A novel sodium signaling complex regulates uterine activity

Abstract

At the end of pregnancy, the uterus transitions from a quiescent state to an excitable, contractile state. These changes are linked to depolarization of the myometrial smooth muscle cell (MSMC) resting membrane potential. The membrane potential is primarily determined by the balance between an outward potassium (K+) leak current and an inward sodium (Na+) leak current. We recently described a Na+-activated K+ channel (SLO2.1) and a non-selective Na+ leak channel (NALCN) in human MSMCs. Here, we asked whether these channels function together. We show that SLO2.1 currents are activated by an inward NALCN-dependent Na+ leak current, leading to MSMC hyperpolarization. The regulation of the membrane potential by NALCN/SLO2.1 activity modulates both Ca2+ entry through VDCCs, and myometrial contractility. Finally, NALCN and SLO2.1 are in proximity to one another in human MSMCs. We conclude that SLO2.1 and NALCN function together to regulate human MSMC membrane potential and excitability.

SLO2.1/NALCN Functional Complex Activity in Mouse Myometrial Smooth Muscle Cells During Pregnancy

“…Treatment with 80 mM Li + , a NALCN-permeable cation that does not activate SLO2.1, depolarized MSMCs from non-pregnant mice (Figure 5c). Similar experiments with the impermeable cation Choline also showed no membrane hyperpolarization, indicating that the Na + effect was not due to osmolarity changes (Ferreira et al 2021). Conversely, MSMCs from d14.5 and d18.5 mice exhibited a statistically significant depolarization after addition of 80 mM Na + (Figure 5b, d).…”

“…This was demonstrated by showing that inhibition of NALCN with gadolinium (Gd 2+ ) or replacing Na + with Lithium (Li + ), which can pass through NALCN but not activate SLO2.1, prevented Na + -activated K + influx. Furthermore, NALCN and SLO2.1 are in proximity to one another in human MSMCs, and functional coupling of NALCN and SLO2.1 regulates contractility (Ferreira et al, 2021). These data support a model that, during pregnancy, Na + influx through NALCN activates SLO2.1, leading to K + influx, Vm hyperpolarization, and MSMC quiescence.…”

“…We next wondered whether Na +mediated activation of SLO2.1 and the ensuing K + efflux could lead to membrane hyperpolarization in mouse MSMCs. Thus, we performed flow cytometry with the cationic voltage-sensitive dye, DiSC3(5) (Ferreira et al, 2021). In MSMCs from non-pregnant and d7.5 mice, treatment with 80 mM extracellular Na + led to a respective increase of 28.22 +/-26.15 and 30.14 +/-13.49 in DiSC3( 5) fluorescence (normalized to Valinomycin), indicating membrane hyperpolarization at these stages (Figure 5b, d).…”

“…Na + influx through NALCN activates SLO2.1 in MSMCs from nonpregnant and early pregnant mice. To determine whether Na + influx through NALCN activates SLO2.1 in mouse MSMCs as in human MSMCs (Ferreira et al, 2021), we conducted whole-cell patch clamp experiments on MSMCs isolated from different stages of pregnancy. Substituting 135 mM Li + with 135 mM extracellular Na + led to a 100-200% increase in K + currents at both +80 mV and -60 mV in MSMCs from non-pregnant and early pregnant (d7.5) mice but not in MSMCs from d14.5 and d18.5 (Figure 4a, c, d).…”

“…3.6 SLO2.1 and NALCN differentially modulate Ca 2+ responses in mouse MSMCs across pregnancy. Membrane depolarization in human and mouse MSMCs enhances Ca 2+ influx via VDCCs (Ferreira et al, 2021),and inhibiting SLO2.1 in human MSMCs leads to increased Ca 2+ entry through VDCCs (Ferreira et al, 2019(Ferreira et al, , 2021. Thus, we wanted to define the effects of the NALCN/SLO2.1 complex on Ca 2+ in mouse MSMCs across pregnancy.…”