Intracellular Cl⁻ concentration in striated intralobular ducts from rabbit mandibular salivary glands

Abstract

Intralobular striated ducts have been isolated from rabbit mandibular salivary glands and maintained in primary culture for up to 2 days. Such ducts were loaded with the Cl⁻-sensitive fluorescent dyeN-(ethoxycarbonylmethyl)-(6-methoxyquinolinium bromide) (MQAE) and intracellular Cl⁻ concentration ([Cl⁻]_i monitored using a fluorescence microscope. Intracellular Cl⁻ could be rapidly and reversibly emptied from striated duct cells by replacing Cl⁻ in the superfusing solution with NO ⁻₃ . [Cl⁻]_i could be lowered by removal of external Na⁺, exposure to 10 μM amiloride or to 10 μM 4,4′-diisothiocyanatostilbene-2,2′-disulphonic acid (DIDS). Both amiloride and DIDS were able to inhibit the recovery of [Cl⁻]_i after an initial exposure to Na⁺- or Cl⁻-free solution. The amiloride derivatives, benzamil (2 μM) and N-isobutyl-N-methylamiloride (MIBA), (10 μM) also lowered [Cl⁻]_i by similar amounts as 10 μM amiloride. Varying external K⁺ concentration ([K⁺]_o) also affected [Cl⁻]_i. Increasing [K⁺]_o increased [Cl⁻]_i, but decreasing [K⁺]_o did not decrease [Cl⁻]_i. Instead, [Cl⁻]_i was also increased when [K⁺]_o was lowered below the control value. Bumetanide (0.1 mM) lowered [Cl⁻]_i by only a small amount, while ouabain (1 mM) had no significant effect on [Cl⁻]_i. These data are consistent with current models of electrolyte transport in salivary ducts which include Cl⁻ channels, Na⁺ channels, and Na⁺/H⁺ exchangers in the apical membrane. The effects of low [K⁺]_o can be interpreted in terms of a K⁺-dependent exit mechanism for Cl⁻.