Aldosterone sensitizes connecting tubule glomerular feedback via the aldosterone receptor GPR30

Abstract

Increasing Na delivery to epithelial Na channels (ENaC) in the connecting tubule (CNT) dilates the afferent arteriole (Af-Art), a process we call connecting tubule glomerular feedback (CTGF). We hypothesize that aldosterone sensitizes CTGF via a nongenomic mechanism that stimulates CNT ENaC via the aldosterone receptor GPR30. Rabbit Af-Arts and their adherent CNTs were microdissected and simultaneously perfused. Two consecutive CTGF curves were elicited by increasing luminal NaCl in the CNT. During the control period, the concentration of NaCl that elicited a half-maximal response (EC50) was 37.0 ± 2.0 mmol/l; addition of aldosterone 10(-8) mol/l to the CNT lumen caused a left-shift (decrease) in EC50 to 19.3 ± 1.3 mmol/l (P = 0.001 vs. control; n = 6). Neither the transcription inhibitor actinomycin D nor the translation inhibitor cycloheximide prevented the effect of aldosterone (control EC50 = 34.7 ± 1.9 mmol/l; aldosterone+actinomycin D EC50 = 22.6 ± 1.6 mmol/l; P < 0.001 and control EC50 = 32.4 ± 4.3 mmol/l; aldosterone+cycloheximide EC50 = 17.4 ± 3.3 mmol/l; P < 0.001). The aldosterone antagonist eplerenone prevented the sensitization of CTGF by aldosterone (control EC50 = 33.2 ± 1.7 mmol/l; aldosterone+eplerenone EC50 = 33.5 ± 1.3 mmol/l; n = 7). The GPR30 receptor blocker G-36 blocked the sensitization of CTGF by aldosterone (aldosterone EC50 = 16.5 ± 1.9 mmol/l; aldosterone+G-36 EC50 = 29.0 ± 2.1 mmol/l; n = 7; P < 0.001). Finally, we found that the sensitization of CTGF by aldosterone was mediated, at least in part, by the sodium/hydrogen exchanger (NHE). We conclude that aldosterone in the CNT lumen sensitizes CTGF via a nongenomic effect involving GPR30 receptors and NHE. Sensitized CTGF induced by aldosterone may contribute to renal damage by increasing Af-Art dilation and glomerular capillary pressure (glomerular barotrauma).

Keywords: GPR30 receptor; MR receptor; aldosterone; arterioles; distal kidney tubules.

Fig. 1.

A: increasing NaCl concentration 2 consecutive times in the connecting tubule (CNT) dilated preconstricted afferent arterioles (Af-Arts) in a similar manner, indicating that connecting tubule glomerular feedback (CTGF) is stable and reproducible over time. B: adding 10⁻⁸ mol/l aldosterone to the CNT perfusate sensitized CTGF (*P < 0.05, **P < 0.01, ***P < 0.001; with vs. without aldosterone).

Fig. 2.

A: adding 5 × 10⁻⁶ mol/l actinomycin D to the CNT perfusate did not prevent the sensitization of CTGF by aldosterone, suggesting that the effect of aldosterone on CTGF is not genomic. **P < 0.01; ***P < 0.001, control vs. aldosterone plus actinomycin D. B: actinomycin D alone did not affect CTGF in the absence of exogenous aldosterone.

Fig. 3.

A: adding 10⁻⁵ mol/l cycloheximide to the CNT perfusate did not prevent the sensitization of CTGF by aldosterone, suggesting that the effect of aldosterone on CTGF is not genomic. **P < 0.01; ***P < 0.001, control vs. aldosterone plus cycloheximide. B: cycloheximide alone did not affect CTGF in the absence of exogenous aldosterone.

Fig. 4.

A: adding 10⁻⁵ mol/l eplerenone to the CNT perfusate prevented the sensitization of CTGF by aldosterone, suggesting that this effect is mediated by either the mineralocorticoid receptor (MR) or G protein-coupled receptor 30 (GPR30). B: eplerenone alone did not affect CTGF in the absence of exogenous aldosterone.

Fig. 5.

A: adding 10⁻⁵ mol/l G36 to the CNT perfusate attenuated the sensitization of CTGF by aldosterone, suggesting that this effect is mediated, at least in part, by GPR30 (**P < 0.01; ***P < 0.001, aldosterone vs. aldosterone plus G36). B: adding G36 to the CNT perfusate completely prevented the sensitization of CTGF by aldosterone, indicating that this effect is mainly mediated by GPR30. C: G36 alone did not affect CTGF in the absence of exogenous aldosterone.

Fig. 6.

A: adding 10⁻⁶ mol/l benzamil to the CNT perfusate did not prevent aldosterone from inducing CTGF, suggesting that this nongenomic effect of aldosterone is, at least in part, independent from ENaC (*P < 0.05, **P < 0.01, aldosterone vs. aldosterone plus benzamil; ##P < 0.01 vs. 0 NaCl). B: adding 10⁻⁴ mol/l dimethylamiloride (DMA) and benzamil to the CNT perfusate completely abolished both basal CTGF and aldosterone-induced sensitization of CTGF (***P < 0.001, aldosterone vs. aldosterone plus DMA).

Fig. 7.

A: adding 10⁻⁴ mol/l DMA to the CNT perfusate completely prevented the sensitization of CTGF by aldosterone, indicating that this effect is mediated by the sodium/hydrogen exchanger. B: DMA alone did not affect CTGF in the absence of exogenous aldosterone, indicating that the sodium/hydrogen exchanger does not participate in basal CTGF.