Genetic tools that target mechanoreceptors produce reliable labeling of bladder afferents

Abstract

Mechanosensitive bladder sensory neurons are critical for sensing bladder distention, but their role in bladder pain and bladder pathology is poorly understood, due in part, to the challenges of identifying mechanoreceptors in tissue sections. A lot of what is known about how disease alters sensory innervation of the bladder comes from studies that traditionally focus on nociceptive nerve terminals. In seeking tools to characterize the role of non-nociceptive afferents in the bladder, we first examined neurofilament heavy (NFH), a marker for medium to large-diameter myelinated A fibers, in combination with the common marker for peptidergic nociceptors, calcitonin gene-related peptide (CGRP). While there was partial overlap between NFH and CGRP, 87% of NFH fibers were CGRP-negative, underscoring the abundance of non-nociceptive A fibers nerve terminals in the bladder. Two mouse lines that have been used for genetic labeling of mechanoreceptors of the skin were tested for their ability to label bladder afferents. Once crossed to Cre-dependent reporter lines, tyrosine kinase B (TrkB) TrkBCreER2 mice can be used to label A-delta mechanoreceptors while receptor tyrosine kinase ret protooncogene (Ret) RetCreER2 mice can label a combination of A-beta mechanoreceptors and non-peptidergic nociceptors in adult mice (late Ret). Both mouse lines produced successful labeling of bladder nerve terminals demonstrating partial overlap with NFH and minimal overlap with CGRP. Thus, we have identified new genetic strategies to investigate CGRP-negative subpopulations of bladder afferents that remain largely uncharacterized in studies that target peptidergic nociceptors. These tools can help elucidate the role of mechanosensitive afferents in bladder pathophysiology and urologic chronic pain.