Alterations of tight and gap junctions in mouse hepatocytes following administration of colchicine

Abstract

Following the administration of colchicine at a dosage (1 mg/mouse) known to cause an antimicrotubular effect, membranes as well as tight and gap junctions of hepatocytes were studied using the thin-sectioning and freeze-fracturing technique. As early as 1 h after administration of colchicine the intercellular spaces were dilated and vacuoles were visible within the cytoplasm. The bile canaliculi became enlarged, and after lanthanum perfusion the tracer was found in the canalicular lumen, i.e., the tight junctions became permeable to the tracer. These findings correlated with a disorganized arrangement of the tight junctional strands of the zonula occludens. In some regions the strands showed interruptions and frequently ended freely in a diffuse pattern on the plasma membrane. Proliferation of tight junctions could be observed at various locations on the plasma membrane. The gap junctions also exhibited alterations. They showed an irregular outline with outpouchings, in addition to an enlargement in their total area from approximately 0.5 microns 2 in controls up to approximately 2 microns 2 in treated mice. The surface area occupied by these junctions increased from 4% (controls) to 20% (treated) of the hepatic plasma membrane. In the cytoplasm of hepatocytes from colchicine-treated mice gap-junctional vesicles were frequently observed. In view of the antimicrotubular effect of colchicine it is tentatively suggested that the intact microtubular system of the cell may play a decisive role in the regular formation of gap and tight junctions, either directly or indirectly via microfilaments.

Visualization of gap junction mobility in living cells

“…The plaque regions are in continuous motion, moving around at considerable speed, changing shape, fusing frequently with each other, and separating into smaller patches. This is in agreement with the view that neither microfilaments nor microtubules are needed for gap junction assembly (Kidder et al 1987;Feldman et al 1997;George et al 1999), although certain aspects of clustering and stability may be regulated by these filament systems (e.g., Rassat et al 1982;Wang and Rose 1995), and connexins may associate with cortical linker and/or signaling molecules (e.g., Giepmans and Moolenaar 1998;Toyofuku et al 1998). However, in contrast to Jordan et al (1999) who were only able to record fluorescence for up to 37.3 min, we should like to stress that the gap junctions formed in our cells were highly motile.…”

Tight junction formation in cultured epithelial cells (MDCK)

“…Freeze-fracture replicas of the (Table, line 1 I) in agreement with previous information obtained in mature monolayers, indicating that they can be opened and resealed by removal and restoration of Ca 2 § in the presence or absence of this drug [23,24]. Meldolesi et al [22] have shown that colchicine has no effect on the pattern of resting tight junctions of acinar pancreatic cells of guinea pig, but it does affect reassembly when they have been subject to removal and restoration of Ca 2 § and Rassat et al [30], have found that this drug disorganizes the strands between rat hepatocytes. Colchicine, therefore, seems to have a distinct effect on tight junctions of different tissues.…”

Gap junction assembly in the preimplantation mouse conceptus is independent of microtubules, microfilaments, cell flattening, and cytokinesis.

“…We thought it reasonable that cytoskeletal elements might exert such a restraining influence. Both microtubules and microfilaments have been implicated in such a role, since treatment of somatic tissues with either colchicine or cytochalasin has been shown to promote the proliferation and expansion of gap junctions (20,21). However, neither cytochalasin nor nocodazole could confer competence for gapjunction assembly on four-cell embryos, leading us to conclude that these elements do not exert restraint to control the timing of this event in mouse embryos.…”

“…A point that must be emphasized is that successful dye transfer between cleavage-blocked embryos occurred only when both embryos in the aggregate were the same age as compacted eight-cell embryos: four-cell embryos (60 hr after human chorionic gonadotropin) treated with cytochalasin for 4 hr, then aggregated immediately with compacted eight-cell embryos (without waiting for controls of the same batch to cleave and compact), were not capable of assembling interembryonic membrane channels ( 20 gm and applies to all parts of the figure. In c the recipient embryo (on the right) was prevented by cytochalasin treatment from initiating cell flattening, whereas the donor embryo has decompacted due to the continued presence of the drug.…”