KV7.1 channel blockade inhibits neonatal renal autoregulation triggered by a step decrease in arterial pressure

Abstract

K_V7 channels, the voltage-gated K⁺ channels encoded by KCNQ genes, mediate heterogeneous vascular responses in rodents. Postnatal changes in the functional expression of K_V7 channels have been reported in rodent saphenous arteries, but their physiological function in the neonatal renal vascular bed is unclear. Here, we report that, unlike adult pigs, only KCNQ1 (K_V7.1) out of the five members of KCNQ genes was detected in neonatal pig renal microvessels. KCNQ1 is present in fetal pig kidneys as early as day 50 of gestation, and the level of expression remains the same up to postnatal day 21. Activation of renal vascular smooth muscle cell (SMC) K_V7.1 stimulated whole cell currents, inhibited by HMR1556 (HMR), a selective K_V7.1 blocker. HMR did not change the steady-state diameter of isolated renal microvessels. Similarly, intrarenal artery infusion of HMR did not alter mean arterial pressure, renal blood flow, and renal vascular resistance in the pigs. An ∼20 mmHg reduction in mean arterial pressure evoked effective autoregulation of renal blood flow, which HMR inhibited. We conclude that 1) the expression of KCNQ isoforms in porcine renal microvessels is dependent on kidney maturation, 2) K_V7.1 is functionally expressed in neonatal pig renal vascular SMCs, 3) a decrease in arterial pressure up to 20 mmHg induces renal autoregulation in neonatal pigs, and 4) SMC K_V7.1 does not control basal renal vascular tone but contributes to neonatal renal autoregulation triggered by a step decrease in arterial pressure.NEW & NOTEWORTHY K_V7.1 is present in fetal pig kidneys as early as day 50 of gestation, and the level of expression remains the same up to postnatal day 21. K_V7.1 is functionally expressed in neonatal pig renal vascular smooth muscle cells (SMCs). A decrease in arterial pressure up to 20 mmHg induces renal autoregulation in neonatal pigs. Although SMC K_V7.1 does not control basal renal vascular resistance, its inhibition blunts neonatal renal autoregulation engendered by a step decrease in arterial pressure.

Keywords: KCNQ; KV7.1; neonates; renal autoregulation; smooth muscle cells.

Graphical abstract

Figure 1.

KCNQ1 is the predominant isoform in the neonatal pig kidney and renal vascular smooth muscle cells (SMCs). A: agarose gel images demonstrating the amplification of KCNQ isoforms in the neonatal pig (3–5 days old) kidneys, brain, heart, and pulmonary arteries. B: bar graphs summarizing mean data for quantitative RT-PCR experiments that compared KNCQ1 mRNA expression levels in fetal pig kidneys at gestational days 50 and 100 (GD50 and GD100, respectively) and postnatal days 1 and 21 (P1 and P21, respectively). C: agarose gels images demonstrating amplification of α-smooth muscle actin (ACTA2), von Willebrand factor (vWF), and KCNQ1 in interlobular arteries (ILA) and ACTA2 and KCNQ1 in ILA SMCs. D: agarose gel image demonstrating the amplification of KCNQ isoforms in adult pigs. E: confocal microscopy images of ILA SMCs immunostained with a selective K_V7.1 blocking peptide (BP) + K_V7.1 and caveolin-1 antibodies. F: images showing colocalization of caveolin-1 and K_V7.1 channels in the plasma membrane of SMCs. Product sizes for KNCQ1, KNCQ2, KNCQ3, KNCQ4, and KNCQ5 are 213, 207, 186, 262, and 110, respectively (one-way ANOVA with a Bonferroni’s post hoc test). ns, not significant; NTC, no template control.

Figure 2.

K_V7.1 is functionally expressed in neonatal pig renal vascular smooth muscle cells (SMCs). A: linopirdine (Lino; 200 µM) inhibited outward currents in renal vascular SMCs. B: current-voltage curves of whole cell currents in control and Lino-treated cells. C: current-voltage curve of Lino-sensitive current. D: HMR1556 (HMR) inhibited outward currents in a concentration-dependent manner. E: concentration-dependent curve fit of HMR shown on a semilog plot. The y axis represents the uninhibited current normalized to the mean of control (I/I_control) at membrane voltages of +20, +30, and +40 mV. F and G: whole cell currents in control and ML277 (1 µM)-treated renal vascular SMCs. H and I: mefenamic acid (MA; 300 nM)-induced whole cell currents in renal vascular SMCs, which were inhibited by HMR (3 µM). *P < 0.05 (two-way ANOVA with a Bonferroni’s post hoc test).

Figure 3.

K_V7.1 does not control the basal renal vascular tone of neonatal pigs. A trace (A) and bar chart (B) showing the effect of phenylephrine (PE; 10 µM) and HMR1556 (HMR; 10 µM) on pressurized interlobular arteries. C–F: traces and bar charts demonstrating that intrarenal artery infusion of HMR (20 ng/kg/min for 20 min) did not alter basal mean arterial pressure (MAP), renal blood flow (RBF), and renal vascular resistance (RVR). *P < 0.05 vs. baseline (two-tailed paired t test). ns, not significant.

Figure 4.

A step decrease in arterial pressure triggered renal autoregulation in neonatal pigs. A and B: traces and bar charts showing that an ∼20-mmHg decrease in mean arterial pressure (MAP) triggered renal autoregulation. C: traces demonstrating that renal autoregulation was ineffective by decreasing MAP by ∼23 mmHg. D: renal autoregulation indexes at ∼20 and 23 mmHg. *P < 0.05 (two-tailed unpaired t test). RBF, renal blood flow.

Figure 5.

Inhibition of K_V7.1 channels attenuated renal autoregulation in neonatal pigs by opposing vasodilation. A and B: traces illustrating the response of renal blood flow (RBF) to an ∼20 mmHg reduction in mean arterial pressure in the absence and presence of HMR1556 (HMR; 20 ng/kg/min for 20 min). C: bar charts summarizing renal autoregulation indexes in the absence and presence of HMR [5 ng/kg/min (a), 10 ng/kg/min (b), and 20 ng/kg/min (c) for 20 min]. D: HMR dose-response fit for the autoregulation index. *P < 0.05 (one-way ANOVA with Bonferroni’s post hoc test). MAP, mean arterial pressure.