Experimental depression of junctional membrane permeability in mammalian cell culture. A study with tracer molecules in the 300 to 800 dalton range

Abstract

Cell-to-cell junctional permeability in mammalian cell cultures was probed with a series of fluorescent tracers ranging 300 to 800 in molecular weight, during treatment with metabolic inhibitors, Ca-transporting ionophore, and carbon dioxide. Treatment with the combination of cyanide and iodoacetic acid (1–2mm each), but not with either one alone, caused reversible junctional blockade to all tracer molecular species, large and small. (Electrical coupling, however, persisted in a proportion of the junctions tested.) Treatment with the ionophore A23187 (2–10 μm) or with CO₂ (an atmosphere of 100% CO₂ equilibrated with the medium) produced selective junctional blockade: transmission of a 688 and an 817-dalton tracer was generally blocked, while that of a 376-dalton tracer and, in certain conditions, that of a 559-dalton one, persisted. The junctional effect of the ionophore required the presence of Ca in the external medium; and effective junctional blockade by CO₂ required pretreatment in medium with high Ca concentration or, interchangeably, pretreatment in medium with high CO₂ concentration. In one cell type, prolonged exposure to medium with high Ca concentration alone sufficed to block transmission of the 688-dalton tracer. These effects are discussed in terms of the Ca hypothesis of junctional permeability regulation. In comparison with mammalian (or other vertebrate and invertebrate) organized tissues or with insect cell cultures, the mammalian cell cultures are more resistant to junctional blockade. This difference in transmission stability is discussed in terms of intracellular Ca-buffering capacities of the junctional locales; in particular, in terms of the electron-microscopic finding in the mammalian cultures of fine, bilateral cell processes connected by gap junctions.