Calretinin Immunoreactive Nerve Endings in the Trachea and Bronchi of the Rat.

Abstract

ABSTRACT. Nerve endings showing calretinin immunoreactivity were examined in the lower respiratory tract of the adult rat. Tree-like nerve endings were immunostained in the tracheal and bronchial smooth muscle layer. The endings that arose from thick nerve fibers and formed corpuscles composed of many arborized nerve terminals. A few of the nerve endings were also observed in the lamina propria of the tracheal mucosa, close to the epithelial layer. Immunoelectron microscopy revealed that the immunoreactive terminals were filled with mitochondria and scattered among the intermuscular collagen fibrils. Schwann cell sheath and collagen fibrils were intercalated between the smooth muscle cells and nerve endings. The calretinin immunoreactive nerve endings observed in the present study seem to be slowly adapting stretch receptors.-KEY WORDS: calretinin, sensory nerve ending, trachea.J. Vet. Med. Sci. 61(3): 267-269, 1999 incubated with polyclonal rabbit antibodies (IgG) against guinea-pig calretinin (diluted 1:5,000; AB149; Chemicon, Temecula, CA, U.S.A.) for 72 hr at 4°C. The specificity of the calretinin antibody has been described previously [11]. After incubation, the specimens were treated with biotinylated antibodies that had been raised in goat against rabbit IgG (Vector, Burlingame, CA, U.S.A.) and reagents from an ABC kit (Elite ABC kit; Vector) for 90 min each at room temperature. Immunoreactions were made visible by incubation with Tris-HCl buffer containing 3,3'-diaminobenzidine tetrahydrochloride and H 2 O 2 . Immunostained specimens were mounted on gelatin-coated slides, air-dried, dehydrated in ethanol, cleared with xylene, and sealed with coverslips. For examination by immunoelectron microscopy, 50-µm-thick cryostat sections of the thoracic trachea and the lung were immunostained with calretinin antibodies without treatment with PBS containing Triton X-100 or H 2 O 2 . Portions of the sections containing immunostained endings were removed under the dissecting microscope, postfixed in 1% osmium tetroxide for 60 min, dehydrated with a graded ethanol series, and embedded in Epon 812. The ultrathin sections, without any contrast staining, were examined by a transmission electron microscope at 75 kV (H-8100; Hitachi, Tokyo, Japan).Tree-like nerve endings showing calretinin immunoreactivity were identified in the smooth muscle layer of the trachea and the larger bronchi (Fig. 1A-C). The endings were distributed mainly in the smooth muscle layer of the thoracic trachea and the principal and lobar bronchi, without the smaller bronchi. Only 1 to 10 endings were found in the whole length of the trachea (Fig. 2). A few endings were also observed in the lamina propria of the tracheal mucosa close to the epithelial layer (Fig. 1D).Thick nerve fibers with or without swellings ramified into several branches to form the tree-like endings (Fig. 1). The axons ended with laminar terminal arborizations. The corpuscles were arranged in parallel to the longitudinal axis of the smooth muscle cells, about 100-200 µm in le...

Neurochemical characterisation of sensory receptors in airway smooth muscle: comparison with pulmonary neuroepithelial bodies

“…CRT IR has been described before in branching nerve endings in smooth muscle of the trachea, principal and lobar bronchi, but not in smaller bronchi in rat lungs (Yamamoto et al 1999). Since SMARs were also found at the level of bronchioli in the present study, immunostaining of the calcium-binding protein may not be a sufficiently reliable marker for SMARs.…”

“…c The combination of the red and green channels clearly shows that the VGLUT1-ir intraepithelial arborisation coincides with the NEB. Note that VGLUT1 (arrows) and CGRP IR (open arrows) are present in different nerve fibre populations b nary stretch receptors' (von Du¨ring et al 1974) and 'SARs' (Yamamoto et al 1999;Yu et al 2004) suggests that most likely the same structures are concerned.…”

“…In rats, for example, calretinin (CRT) IR showed extensively branching nerve endings in the smooth muscle layer of the trachea and in the principal and lobar bronchi (Yamamoto et al 1999). Also, the Na + /K + -ATPase a3-immunostaining in rat and rabbit lungs revealed nerve terminals with multiple branches, presumed to be embedded in the airway smooth muscle or in the lamina propria (Yu et al 2003(Yu et al , 2004.…”

“…Based on their location, appearance and presumed relationship to physiologically characterised sensory receptors, the branching receptor-like terminals that were supposed to be associated with airway smooth muscle have been named 'smooth muscle nerve spindles' (Larsell 1922), 'pulmonary stretch receptors' (von Du¨r-ing et al 1974) and 'slowly adapting stretch receptors' (SARs) (Yamamoto et al 1999;Yu et al 2004). However, because of the limited data that are available on the morphology of putative airway receptors, correlation between physiologically and morphologically defined lung receptors should be made with great care.…”

“…The present study intended to further characterise the receptor terminals that have been described in the wall of rat bronchi (Yamamoto et al 1999;Yu et al 2004). To this end, airway smooth muscle was visualised using antibodies against a smooth muscle actin (aSMA) and multiple immunocytochemical stainings were performed to clarify the neurochemical coding of receptor-like structures in rat airways.…”