Immunohistochemical demonstration of the NK1 tachykinin receptor on muscle and epithelia in guinea pig intestine

Abstract

Background & Aims: Previous immunohistochemical studies failed to reveal neurokinin (NK)1 tachykinin receptors on intestinal muscle, despite convincing pharmacologic data indicating their presence. This study aimed to apply optimal immunohistochemical methods to reveal the receptors. Methods: NK1-receptor immunoreactivity was examined by confocal microscopy in tissue incubated with or without 10-7 mol/L substance P (SP), 10-7 mol/L SP plus 10-6 mol/L NK1 receptor antagonist (CP99994), or with fluorescent cyanine 3.18 (Cy3) SP. Results: Without incubation, NK1-receptor immunoreactivity was strong on muscle of the rectum and distal colon and weak in proximal colon and small intestine. NK1 receptor was located on the surface of muscle cells in all gut regions. Exposure to SP increased the intensity of immunoreactivity, and the receptor moved into the cytoplasm. Mobilization of the receptor by SP was blocked by the NK1-receptor antagonist CP99994. Cy3-SP was internalized by muscle cells and colocalized with the receptor. NK1-receptor immunoreactivity occurred on crypt epithelial cells in the small intestine and the base of glands in the proximal colon. Conclusions: The NK1 receptor occurs on the external muscle throughout the small and large intestines. SP binds and triggers NK1-receptor aggregation and internalization in the muscle.