Corticotropin-Releasing Factor Receptors in Mouse Spleen: Identification of Receptor-Bearing Cells as Resident Macrophages

Abstract

CRF is a primary integrator of the organism's coordinated neuroendocrine, autonomic, behavioral, and immune responses to stress. In the present study the identity of the cell type(s) expressing CRF receptors in mouse spleen was determined using a combination of cell fractionation and receptor-binding techniques. Autoradiographic studies of the distribution of [125I]Tyro-ovine CRF [( 125I]oCRF)-binding sites in spleen localized CRF receptors primarily to the red pulp and marginal zones. The distribution pattern of [125I]oCRF-binding sites closely resembled the pattern of India ink accumulated in phagocytic cells in the same sections. To identify the specific cell type(s) expressing CRF receptors, [125I]oCRF-binding activity was evaluated in splenic cell populations fractionated on the basis of their physical and functional properties. Macrophages were identified in each fraction by their phagocytosis of polystyrene beads and membrane labeling with MONTS-4, a monoclonal antibody specific for resident macrophages. Spleen cells were fractionated by adherence to glass bead or Sephadex G-10 columns, phagocytosis of carbonyl iron particles, and centrifugation on discontinuous Percoll gradients. By all fractionation methods, there was a significant correlation of [125I]oCRF binding with both phagocytic activity (r = 0.75; P less than 0.001) and MONTS-4 staining (r = 0.84; P less than 0.001), strongly suggesting that CRF receptors are primarily expressed on resident splenic macrophages. However, there was essentially no specific binding of [125I]oCRF to either resident or elicited peritoneal macrophages or to several monocyte/macrophage, B-cell, or T-cell lines. While these results suggest that the expression of CRF receptors may be restricted to a population of splenic macrophages, they do not exclude the possibility that CRF receptors may be induced on resident macrophages in spleen and other immune system-related tissues by factors present in the microenvironment.