Labeling Phrenic Afferents with Intrapleural AAV-PHP.S

Authors

Amanda C. Morales, Marquette University
Taylor C. Holmes, Marquette University
Felix T. Sanchez, Marquette University
Haozhi Huang, Marquette University
Jordan J. Williams, Marquette University
Kristi A. Streeter, Marquette UniversityFollow

Document Type

Article

Publication Date

7-2025

Publisher

Elsevier

Source Publication

Journal of Neuroscience Methods

Source ISSN

0165-0270

Original Item ID

DOI: 10.1016/j.jneumeth.2025.110466

Abstract

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.

Comments

Accepted version. Journal of Neuroscience Methods, Vol. 419 (July 2025). DOI. © 2025 Elsevier. Used with permission.

Recommended Citation

Morales, Amanda C.; Holmes, Taylor C.; Sanchez, Felix T.; Huang, Haozhi; Williams, Jordan J.; and Streeter, Kristi A., "Labeling Phrenic Afferents with Intrapleural AAV-PHP.S" (2025). Physical Therapy Faculty Research and Publications. 228.
https://epublications.marquette.edu/phys_therapy_fac/228