MODULATION OF NA-H+ EXCHANGE BY ALTERED CELL-VOLUME IN PERFUSED RAT MANDIBULAR SALIVARY-GLAND()

1. Intracellular pH (pH(i)) was measured by spectrofluorometry in perf used mandibular salivary glands isolated from the rat and loaded with the pH-sensitive fluoroprobe 2',7'-bis(2-carboxyethyl)-5(6)-carboxyflu orescein (BCECF). Cell volume changes were estimated from changes in i ntracellular water content measured by proton NMR spectroscopy. 2. Sti mulation with 1 mu M acetylcholine (ACh) led to a 15 +/- 2% decrease i n cell volume. A transient decrease in pH(i) was followed by a sustain ed increase (0.17 +/- 0.03 pH units) that has previously been attribut ed to the upregulation of the Na+-H+ exchanger. 3. Increasing perfusat e osmolarity by addition of 60 mM sucrose caused a 19 +/- 2% decrease in cell volume and a sustained increase in pH(i) (0.12 +/- 0.01 pH uni ts) that was abolished by 1 mM amiloride. Acid loading experiments ind icated that the increase in pH(i) was due to an alkaline shift in the pH dependence of the Na+-H+ exchanger. 4. A 20% reduction in perfusate osmolarity prevented the cell shrinkage normally associated with ACh stimulation and largely abolished the ACh-induced increase in pH(i). 5 . Steady-state Na+-H+ exchanger activity, estimated from the initial r ate of change in pH(i) following addition of amiloride, increased 9-fo ld during stimulation with ACh. When cell shrinkage was prevented by s imultaneous exposure to the hypotonic solution, the activity of the ex changer still increased 7-fold in response to ACh. 6. We conclude that , although cell shrinkage leads to upregulation of the Na+-H+ exchange r, this factor alone is insufficient to account for the marked increas e in exchanger activity that follows muscarinic stimulation.