Calcitonin gene-related peptide in visceral afferent nerve fibres: Quantification by radioimmunoassay and determination of axonal transport rates

Abstract

An antibody specific for the C-terrainus of rat alpha calcitonin gene-related peptide has been used in radioimmunoassay to measure concentrations of immunoreactive peptide in the upper gastrointestinal tract of capsaicin-treated and coeliac ganglionectomized rats and to measure axonal transport velocities in the vagus and splanchnic nerves. In adult rats that had been treated soon after birth with capsaicin, immunoreactive calcitonin gene-related peptide in the stomach and duodenum was undetectable (<0.1 pmol/g) compared with 4–10 pmol/g in control rats. Removal of the coeliac ganglion also reduced concentrations of immunoreactive calcitonin gene-related peptide by 5-fold, but Leu-enkephalin and Met-enkephalin Arg 6Gly 7Leu 8-immunoreactivities (which are thought to occur in intrinsic gut neurons) were unchanged by coeliac ganglionectomy. Concentrations of calcitonin gene-related peptide immuno-reactivity in coeliac ganglia were depressed by 90% in capsaicin-treated rats but concentrations of opioid peptide immunoreactivity were similar to control. The results suggest calcitonin gene-related peptide-immunoreactivity in the upper gastrointestinal tract in the rat is predominantly of extrinsic afferent origin. Chromatographie separation on Sephadex G50, or high-performance liquid chromatography revealed that the major immunoreactive form in stomach extracts corresponded to intact calcitonin gene-related peptide, although there was evidence of smaller, less hydrophobic C-terminal fragments. Direct evidence of transport of calcitonin gene-related peptide towards the gut was obtained by ligation of the cervical vagus and greater splanchnic nerves. There was accumulation on the central side of ligatures, which suggested axonal transport velocities in the vagus of about 1.5 mm/h and 0.7 mm/h in splanchnic nerves. The present results are compatible with immunohistochemical findings that suggest the spinal afferent innervation is the major source of calcitonin gene-related peptide in the rat stomach.