Absorption of NH(4)(+) by the medullary thick ascending limb (MTAL) is a key event in the renal handling of NH(4)(+), leading to accumulation of NH(4)(+)/NH(3) in the renal medulla, which favors NH(4)(+) secretion in medullary collecting ducts and excretion in urine. The Na(+)-K(+)(NH(4)(+))-2Cl(-) cotransporter (BSC1/NKCC2) ensures approximately 50-65% of MTAL active luminal NH(4)(+) uptake under basal conditions. Apical barium- and verapamil-sensitive K(+)/NH(4)(+) antiport and amiloride-sensitive NH(4)(+) conductance account for the rest of active luminal NH(4)(+) transport. The presence of a K(+)/NH(4)(+) antiport besides BSC1 allows NH(4)(+) and NaCl absorption by MTAL to be independently regulated by vasopressin. At the basolateral step, the roles of NH(3) diffusion coupled to Na(+)/H(+) exchange or Na(+)/NH(4)(+) exchange, which favors NH(4)(+) absorption, and of Na(+)/K(+)(NH(4)(+))-ATPase, NH(4)(+)-Cl(-) cotransport, and NH(4)(+) conductance, which oppose NH(4)(+) absorption, have not been quantitatively defined. The increased ability of the MTAL to absorb NH(4)(+) during chronic metabolic acidosis involves an increase in BSC1 expression, but fine regulation of MTAL NH(4)(+) transport probably requires coordinated effects on various apical and basolateral MTAL carriers.

No abstract

No abstract

To assess whether glucocorticoids regulate rBSC1, the apical Na ؉ -K ؉ (NH 4 ϩ )-2Cl ؊ cotransporter of kidney medullary thick ascending limb (MTAL), studies were performed in normal rats, adrenalectomized (ADX) rats, and ADX rats infused with dexamethasone for 6 days. The effects of dexamethasone on rBSC1 were also studied in vitro using isolated rat MTAL segments. Cotransport activity was estimated by intracellular pH measurements; rBSC1 protein was quantified in MTAL crude membranes by immunoblotting analysis, and mRNA was quantified by quantitative reverse transcription-polymerase chain reaction. The abundance of rBSC1 protein and mRNA increased in ADX rats infused with dexamethasone compared with ADX rats (p < 0.04). In addition, application of dexamethasone for 1-3 h to MTALs caused rBSC1 protein and mRNA abundance and cotransport activity to significantly increase in a hyperosmotic medium (450 mosmol/kg of H 2 O) containing 0.7 nM arginine vasopressin, which is an in vitro experimental condition that resembles the in vivo MTAL environment. Results obtained in various media and with 8-bromocAMP indicated that stimulation of rBSC1 expression by glucocorticoids required interactions between glucocorticoid receptor-and cAMP-dependent factors. Up to 100 nM d-aldosterone had no effect on cotransport activity in vitro. Thus glucocorticoids directly stimulate MTAL rBSC1 expression and activity, which contributes to glucocorticoid-dependent effects on the renal regulation of acid-base balance and urinary concentrating ability.ϩ )-2Cl Ϫ cotransport is responsible for the apical step of NaCl and ammonia transport by the thick ascending limb (TAL) 1 of the nephron. NaCl and ammonia absorption without water by the medullary TAL (MTAL) causes transepithelial concentration differences of these solutes, which constitutes the "single effects" responsible for NaCl and ammonia accumulation in the renal medulla. This is critical both to the level of renal medullary hyperosmolality and thus to the urinary concentrating ability of the kidney and to urinary ammonia excretion and thus to the renal regulation of acid-base balance (1, 2). The MTAL apical Na ϩ -K ϩ (NH 4 ϩ )-2Cl Ϫ cotransporter (BSC1 (bumetanide-sensitive cotransporter) or NKCC2 (Na ϩ -K ϩ -Cl Ϫ cotransporter)) was recently cloned from rat (3), mouse (4, 5), rabbit (6), and human (7) kidneys. The transporter protein has been localized at the apical membrane of the TAL as well as at the macula densa (8 -10). BSC1 was recently shown to be up-regulated by chronic saline loading (9), restriction of water intake and arginine vasopressin (AVP) administration (11), and metabolic acidosis (12) and down-regulated by potassium depletion (13). However, the stimuli and cellular mechanisms of these adaptations of rBSC1 expression were not specified in the latter in vivo studies.The presence of specific glucocorticoid receptors (GR) in the MTAL has been demonstrated by binding, immunological, and mRNA detection methods (14 -17). A number of studies have suggested that in vivo gl...