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Gastrointestinal Division, Departments of Medicine and Physiology, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205-2195
Intestinal neutral NaCl absorption, which is made up of
brush-border (BB)
Na+/H+
exchange linked to BB
Cl
/HCO3
exchange, is up- and downregulated as part of digestion and diarrheal
diseases. Glucocorticoids stimulate ileal NaCl absorption and BB
Na+/H+
exchange. Intestinal BB contains two
Na+/H+
exchanger isoforms, NHE2 and NHE3, but their relative roles in rabbit
ileal BB
Na+/H+
exchange has not been determined. A technique to separate the contribution of NHE2 and NHE3 to ileal BB
Na+/H+
exchange activity was standardized by using an amiloride-related compound, HOE-694. Under basal conditions, both NHE2 and NHE3 contribute ~50% to ileal
Na+/H+
exchange. Glucocorticoids (methylprednisolone) increase BB
Na+/H+
exchange (2.5 times) but increase only ileal NHE3 activity (4.1 times),
without an effect on NHE2 activity. Thus ileal BB
Na+/H+
exchange in animals treated with glucocorticoids is 69% via NHE3. A
quantitative Western analysis for NHE3 was developed, using as an
internal standard a fusion protein of the COOH-terminal 85 amino acids
of NHE3 and maltose binding protein. Glucocorticoid treatment increased
the amount of BB NHE3. The quantitative Western analysis showed that
NHE3 makes up 0.018% of ileal BB protein in control rabbits and
0.042% (2.3 times as much) in methylprednisolone-treated rabbits.
Methylprednisolone treatment did not alter the amount of ileal BB NHE2
protein. NHE3 turnover number was estimated to be 458 cycles/s under
basal conditions and 708 cycles/s in glucocorticoid-treated ileum. Thus
methylprednisolone stimulates ileal BB
Na+/H+
exchange activity only by an effect on NHE3 and not on NHE2; it does so
primarily by increasing the amount of BB NHE3, although it also
increases the NHE3 turnover number.
HOE-694; plasma membrane vesicle; quantitative Western analysis; turnover number
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