Ontogeny of Purinergic Receptor-Regulated Ca²⁺ Signaling in Mouse Cortical Collecting Duct Epithelium

Abstract

Changes in ATP-induced increase in [Ca²⁺]_i during collecting duct ontogeny were studied in primary monolayer cultures of mouse ureteric bud (UB) and cortical collecting duct (CCD) cells by Fura-PE3 fluorescence ratio imaging. In UB (embryonic day E14 and postnatal day P1) the ATP-stimulated increase (EC₅₀ ≈ 1 µM) in fluorescence ratio (ΔR_ATP) was independent of extracellular Ca²⁺ and insensitive to the P2 purinoceptor-antagonist suramin (1 mM). From day P7 onward when CCD morphogenesis had been completed ΔR_ATP increased and became dependent on extracellular Ca²⁺. This ATP-stimulated Ca²⁺ entry into CCD cells was non-capacitative and suramin (1 mM)-insensitive, but sensitive to nifedipine (30 µM) and enhanced by Bay K8644 (15 µM), a blocker and an agonist of L-type Ca²⁺ channels, respectively. Quantitative RT-PCR demonstrated similar mRNA expression of L-type Ca²⁺ channel α1-subunit, P2Y₁, P2Y₂, and P2X_4b purinoceptors in UB and CCD monolayers while the abundance of P2X₄ mRNA increased with CCD morphogenesis. In conclusion, both embryonic and postnatal cells express probably P2Y₂-stimulated Ca²⁺ release from intracellular stores. With development, the CCD epithelium acquires ATP-stimulated Ca²⁺ entry via L-type Ca²⁺ channels. This pathway might by mediated by the increasing expression of P2X₄-receptors resulting in an increasing ATP-dependent membrane depolarization and activation of L-type Ca²⁺ channels.

Sodium-Dependent Regulation of Renal Amiloride-Sensitive Currents by Apical P2 Receptors

“…6,[27][28][29][30][31][32][33][34][35][36][37][38][39] They also extend the findings of an earlier study in which, using Na ϩ -replete rats and a limited number of commercially available P2R antibodies, we demonstrated a variety of P2R expressed throughout the nephron. 40 …”

Mechanoregulation of intracellular Ca2+concentration is attenuated in collecting duct of monocilium-impairedorpkmice

“…Furthermore, baseline [Ca 2ϩ ] i in 1-and 2-wk old normal mouse collecting duct cells was similar, a finding that contrasts with the developmental increase in resting [Ca 2ϩ ] i reported in rabbit collecting duct cells after the first week of postnatal life (43). In an analysis of purinergic-induced Ca 2ϩ signaling in mouse ureteric bud (E14 -P1) and neonatal CCD (P7 to P20) principal cells, Tschop et al (38) observed similar resting [Ca 2ϩ ] i during all developmental stages. Relevant to the present study was the observation by the latter investigators that, when cells were bathed in a Ca 2ϩ -free solution, ATP induced similar increases in [Ca 2ϩ ] i at all developmental stages (38), suggesting that the magnitude of internal Ca 2ϩ stores did not change with development.…”

“…In an analysis of purinergic-induced Ca 2ϩ signaling in mouse ureteric bud (E14 -P1) and neonatal CCD (P7 to P20) principal cells, Tschop et al (38) observed similar resting [Ca 2ϩ ] i during all developmental stages. Relevant to the present study was the observation by the latter investigators that, when cells were bathed in a Ca 2ϩ -free solution, ATP induced similar increases in [Ca 2ϩ ] i at all developmental stages (38), suggesting that the magnitude of internal Ca 2ϩ stores did not change with development. In contrast, ATP-induced external Ca 2ϩ entry, mediated by L-type Ca 2ϩ channels, was acquired only after P7, when collecting duct morphogenesis was complete (38).…”

Ontogeny of flow-stimulated potassium secretion in rabbit cortical collecting duct: functional and molecular aspects

“…The modest response of the neonatal principal cell to flow cannot be explained by the absence of an apical central cilium; indeed, scanning electron microscopic analyses reveal that the principal cell cilia at birth are longer than those in the adult (10). Functional expression of Ca 2ϩ -conducting channels in the collecting duct may be low in early renal development (53). Studies in developing neurons suggest that IP 3 receptors and Ca 2ϩ channels appear to be present early in life but are not assembled to allow IP 3 -assisted, Ca 2ϩ -induced Ca 2ϩ release (62).…”