Innervation of bovine pulmonary artery endothelium by nerve fibres arising from a supernumerary artery-associated neurovascular bundle A. Tracey, A. MacDonald, R.A. Davidson, A.D. Corbett, I.C. Wilkie, A.M. Shaw. Department of Biological & Biomedical Sciences, Glasgow Caledonian University, Glasgow. G40BA.	Print Abstract Search PubMed for: Tracey A MacDonald A Davidson RA Corbett AD Wilkie IC Shaw MA

Pulmonary artery branching follows the airway branching pattern (conventional arteries, CA). In addition, small arteries (supernumerary arteries, SA) arise along the length of the conventional arteries and branch, without an accompanying airway, into the adjacent respiratory tissue. Blood flow into the SA may be regulated by a cushion-like structure located at its origin with the CA (Shaw et al., 1999). This histological and ultrastructural study investigated the innervation of CA and their associated SA.

Bovine lungs (either sex) were obtained fresh from the abattoir. Segments of CA (3-4 mm in diameter) and SA (diameter 0.5-1 mm) were dissected, fixed and processed using routine methods. Immunohistochemical studies employed the avidin-biotin immunoperoxidase technique and primary antibodies raised against the neuronal marker protein gene product 9.5 (PGP 9.5, Dako, 1:800 dilution). Sections (5 µm thick) were exposed to the primary antibody for 1 hour at 37^oC, rinsed and exposed to the secondary antibody, which was a 2% biotinylated goat anti-rabbit antibody (Vector Laboratories). In some studies the secondary antibody was a goat anti-rabbit antibody conjugated to either a 1 nm gold particle (British Biocell) or coupled to Alexa Fluor^® 488 (Molecular Probes).

Dissection of the collagen sheath surrounding each SA exposed additional vascular elements that followed the SA into the respiratory tissue in one direction and in the other branched over the adventitial surface of the CA in the region from which the SA originates. Small branches were seen to penetrate more deeply into the wall of the CA. Histological studies identified these additional vascular elements as a neurovascular bundle consisting of a nerve, artery, vein and several smaller vascular components.

In CA, PGP 9.5-like immunoreactivity was detected in association with nerve-like structures in the adventitia and in the media, extending as far as the inner muscularis. Intense immunoreactivity was also observed in association with the intima. This immunoreactivity was localised to the basal side of the endothelium using the immunogold technique, and immunofluorescence microscopy highlighted fibrous structures in the subendothelium as well as bulbous structures associated with the basal edge of the endothelial cells. A similar pattern of immunoreactivity was observed using primary antibodies directed against neurofilaments and neurone-specific enolase. In ultrastructural studies structures resembling nerve terminals were frequently observed in close apposition to, and partially surrounded by, the basal plasma membrane of endothelial cells. These structures were always anuclear and had an electron lucent cytoplasm containing mitochondria and an abundance of circular agranular vesicles (mean max diameter per field of view±SD 66±11 nm, n=16, range 50-90 nm). Microfilaments (7-10 nm in diameter) and microtubules (20-30 nm in diameter) were also present. These observations suggest that bovine pulmonary artery endothelium is innervated.

Shaw, A.M., Bunton, D.C., Fisher, A. et al. (1999). J. Appl. Physiol., 87, 2348-2356.