title: Functional and morphological assessment of diaphragm innervation by phrenic motor neurons.
abstract: This protocol specifically focuses on tools for assessing phrenic motor neuron (PhMN) innervation of the diaphragm at both the electrophysiological and morphological levels. Compound muscle action potential (CMAP) recording following phrenic nerve stimulation can be used to quantitatively assess functional diaphragm innervation by PhMNs of the cervical spinal cord in vivo in anesthetized rats and mice. Because CMAPs represent simultaneous recording of all myofibers of the whole hemi-diaphragm, it is useful to also examine the phenotypes of individual motor axons and myofibers at the diaphragm NMJ in order to track disease- and therapy-relevant morphological changes such as partial and complete denervation, regenerative sprouting and reinnervation. This can be accomplished via whole-mount immunohistochemistry (IHC) of the diaphragm, followed by detailed morphological assessment of individual NMJs throughout the muscle. Combining CMAPs and NMJ analysis provides a powerful approach for quantitatively studying diaphragmatic innervation in rodent models of CNS and PNS disease.
authors.affiliation: Department of Neuroscience, Farber Institute for Neurosciences, Sidney Kimmel Medical College at Thomas Jefferson University; Department of Biology, Arcadia University.
authors.forename: Melanie
authors.initials: M
authors.lastname: Martin
authors.affiliation: Department of Neuroscience, Farber Institute for Neurosciences, Sidney Kimmel Medical College at Thomas Jefferson University.
authors.forename: Ke
authors.initials: K
authors.lastname: Li
authors.affiliation: Department of Biology, Arcadia University; wrightm@arcadia.edu.
authors.forename: Megan C
authors.initials: MC
authors.lastname: Wright
authors.affiliation: Department of Neuroscience, Farber Institute for Neurosciences, Sidney Kimmel Medical College at Thomas Jefferson University; Angelo.Lepore@jefferson.edu.
authors.forename: Angelo C
authors.initials: AC
authors.lastname: Lepore
mesh_terms: D000818:Animals; D001369:Axons / Q000502:physiology; D003964:Diaphragm / Q000294:innervation*; D004195:Disease Models, Animal; D051379:Mice; D009046:Motor Neurons / Q000166:cytology / Q000502:physiology*; D009469:Neuromuscular Junction / Q000166:cytology / Q000502:physiology; D010791:Phrenic Nerve / Q000166:cytology / Q000502:physiology*; D051381:Rats; D013119:Spinal Cord Injuries / Q000473:pathology / Q000503:physiopathology*