Abstract
Voltage-gated calcium channels and estrogen receptors are essential players in uterine physiology, and their association with different calcium signaling pathways contributes to healthy and pathological conditions of the uterine myometrium. Among the properties of the various cell subtypes present in human uterine myometrium, there is increasing evidence that calcium oscillations in telocytes (TCs) contribute to contractile activity and pregnancy. Our study aimed to evaluate the effects of beta-estradiol on voltage-gated calcium channels and estrogen receptors in TCs from human uterine myometrium and to understand their role in pregnancy. For this purpose, we employed patch-clamp recordings, ratiometric Fura-2-based calcium imaging analysis, and qRT-PCR techniques for the analysis of cultured human myometrial TCs derived from pregnant and non-pregnant uterine samples. In human myometrial TCs from both non-pregnant and pregnant uterus, we evidenced by qRT-PCR the presence of genes encoding for voltage-gated calcium channels (Cav3.1, Ca3.2, Cav3.3, Cav2.1), estrogen receptors (ESR1, ESR2, GPR30), and nuclear receptor coactivator 3 (NCOA3). Pregnancy significantly upregulated Cav3.1 and downregulated Cav3.2, Cav3.3, ESR1, ESR2, and NCOA3, compared to the non-pregnant condition. Beta-estradiol treatment (24 h, 10, 100, 1000 nM) downregulated Cav3.2, Cav3.3, Cav1.2, ESR1, ESR2, GRP30, and NCOA3 in TCs from human pregnant uterine myometrium. We also confirmed the functional expression of voltage-gated calcium channels by patch-clamp recordings and calcium imaging analysis of TCs from pregnant human myometrium by perfusing with BAY K8644, which induced calcium influx through these channels. Additionally, we demonstrated that beta-estradiol (1000 nM) antagonized the effect of BAY K8644 (2.5 or 5 µM) in the same preparations. In conclusion, we evidenced the presence of voltage-gated calcium channels and estrogen receptors in TCs from non-pregnant and pregnant human uterine myometrium and their gene expression regulation by beta-estradiol in pregnant conditions. Further exploration of the calcium signaling in TCs and its modulation by estrogen hormones will contribute to the understanding of labor and pregnancy mechanisms and to the development of effective strategies to reduce the risk of premature birth.
Highlights
Uterine contractions represent a key point throughout pregnancy
We demonstrated the immunopositivity for T-type calcium channels (i.e., Cav3.1 and Cav3.2) in human myometrial TCs from pregnant and non-pregnant uterus [35]
We evidenced the presence of T-type calcium currents and high-voltage-activated Ca2+ channels (HVA) currents in human myometrial TCs by employing patch-clamp recordings [35]
Summary
Uterine contractions represent a key point throughout pregnancy. Considered to be normal during pregnancy, uterine contractions may be responsible for triggering premature or dysfunctional labor. The myometrium suffers morphological and adaptive changes which enable it to become a forceful organ necessary for delivery. Myometrium contractility involves, among other physiological mechanisms that produce excitation in the uterus, changes in Ca2+ signals. Smooth myofiber contraction requires actin and myosin myofilaments and their interaction. The cross-bridge formation and contraction are mediated by elevated levels of intracellular Ca2+ and myosin light-chain phosphorylation
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