human check cell
Springer Online Journal Archives 1860-2000
Chemistry and Pharmacology
Abstract Proton-dependent, ethylisopropylamiloride (EIPA)-sensitive Na+ uptake (Na+/H+ antiporter) studies were performed to examine if saliva, and ionophores which alter cellular electrolyte balance, could influence the activity of the cheek cell Na+/H+ antiporter. Using the standard conditions of 1 mmol/1 Na+, and a 65:1 (inside:outside) proton gradient in the assay, the uniport ionophores valinomycin (K+) and gramicidin (Na+) increased EIPA-sensitive Na+ uptake by 177% (p 〈 0.01) and 227% (p 〈 0.01), respectively. The dual antiporter ionophore nigericin (K+-H+) increased EIPA-sensitive Na+ uptake by 654% (p 〈 0.01), with maximal Na+ uptake achieved by 1 min and at an ionophore concentration of 50 μmol/l, with an EC 50 value 6.4 μmol/l. Preincubation of cheek cells with saliva or the low molecular weight (MW) components of saliva (saliva activating factors, SAF) for 2 h at 37°C, also significantly stimulated EIPA-sensitive Na+ uptake. This stimulation could be mimicked by pre-incubation with 25 mmol/l KCl or K+-phosphate buffer. Pre-incubating cheek cells with SAF and the inclusion of 20 μmol/1 nigericin in the assay, produced maximum EIPA-sensitive Na+ uptake. After pre-incubation with water, 25 mmol/1 K+-phosphate or SAF, with nigericin in all assays, the initial rate of proton-gradient dependent, EIPA-sensitive Na+ uptake was saturable with respect to external Na+ with Km values of 0.9, 1.7, and 1.8 mmol/l, and V max values of 13.4, 25.8, and 31.1 nmol/mg protein/30 sec, respectively. With 20 μmol/1 nigericin in the assay, Na+ uptake was inhibited by either increasing the [K+]o in the assay, with an ID 50 of 3 mmol/l. These results indicate that nigericin can facilitate K+ i exchange for H+ o and the attending re-acidification of the cheek cell amplifies IINa+ uptake via the Na+/H+ antiporter. The degree of stimulation of proton-dependent, EIPA-sensitive Na+ uptake is therefore dependent, in part, on the intracellular K+ i.
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