The permeability of the cell-to-cell membrane channels in salivary gland cell junction (Chironomus thummi) was probed with fluorescent-labeled amino acids and synthetic or natural peptides. Molecules up to 1200 daltons pass through the channels with velocities depending on molecular size. Molecules of 1900 daltons or greater do not pass. This passage failure seems to reflect the normal size limit for junctional channel permeation; the channels continue to be permeated by the molecules up to 1200 daltons when these are mixed with the nonpermeant molecules. From this size limit a channel diameter of 10 to 14 angstroms is estimated.

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Incorporation of the gene for connexin43, a cell-cell channel protein of gap junction, into the genome of communication-deficient transformed mouse 10T1/2 cells restored junctional communication and inhibited growth. Growth was slowed, saturation density reduced and focus formation suppressed, and these effects were contingent on overexpression of the exogenous gene and the consequent enhancement of communication. In coculture with normal cells the growth of the connexin overexpressors was completely arrested, as these cells established strong communication with the normal ones. Thus, in culture by themselves or in coculture, the connexin overexpressor cells grew like normal cells. These results demonstrate that the cell-cell channel is instrumental in growth control; they are the expected behavior if the channel transmits cytoplasmic growth-regulatory signals.

A technique is devised to determine the spatial distribution of the free ionized cytoplasmic calcium concentration ([Ca2+]i) inside a cell: Chironomus salivary gland cells are loaded with aequorin, and hte Ca2+-dependent light emission of the aequorin is scanned with an image-intensifier/television system. With this technique, the [Ca2+]i is determined simultaneously with junctional electrical coupling when Ca2+ is microinjected into the cells, or when the cells are exposed to metabolic inhibitors, Ca-transporting ionophores, or Ca-free medium. Ca microinjections elevating the [Ca2+]i in the junctional locale produce depression of junctional membrane conductance. When the [Ca2+]i elevation is confined to the vicinity of one cell junction, the conductance of that junction alone is depressed; other junctions of the same cell are not affected. The depression sets in as the [Ca2+]i rises in the junctional locale, and reverses after the [Ca2+]i falls to baseline. When the [Ca2+]i elevation is diffuse throughout the cell, the conductances of all junctions of the cell are depressed. The Ca injections produce no detectable [Ca2+]i elevations in cells adjacent to the injected one; the Ca-induced change in junctional membrane permeability seems fast enough to block appreciable transjunctional flow of Ca2+. Control injections of Cl- or K+ do not affect junctional conductance. The Ca injections that elevate [Ca2+]i sufficiently to depress junctional conductance also produce under the usual conditions an increase in nonjunctional membrane conductance and, hence, depolarization. But injections that elevate [Ca2+]i at the junction while largely avoiding nonjunctional membrane cause depression of junctional conductance with little or no depolarization. Moreover, elevations of [Ca2+]i in cells clamped near resting potential produce the depression, too. On the other hand, complete depolarization in K medium does not produce the depression, unless accompanied by [Ca2+]i elevation. Thus, the depolarization is neither necessary nor sufficient for depression of junctional conductance. Treatment with cyanide, dinitrophenol and ionophores X537A or A23187 produces diffuse elevation of [Ca2+]i associated with depression of junctional conductance. Prolonged exposure to Ca-free medium leads to fluctuation in [Ca2+]i where rise and fall of [Ca2+]i correlate respectively with fall and rise in junctional conductance.