Genomic and nongenomic stimulatory effect of aldosterone on H⁺-ATPase in proximal S3 segments

Abstract

The genomic and nongenomic effects of aldosterone on the intracellular pH recovery rate (pHirr) via H+-ATPase and on cytosolic free calcium concentration ([Ca2+]i) were investigated in isolated proximal S3 segments of rats during superfusion with an Na+-free solution, by using the fluorescent probes BCECF-AM and FLUO-4-AM, respectively. The pHirr, after cellular acidification with a NH4Cl pulse, was 0.064 ± 0.003 pH units/min ( n = 17/74) and was abolished with concanamycin. Aldosterone (10−12, 10−10, 10−8, or 10−6 M with 1-h or 15- or 2-min preincubation) increased the pHirr. The baseline [Ca2+]i was 103 ± 2 nM ( n = 58). After 1 min of aldosterone preincubation, there was a transient and dose-dependent increase in [Ca2+]i and after 6-min preincubation there was a new increase in [Ca2+]i that persisted after 1 h. Spironolactone [mineralocorticoid (MR) antagonist], actinomycin D, or cycloheximide did not affect the effects of aldosterone (15- or 2-min preincubation) on pHirr and on [Ca2+]i but inhibited the effects of aldosterone (1-h preincubation) on these parameters. RU 486 [glucocorticoid (GR) antagonist] and dimethyl-BAPTA (Ca2+ chelator) prevented the effect of aldosterone on both parameters. The data indicate a genomic (1 h, via MR) and a nongenomic action (15 or 2 min, probably via GR) on the H+-ATPase and on [Ca2+]i. The results are compatible with stimulation of the H+-ATPase by increases in [Ca2+]i (at 10−12-10−6 M aldosterone) and inhibition of the H+-ATPase by decreases in [Ca2+]i (at 10−12 or 10−6 M aldosterone plus RU 486).

Action of ANP on the nongenomic dose-dependent biphasic effect of aldosterone on NHE1 in proximal S3 segment

“…These results are in accordance with previous data published by our laboratory [5]. Our results also indicate that during the superfusion of the S3 segment with a zero Na + solution (which inhibits the activity of the Na + /H + exchanger), a small pHirr still was observed, which was abolished by concanamycin (a H + -ATPase inhibitor); these data agree with recently published results [27] showing that in the S3 segment the pHi recovery also occurs via H + -ATPase. However, in our present study and recently published work [27], the activity of this transporter begins about 2.5 min after the acid pulse and does not reach the basal pHi.…”

ANG II, ANG-(1-7), ALDO and AVP biphasic effects on Na+/H+ transport: the role of cellular calcium

“…Apical Na + channel activity is the limiting step in the transport process, and it is likely that ALDO ultimately acts to increase the open time of existing ion channels and/ or increase the total number of such channels [225]. However, other protein targets have also been identified, including i) a luminal NHE3 exchanger in the colon [226,227] and the proximal tubule [228] and a basolateral NHE1 in the proximal tubule [229,230], ii) an Na + /K + -ATPase in human kidney proximal tubule (HKC11) cells [231], iii) a luminal thiazide-sensitive Na + /Cl − cotransporter in the distal renal tubule that appears to mediate Na+ reabsorption in response to volume depletion [232], and iv) a renal proximal H + -ATPase [233].…”

“…The physiological and clinical relevance supporting these rapid effects remains unclear, but their existence has been described in various target organs and cells, including amphibian skin and urinary bladders [258,259], vascular smooth muscle cells and endothelial cells [260], skeletal muscle cells [261], human mononuclear leukocytes [262], cardiac myocytes [263], skin fibroblasts from MR-knockout mice [256], colonic epithelial cells [264] and isolated colonic crypts [265]. Several sites in the kidney, particularly cultured kidney cells, have been shown to be sensitive to non-genomic ALDO action [266], including principal cells that were freshly isolated from rabbits [267], the human distal colon [268], in vivo renal proximal tubules (S2 segment) [228], isolated renal proximal tubules (S3 segment) [229,233], medullary thick ascending limbs [269] and renal collecting duct cells [270]. Its non-genomic actions include effects on signal transduction pathways and ion transporters, such as the epithelial Na + channel [267], the Na + /H + exchanger [228,229,271,272] and the vacuolar H + -ATPase [230,233,258,259,273].…”

“…Several sites in the kidney, particularly cultured kidney cells, have been shown to be sensitive to non-genomic ALDO action [266], including principal cells that were freshly isolated from rabbits [267], the human distal colon [268], in vivo renal proximal tubules (S2 segment) [228], isolated renal proximal tubules (S3 segment) [229,233], medullary thick ascending limbs [269] and renal collecting duct cells [270]. Its non-genomic actions include effects on signal transduction pathways and ion transporters, such as the epithelial Na + channel [267], the Na + /H + exchanger [228,229,271,272] and the vacuolar H + -ATPase [230,233,258,259,273].…”

“…Additionally, it has been shown that through both genomic and non-genomic mechanisms, ALDO stimulates Na + reabsorption, K + secretion [280], Na + /H + exchange [230] and H + -ATPase [233] in the kidney, and Na + /H + exchange in vascular smooth muscle cells [281].…”