Characterization of inositol 1,4,5-trisphosphate (IP3) receptor subtypes at rat colonic epithelium

Abstract

The aim of the present study was the characterization of the subtypes of inositol 1,4,5-trisphosphate receptors (IP3R) in rat colonic epithelium. A monoclonal antibody against IP3R1 did not stain the colonic epithelial cells. In contrast, IP3R2 and IP3R3 were found within the epithelium; however, with a distinct intracellular localization and differences in their distribution along the crypt axis. IP3R2 immunoreactivity was found within the nuclei of the epithelial cells. The signal was distributed all over the nucleus and not restricted to the nuclear envelope as demonstrated by counterstaining with lamin B1 and electron microscopical examination after immunogold labelling. In contrast, an antibody against IP3R3 stained the epithelial cells mostly in their apical half in accordance with the typical localization of IP3R in organelles such as the endoplasmic reticulum. In addition, there was a gradient from the surface region towards the crypt fundus, where the IP3R3 signal could not be detected. Despite the strong IP3R3-gradient, in saponin-permeabilized colonic crypts exogenously administered IP3 or adenophostin A evoked a similar depletion of mag-fura-2-loaded intracellular Ca2+ stores in crypt and surface cells suggesting a contribution of the nuclear IP3R2 to the Ca2+ release. This conclusion was confirmed by experiments with isolated nuclei from colonic epithelium, at which IP3 was able to induce changes in the Ca2+ concentration, which were inhibited by 2-aminoethoxy-diphenylborate (2-APB), a blocker of IP3 receptors. These results demonstrate that the colonic epithelial cells undergo changes in IP3R subtype expression during differentiation.