The kidneys play pivotal roles in acid-base homeostasis, and the acid-secreting (␣-type) and bicarbonate-secreting (␤-type) intercalated cells in the collecting ducts are major sites for the final modulation of urinary acid secretion. Since the H ؉ -ATPase and anion exchanger activities in these two types of intercalated cells exhibit opposite polarities, it has been suggested that the ␣-and ␤-intercalated cells are interchangeable via a cell polarity change. Immunohistological studies, however, have failed to confirm that the apical anion exchanger of ␤-intercalated cells is the band 3 protein localized to the basolateral membrane of ␣-intercalated cells. In the present study, we show the evidence that a novel member of the anion exchanger and sodium bicarbonate cotransporter superfamily is an apical anion exchanger of ␤-intercalated cells. Cloned cDNA from the ␤-intercalated cells shows about 30% homology with anion exchanger types 1-3, and functional expression of this protein in COS-7 cells and Xenopus oocytes showed sodium-independent and 4,4-diisothiocyanostilbene-2,2-disulfonic acid-insensitive anion exchanger activity. Furthermore, immunohistological studies revealed that this novel anion exchanger is present on the apical membrane of ␤-intercalated cells, although some ␤-intercalated cells were negative for AE4 staining. We conclude that our newly cloned transporter is an apical anion exchanger of the ␤-intercalated cells, whereas our data do not exclude the possibility that there may be another form of anion exchanger in these cells.

Vacuolar H+-ATPases have an essential role in renal hydrogen ion secretion in the proximal tubule, collecting duct, and other segments of the nephron. Control of H+ transport is achieved by variations in the intrinsic properties of the renal H+-ATPases and by several cellular regulatory mechanisms, including redistribution of the enzyme both by vesicular traffic and regulated assembly and disassembly, and cytosolic regulatory proteins that interact directly with H+-ATPase. These mechanisms may provide a means for fine control of net acid excretion and for regulating vacuolar H+-ATPases residing on the plasma membrane independently from those in intracellular compartments.

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Since Smith's time, the essential role of collecting duct intercalated cells in controlling net acid excretion has been recognized. Rather than employing an H(+)-exchange mechanism, intercalated cells have V-ATPase on the plasma membrane and in plasmalemma-associated tubulovesicles, which functions in the bicarbonate reabsorption, regeneration, and bicarbonate secretion required for acid-base homeostasis. Several distinct mechanisms participate in regulating V-ATPase-driven H+ secretion in different cell types: (1) Renal epithelial cells have the capacity to express different structural forms of V-ATPase that have intrinsic differences in their enzymatic properties. 2) The kidney produces cytosolic regulatory proteins, capable of interacting directly with the V-ATPase, that may modify its activity. V-ATPases in different cell types may differ in the degree to which their activity is affected by regulatory factors, as a result of variations in V-ATPase structure. (3) In the alpha intercalated cell, the number of active V-ATPases on the luminal membrane is controlled in vivo by membrane vesicle-mediated traffic that may require unidentified mediators. In the beta intercalated cell, the number of active V-ATPases on the basolateral membrane may be controlled by regulated assembly and disassembly, responding directly to extracellular pH.

To examine the mechanisms which regulate the functions ofthe intercalated cells (ICs) in the cortical collecting duct (CCD), the effect of isoproterenol on intracellular pH (pHi) of ICs was studied with the in vitro microperfused rabbit CCD, using the single cell pHi determination technique with fluorescent dye, 2',7'-bis-(2-carboxyethyl)-5(and-6)carboxyfluorescein. The pHi of #-IC was significantly decreased with the addition of basolateral 106 M isoproterenol (7.21±0.04 to 7.05±0.04), whereas a-IC did not show any change. This response of t-IC to isoproterenol was dose-dependent and completely inhibited by the j8blockers, atenolol or propranolol. The addition offorskolin or 8-Br-cAMP mimicked the effects of isoproterenol, suggesting that the activation of adenylate cyclase induced the decrease in pHi. The rate of pHi changes after the Cl-removal from the perfusate, which is considered to reflect the activity of luminal anion exchanger, was significantly higher with isoproterenol (0.032±0.009 pH unit/s) than that in the control (0.023±0.009 pH unit/s). The present studies provide direct evidence for the regulation of #-IC function by fl-adrenergic receptor, and the luminal Cl-/HCO; exchanger was considered to be stimulated by e-agonist, directly. (J. Clin. Invest. 1991.