The visualisation of cardiovascular innervation in the guinea pig using antiserum to protein gene product 9.5 (pgp 9.5)

Abstract

The various subpopulations of autonomic and sensory nerves supplying the mammalian cardiovascular system may be demonstrated using specific immunocytochemical and histochemical techniques, but no single marker has previously been available for the visualisation of the entire innervation. Protein gene product (PGP) 9.5 was first identified in extracts of human brain and found to represent a major protein component of the neuronal cytoplasm. We have demonstrated that PGP 9.5 immunoreactivity occurs in the guinea pig cardiovascular innervation and is present in more individual nerve fibres than other general neuronal markers (neuron-specific enolase and neurofilaments). PGP 9.5 immunoreactivity was localized to both intrinsic neurones and nerve fibres in the guinea pig heart. In the vascular system PGP 9.5-immunoreactivity occurred in an extensive plexus of fine perivascular nerve fibres and fascicles running around and along both arteries and veins, mainly at the adventitial-medial border. At the ultrastructural level, this immunoreactive material was localized to the axonal cytoplasm and did not appear to be associated with cytoskeletal elements or secretory vesicles. 6-Hydroxydopamine (6-OHDA) pretreatment resulted in the degeneration of noradrenergic axon terminals containing PGP 9.5, tyrosine hydroxylase (TH) and neuropeptide tyrosine (NPY) immunoreactivities. Most of the perivascular nerve fibres which remained displayed substance P- and calcitonin gene-related peptide (CGRP) immunoreactivity, as well as PGP 9.5 immunoreactivity. Capsaicin pretreatment resulted in a depletion of both substance P and CGRP immunoreactivity, but had no apparent effect on PGP 9.5 immunostaining. In the heart PGP 9.5 immunoreactivity also appeared to be present in presumed postganglionic cholinergic nerves. PGP 9.5 may be a useful marker when examining regional variations in cardiovascular innervation and for determining the relative proportions of nerve subpopulations.