In cardiac myocytes of newborn rats, the degree of intercellular communication through gap junctional channels closely depends on the metabolic state of the cells. In contrast, in stably transfected HeLa cells expressing rat cardiac Cx43 (the main channel-forming protein present in ventricular myocytes), a major part of junctional communication persists in ATP-depleted conditions or in the presence of a broad spectrum inhibitor of protein kinases (H7, 1 mM). However, a progressive closure of junctional channels is observed when cells are exposed to antimycin A, an inhibitor of cytochrome reductive electron transport, known to cause ATP to fall rapidly. The degree of cell-to-cell communication has been determined in dual voltage-clamp conditions by means of either the electrical junctional conductance or the junctional permeability for a fluorescent dye in both HeLa cells and ventricular myocytes obtained from neonatal rats killed by cervical dislocation followed by decapitation.

In perforated patch clamp conditions, the introduction of antimycin A (10 µM) in the external bath led to a complete interruption of cell-to-cell communication between HeLa cells in 15-20 min (n = 13). When intracellular ATP concentration was kept constant (5 mM) in whole-cell conditions, antimycin A also elicited, within 35-40 min, a complete interruption of the cell-to-cell communication (n = 11), showing that ATP supply was insufficient to prevent channel closure. A second metabolic inhibitor, KCN, was able to significantly reduce the intercellular dye diffusion between rat cardiac myocytes, and had negligible effects on HeLa cells. Indeed, in the first cells, the relative permeability constant for dye transfer was lowered after 20 min from 0.3725 ± 0.0221 min^-1 (mean ± S.E.M.) to 0.0352 ± 0.0261 min^-1 (P < 0.01 by Student’s paired t test, n = 27), whereas it was only slightly reduced, from 0.1513 ± 0.0213 to 0.1289 ± 0.0353 min^-1 (n = 22), in Cx43-HeLa cells.

The possibility of an indirect action of antimycin A, via an important increase in cytosolic calcium concentration ([Ca²⁺]_i), has been examined by means of the fluorescent calcium indicator indo-1. Antimycin A caused an increase in [Ca²⁺]_i; however, a high-K⁺ solution, which does not alter the junctional coupling, elicited a still higher rise in the [Ca²⁺]_i, showing that the latter was not responsible for the inhibition of gap junctional communication. Altogether, these results suggest that, besides its depleting effects on the intracellular ATP content, antimycin A is able to directly interrupt cell-to-cell communication, even when both ATP and calcium concentrations are kept stable.

This study was supported in part by grants from the European Community RDT action QLG1-CT-1999-00516.