Abstract
Intracellular pH (pHi) was measured in acini isolated from rabbit mandibular salivary glands using the fluorescent pH-sensitive probe 2,7-bis(carboxyethyl)-5(6)-carboxyfluorescein (BCECF). Resting pHi was estimated to be 7.13 +/- 0.01 (mean +/- SE of 29 experiments). Stimulation with acetylcholine (ACh) caused an intracellular acidosis followed by a return of pHi toward the control value with a half time of approximately 3 min. The intracellular acidosis was dose dependent and could be abolished by pretreatment of the acini with atropine (10 microM), suggesting that it was due to a receptor-mediated event. Incubation of the acini in HCO3- -free solutions or treatment of the acini with the carbonic anhydrase inhibitor acetazolamide (1 mM) abolished the acidosis, suggesting that the acidosis might be caused by loss of HCO3- from the cell. The acidosis was not affected by either 1) pretreatment of the acini with the anion exchange inhibitor 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS), or 2) equilibration of the acini in Cl- -free solution (Cl- substituted with glucuronate). These results suggest that the postulated HCO3- efflux does not occur by Cl- -HCO3- exchange. However, Cl- -HCO3- exchange did appear to be present because replacement of Cl- caused a large DIDS-sensitive alkalinization of pHi, presumably caused by HCO3- uptake in exchange for Cl-. The recovery of pHi after the initial acidosis on stimulation with ACh could be blocked by 1 mM amiloride, suggesting that the recovery phase was mediated by Na+-H+ exchange.
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