Morphological evidence of paracellular transport in perfused rat submandibular glands

The morphological change of the paracellular route for fluid secretion is still a long-standing question. The purpose of this study was to visualize alterations in the cytoskeleton structure of tight junctions caused by carbachol (CCh) and isoproterenol (IPR) treatment of perfused rat submandibular glands (SMGs), using freeze-fracture (FF) replicas of rapidly frozen tissues. Isolated SMGs from male Wistar rats were perfused and stimulated with 1 microM CCh and IPR. Specimens were immediately rapidly frozen with liquid helium by metal contact. After cutting and deep etching, FF replicas were obtained by rotary shadowing and were examined by transmission electron microscopy. After CCh/IPR stimulation, the strand particles of TJs rearranged with free ends and terminal loops. In the vertical fracture surface, cytoskeletal filaments beneath the plasma membrane were arranged in a thicker layer than those of the gland without stimulation. Contraction of the submembranous actin cytoskeleton during exocytosis elicited by CCh/IPR may cause rearrangement of TJ strands due to direct interactions between the TJ membrane particles and actin filaments via the tiny bridging structures. The rearrangement and movement of TJ membrane particles involves reconstruction of the subluminal membranous actin filament network through the intermediary of interstitial molecules and may modulate increased paracellular permeability after CCh/IPR stimulation.