Differential effects of intestinal ischemia and reperfusion in rat enteric neurons and glial cells expressing P2X2 receptors

Abstract

Background. Intestinal ischemia followed by reperfusion (I/R) may occur following intestinal obstruction. In rats, I/R in the small intestine leads to structural changes accompanied by neuronal death. Aim. The objective was to analyze the impact of I/R injury on different neuronal populations in the myenteric plexus of the rat ileum after different periods of reperfusion. Methods. The superior mesentery artery was occluded for 45 minutes, and animals were euthanized after 24 hours and 1 week of reperfusion. Immunohistochemical analyses were performed with antibodies against the P2X2 receptor in combination with antibodies against nitric oxide synthase (NOS), choline acetyltransferase (ChAT), calbindin, calretinin, the panneuronal marker anti-HuC/D, or S100β (glial marker). Results. Dual immunolabeling demonstrated that approximately 100% of NOS-, ChAT-, calbindin-, and calretinin-immunoreactive neurons in all groups expressed the P2X2 receptor. Following I/R, the neuronal density decreased in the P2X2 receptor-, ChAT- , calretinin-, and HuC/D-immunoreactive neurons at 24 hours and 1 week following injury compared to the densities in the control and sham groups. The calbindinimmunoreactive neuron density was not reduced in any of the groups. The density of enteric glial cells increased by 40% in the I/R group compared to the density in the sham groups. We also observed increases of 12%, 16%, and 23% in the neuronal cell body profile areas of the NOS-, ChAT-, and calbindin-immunoreactive neurons, respectively, at 1 week following I/R. However, the average size of the calretinin-immunoreactive neurons was reduced by 12% in the I/R group at 24 hours. Conclusions This work demonstrates that I/R is associated with a significant loss of different classes of neurons in the myenteric plexus accompanied by morphological changes and an increased density of enteric glial cells; all of these effects may underlie conditions related to intestinal motility disorder.