Labeling phrenic afferents with intrapleural AAV-PHP.S. J Neurosci Methods 2025 Jul;419:110466
Date
05/08/2025Pubmed ID
40334752Pubmed Central ID
PMC12147650DOI
10.1016/j.jneumeth.2025.110466Scopus ID
2-s2.0-105004266981 (requires institutional sign-in at Scopus site) 2 CitationsAbstract
BACKGROUND: Understanding the role of musculoskeletal afferents in health and disease relies on the ability to selectively label afferents. Traditional approaches involve using adeno-associated viral (AAV) tools to transduce afferents with intrathecal, intramuscular, or direct dorsal root ganglion (DRG) injections. However, these approaches are surgically invasive, have non-specific labeling, or do not target functional groups of afferents. For example, labeling phrenic afferents arising from the diaphragm muscle is challenging due to the presence of musculoskeletal and cutaneous afferents from the forelimb, neck, and shoulder in the C3-C5 DRGs.
NEW METHOD: Using a new capsid variant of AAV9 with enhanced tropism toward afferents (AAV-PHP.S), we investigated if intrapleural injection of AAV-PHP.S transduces phrenic afferents in the cervical DRGs and spinal cord.
RESULTS: In animals receiving AAV-PHP.S, we observed robust tdTomato labeling in the DRGs, dorsal roots, dorsal columns, and spinal projections throughout the spinal gray matter. We did not see the same pattern of afferent labeling when we transected the phrenic nerve prior to intrapleural injection, nor did we find any evidence for motor neuron labeling. Classification of labeled afferents suggests preferential labeling of large diameter proprioceptive neurons. Time-course experiments show tdTomato expression in DRG neurons plateaued by 2 weeks.
COMPARISON WITH EXISTING METHODS: To our knowledge this is the first AAV-based method that preferentially targets phrenic afferents without also labeling phrenic motor neurons.
CONCLUSIONS: This approach labels phrenic afferents and may be used in combination with optogenetic or chemogenetic tools to advance our understanding of the functional role of phrenic afferents.
Author List
Morales AC, Holmes TC, Sanchez FT, Huang H, Williams JJ, Streeter KAAuthors
Taylor C. Holmes PhD, BS, DPT Postdoctoral Researcher in the Physiology department at Medical College of WisconsinJordan J. Williams MD, PhD Assistant Professor in the Biomedical Engineering department at Marquette University
MESH terms used to index this publication - Major topics in bold
AnimalsDependovirus
Ganglia, Spinal
Genetic Vectors
Male
Mice
Neurons, Afferent
Phrenic Nerve
Spinal Cord
