FC018: The B1 H+-ATPASE (ATP6V1B1) Subunit is Required for Non-Type a Intercalated Cell Function During Alkalosis
Authors: Bourgeois Soline , Kovacikova Jana , Bugarski Milika , Bettoni Carla , Hall Andrew , Wagner Carsten 
Published: May 2022

Abstract

BACKGROUND AND AIMS
Non-type A intercalated cells (IC) in the connecting tubule (CNT) and cortical collecting duct (CCD) express the luminal Cl−/HCO3− exchanger pendrin and apical and/or basolateral vacuolar H+-ATPases containing the B1 subunit isoform. One of the main functions of non-type A ICs is the secretion of bicarbonate during metabolic alkalosis. Mutations in the B1 subunit (ATP6V1B1) in man cause distal renal tubular acidosis due to its importance in acid secretion by type A ICs. However, the function of the B1 isoform in non-type A ICs has remained elusive. In this work, we investigated the role of the B1 H+-ATPase subunit in non-type A intercalated in the control of alkalosis.

METHOD
Acid-base metabolism of Atp6v1b1 mice was studied during an alkali load (0.28 M NaHCO3 in drinking water and 2 mg deoxycorticosterone i.p for 4 days) or during a chronic furosemide treatment (20 mg i.p. injection daily for 9 days).

RESULTS
Induction of metabolic alkalosis by 0.28 M NaHCO3 resulted in a more pronounced alkalosis in ATP6v1b1−/− mice with increased blood bicarbonate, hypokalaemia and hypochloraemia despite a reduced lung minute volume. Determination of the relative abundance of the different subtypes of cells in the collecting duct system revealed a remodelling of CNT and CCD in the cortex with an increase of type A ICs and a compensatory increase of non-type A ICs in collecting ducts of the outer medullary in ATP6v1b1−/− kidneys. Total pendrin expression in the kidney and activity in non-type A cells of ex vivo microperfused CCD were reduced in ATP6v1b1−/− mice. Moreover, the stimulating effect of Isoproterenol, a β2-adrenergic receptor agonist stimulating cAMP production, on pendrin activity was blunted in non-type A ICs from ATP6v1b1−/− mice. Basolateral H+-ATPase activity in pendrin-expressing cells was strongly reduced, even though the expression of the B2 isoform was increased at their basolateral side. Moreover, the E1 and A H+-ATPase subunits of the V0 domain did only partially colocalize with the a4 H+-ATPase subunit of the V1 domain of H+-ATPases at the basolateral pole of ATP6v1b1−/− non-type A ICs, indicating impaired assembly of V0 and V1 H+-ATPase domains. Finally, when exposed to a chronic furosemide treatment, a treatment associated with a high risk to develop metabolic alkalosis in patients, ATP6v1b1−/− mice developed a more pronounced hypokalaemic hypochloraemic alkalosis with polyuria and loss of urine chloride compared with their wildtype littermates. In ATP6v1b1−/− kidneys, pendrin expression was drastically reduced as well as the water channel AQP2 and the α and γ subunits of the sodium channel ENaC. An increase of BUN and a decrease of kidney klotho expression were also observed in ATP6v1b1−/− mice which might indicate a worsening of their kidney function.

CONCLUSION
Thus, this study demonstrates for the first time that i) the B1 H+-ATPase subunit is critical for normal non-type A ICs function and protects against alkalosis. ii) Moreover, the B1 subunit is required for the assembly of complete and functional basolateral vacuolar H+-ATPases complexes in intercalated cells.