The effects of Na+ replacement on intracellular pH and [Ca2+] in rabbit salivary gland acinar cells.

Abstract

SUMMARY1. The role of Na+-dependent mechanisms in regulating the intracellular pH (pHi) and free calcium concentration ([Ca2+]i) in acinar cells of the rabbit mandibular salivary gland was examined. The fluorescent dyes BCECF and Fura-2 were used to measure pHi and [Ca2+]i respectively in suspensions of isolated acini.2. Replacement of all of the extracellular Na+ with N-methyl-D-glucamine (NMDG) decreased resting pHi from a control value of 741-7-2 to 6-8-6-9. Re-addition of Na+ or Li+ caused a recovery of pHi towards control values. This recovery was blocked by 10-50 ,uM-ethylisopropylamiloride (EIPA), suggesting that it was mediated by Na+-H+ exchange. The rate of recovery of pHi when Na+ was reintroduced increased with Na+ concentration with an apparent Km for Na+ of around 30 mM.3. Replacement of all of the extracellular Na+ with Li+ caused only a small decrease in resting pHi.4. Stimulation of acini with 1 uM-acetylcholine (ACh) evoked an intracellular acidosis both under control conditions and when acini were bathed in Na+-free media. Following the acidosis pHi recovered in acini bathed in either control medium or Na+-free (Li+) medium, but not in acini bathed in Na+-free (NMDG) medium or in control medium containing EIPA.5. Stimulation of acini bathed in Na+-free, HC03--free medium with ACh did not cause any change in pHi.6. Re-addition of Na+ to acini bathed in Na+-free, HC03--free medium evoked the same rate of alkalinization whether or not the acini had been stimulated with ACh, suggesting that receptor stimulation per se did not lead to an activation of acid extrusion.7. Resting [Ca2+]i was elevated in acini bathed in Na+-free (NMDG) medium, but not in acini bathed in Na+-free (Li+) medium.8. ACh evoked a maintained rise in [Ca2+]i in acini bathed in control medium and in Na+-free media with either NMDG or Li' as the Na+ substitute.9. Experiments in which external Ca2+ was reduced to low levels (by the addition of EGTA) just prior to addition of ACh showed that ACh released intracellular Ca2+ stores under both control and Na+-free conditions. 10. In acini bathed in Na+-free (NMDG) [Ca2+]i could also be reduced under these conditions by alkalinizing the cytosol using the weak base trimethylamine. 11. Our results suggest that Na+-H+ exchange is the major acid extrusion mechanism in isolated rabbit mandibular acinar cells. The exchanger is responsible for maintaining pHi above equilibrium, and also underlies the recovery of pH, when the cytosol is acidified by stimulation with ACh.12. Removal of external Na+ does not appear to interfere with receptor-operated Ca mobilization in rabbit mandibular acinar cells. The effects of Na+ removal and readdition on [Ca2+]i appear to be secondary to the changes in pHi, and are most easily explained by a direct interaction between Ca2+ and H+.

Lowering Extracellular pH Raises Intracellular Calcium in Cultured Rat Astrocytes.

“…The close interrelationship has been reported between intracellular calcium concentration ([Ca2+]i) and intracellular pH (pHi) in various cells [3,10,16,18]. The effect of pHi is possibly elicited through the effect on [Ca2+]i. pHo may also affect the glial cells via the regulation of pHi and [Ca2+]i.…”

“…Previous studies on a variety of cells have shown that a reduction in pHo results in increases in [Ca2+]i [3,10,16,18,25,29,31]. It is difficult, however, to investigate the effect on changes in [Ca2+]i of pHo or pHi separately, since the latter seems to be highly dependent on the former [19,20,28].…”

Physiological Pathway of Magnesium Influx in Rat Ventricular Myocytes

“…This apparent dependence on extracellular Na þ could be attributed to intracellular acidosis, which inhibits TRPM7 channel activities (21). Elliott et al (29) reported that the inhibition of Na þ /H þ exchange by replacement of extracellular Na þ with NMDG caused a decrease in intracellular pH (pH i ) to~6.8 in rabbit salivary gland acinar cells. Wu et al (30) also reported that similar acidification was induced by Na þ -free solution in single rat cardiac myocytes.…”

Intracellular alkalinization mobilizes calcium from agonist‐sensitive pools in rat lacrimal acinar cells.

“…One possibility is that the intracellular alkalinization might inhibit the Ca2P entry pathway. Alkalinizing the cytoplasm with trimethylamine has previously been shown to reduce [Ca2+]i in agonist-stimulated salivary acinar cells, consistent with such an inhibitory effect (Elliott, Lau & Brown, 1991). In order to test whether intracellular alkalinization altered Ca2+ entry following depletion of intracellular Ca2P pools we employed thapsigargin.…”