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Nervous or hormonal stimulation of salivary secretion in vivo is associated with a pronounced efflux of K+ from the secretory, acinar cells into the blood. This K+ efflux is followed in the post-stimulus period by a reuptake of K+ into the glandular tissue. In the present study we monitor the changes in [K+] of physiological solutions perfusing a flow chamber containing isolated segments of mouse submandibular glands. Nervous stimulation or the application of exogenous acetylcholine (ACh, 10(-5) M) to the isolated glandular tissue results in characteristic changes in the [K+] of the superfusate, indicating net K+ release followed by K+ reuptake. The post-stimulus reuptake of K+ is shown to be susceptible to blockade by either ouabain (10(-3) M) or piretanide (10(-4) M). The reuptake was markedly attenuated if Cl- in the superfusate was replaced by either NO3- or SO4(2-). The K+ uptake was, however, unaffected when Br- replaced Cl- in the superfusate. Similar effects were observed in the unstimulated glandular tissues. The introduction of Cl-(-)free media containing either NO3- or SO4(2-) resulted in a loss of K+ from the tissue which was followed, upon reintroduction of Cl-, by a pronounced uptake of K+. When Br- was substituted for Cl- there was very little change in [K+] upon removal or reintroduction of Cl-. The uptake of K+ induced by reintroduction of Cl- after a period of NO3- or SO4(2-) superfusion was blocked by both ouabain and piretanide. This uptake of K+ was also dependent on the presence of extracellular Na+. Both Cl- and Na+ had to be present in the superfusing medium for K+ uptake to be fully manifest. These findings indicate that the K+ uptake observed in both the resting and stimulated submandibular gland cannot be explained as solely due to the activity of the Na+-K+-adenosine triphosphatase (Na+-K+-ATPase). The demonstrated anionic selectivity, dependence on extracellular Na+ and susceptibility to blockade by the diuretic piretanide would strongly suggest that a coupled Na+-K+-Cl- co-transport system operates in submandibular glands as it does in other transporting epithelia to achieve K+ uptake.