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Spontaneous calcium activity in metanephric mesenchymal cells regulates branching morphogenesis in the embryonic kidney.

ABSTRACT: The central role of calcium signaling during development of early vertebrates is well documented, but little is known about its role in mammalian embryogenesis. We have used immunofluorescence and time-lapse calcium imaging of cultured explanted embryonic rat kidneys to study the role of calcium signaling for branching morphogenesis. In mesenchymal cells, we recorded spontaneous calcium activity that was characterized by irregular calcium transients. The calcium signals were dependent on release of calcium from intracellular stores in the endoplasmic reticulum. Down-regulation of the calcium activity, both by blocking the sarco-endoplasmic reticulum Ca²⁺-ATPase and by chelating cytosolic calcium, resulted in retardation of branching morphogenesis and a reduced formation of primitive nephrons but had no effect on cell proliferation. We propose that spontaneous calcium activity contributes with a stochastic factor to the self-organizing process that controls branching morphogenesis, a major determinant of the ultimate number of nephrons in the kidney.-Fontana, J. M., Khodus, G. R., Unnersjö-Jess, D., Blom, H., Aperia, A., Brismar, H. Spontaneous calcium activity in metanephric mesenchymal cells regulates branching morphogenesis in the embryonic kidney.

SUBMITTER: Fontana JM

PROVIDER: S-EPMC6404591 | biostudies-literature | 2019 Mar

REPOSITORIES: biostudies-literature

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Spontaneous calcium activity in metanephric mesenchymal cells regulates branching morphogenesis in the embryonic kidney.

Fontana Jacopo M JM Khodus Georgiy R GR Unnersjö-Jess David D Blom Hans H Aperia Anita A Brismar Hjalmar H

FASEB journal : official publication of the Federation of American Societies for Experimental Biology 20181129 3

The central role of calcium signaling during development of early vertebrates is well documented, but little is known about its role in mammalian embryogenesis. We have used immunofluorescence and time-lapse calcium imaging of cultured explanted embryonic rat kidneys to study the role of calcium signaling for branching morphogenesis. In mesenchymal cells, we recorded spontaneous calcium activity that was characterized by irregular calcium transients. The calcium signals were dependent on release ...[more]

PMID: 30496703

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Project description:Airway branching morphogenesis in utero is essential for optimal postnatal lung function. In the fetus, branching morphogenesis occurs during the pseudoglandular stage (weeks 9-17 of human gestation, embryonic days (E)11.5-16.5 in mouse) in a hypercalcaemic environment (~1.7 in the fetus vs. ~1.1-1.3 mM for an adult). Previously we have shown that fetal hypercalcemia exerts an inhibitory brake on branching morphogenesis via the calcium-sensing receptor. In addition, earlier studies have shown that nifedipine, a selective blocker of L-type voltage-gated Ca(2+) channels (VGCC), inhibits fetal lung growth, suggesting a role for VGCC in lung development. The aim of this work was to investigate the expression of VGCC in the pseudoglandular human and mouse lung, and their role in branching morphogenesis. Expression of L-type (CaV1.2 and CaV1.3), P/Q type (CaV2.1), N-type (CaV2.2), R-type (CaV2.3), and T-type (CaV3.2 and CaV3.3) VGCC was investigated in paraffin sections from week 9 human fetal lungs and E12.5 mouse embryos. Here we show, for the first time, that Cav1.2 and Cav1.3 are expressed in both the smooth muscle and epithelium of the developing human and mouse lung. Additionally, Cav2.3 was expressed in the lung epithelium of both species. Incubating E12.5 mouse lung rudiments in the presence of nifedipine doubled the amount of branching, an effect which was partly mimicked by the Cav2.3 inhibitor, SNX-482. Direct measurements of changes in epithelial cell membrane potential, using the voltage-sensitive fluorescent dye DiSBAC2(3), demonstrated that cyclic depolarisations occur within the developing epithelium and coincide with rhythmic occlusions of the lumen, driven by the naturally occurring airway peristalsis. We conclude that VGCC are expressed and functional in the fetal human and mouse lung, where they play a role in branching morphogenesis. Furthermore, rhythmic epithelial depolarisations evoked by airway peristalsis would allow for branching to match growth and distension within the developing lung.

Project description:Prostatic branching morphogenesis is an intricate event requiring precise temporal and spatial integration of numerous hormonal and growth factor-regulated inputs, yet relatively little is known about the downstream signaling pathways that orchestrate this process. In this study, we use a novel mesenchyme-free embryonic prostate culture system, newly available mTOR inhibitors and a conditional PTEN loss-of-function model to investigate the role of the interconnected PI3K and mTOR signaling pathways in prostatic organogenesis. We demonstrate that PI3K levels and PI3K/mTOR activity are robustly induced by androgen during murine prostatic development and that PI3K/mTOR signaling is necessary for prostatic epithelial bud invasion of surrounding mesenchyme. To elucidate the cellular mechanism by which PI3K/mTOR signaling regulates prostatic branching, we show that PI3K/mTOR inhibition does not significantly alter epithelial proliferation or apoptosis, but rather decreases the efficiency and speed with which the developing prostatic epithelial cells migrate. Using mTOR kinase inhibitors to tease out the independent effects of mTOR signaling downstream of PI3K, we find that simultaneous inhibition of mTORC1 and mTORC2 activity attenuates prostatic branching and is sufficient to phenocopy combined PI3K/mTOR inhibition. Surprisingly, however, mTORC1 inhibition alone has the reverse effect, increasing the number and length of prostatic branches. Finally, simultaneous activation of PI3K and downstream mTORC1/C2 via epithelial PTEN loss-of-function also results in decreased budding reversible by mTORC1 inhibition, suggesting that the effect of mTORC1 on branching is not primarily mediated by negative feedback on PI3K/mTORC2 signaling. Taken together, our data point to an important role for PI3K/mTOR signaling in prostatic epithelial invasion and migration and implicates the balance of PI3K and downstream mTORC1/C2 activity as a critical regulator of prostatic epithelial morphogenesis.

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Spontaneous calcium activity in metanephric mesenchymal cells regulates branching morphogenesis in the embryonic kidney.

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Spontaneous calcium activity in metanephric mesenchymal cells regulates branching morphogenesis in the embryonic kidney.

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