fluorescent Proteins; D002795:Choline O-Acetyltransferase