Mechanisms of electrolyte transport across the endometrium. II. Regulation by GRP and substance P

The purpose of this study was to investigate the regulation of electrolyte transport across the porcine endometrium by gastrin-releasing peptide (GRP) and substance P (SP). Luminal addition of GRP, neuromedin B (NMB), SP, or neurokinin A (NKA) to mucosal tissues mounted in Ussing chambers produced a multiphasic change in short-circuit current (I(sc)) characterized by an initial rapid increase and subsequent decrease in current. A similar response was obtained after addition of ionomycin or thapsigargin to the tissues. The I(sc) response to the peptides or Ca ionophore was inhibited by pretreatment of the tissues with luminal amiloride or benzamil. GRP and SP were more potent [50% effective concentration (EC₅₀) of 3 nM] than NMB or NKA (EC₅₀ values of 46 and 26 nM, respectively) in producing the decrease in I(SC). Pretreatment with the GRP receptor antagonist 3-Phe-His-Trp-Ala-Val-D-Ala-His-D-Pro-ΨPhe-NH₂ blocked the I(sc) response to GRP and NMB but not to SP or NKA, whereas the NMB receptor antagonist D-Nal-[Cys-Try-D-Trp-Orn-Val-Cys]-Nal-NH₂ was ineffective in inhibiting the I(sc) response to any of the peptides. In contrast, pretreatment of the tissue with the nonpeptide SP receptor antagonist CP-99,994 blocked the I(sc) response to SP and NKA but not to GRP or NMB. Experiments with amphotericin B-permeabilized tissues showed that GRP, SP, ionomycin, and thapsigargin increased current through an outwardly rectifying K conductance located on the apical membrane of the cells. The K- to-Na selectivity ratio of this conductance was calculated to be 2.5:1. These experiments showed that GRP and SP, acting through different receptors; produced an increase in K efflux through a Ca-dependent K conductance present in the apical membrane of surface endometrial epithelial cells. In addition, immunohistochemistry data showed that GRP-like immunoreactivity was localized to surface and glandular epithelial cells, whereas GRP receptor antibody labeling was observed in both epithelial and stromal cells. These results suggest that GRP functions as both an autocrine and paracrine regulatory peptide in the endometrium.