High pH-Sensitive Store-Operated Ca2+ Entry Mediated by Ca2+ Release-Activated Ca2+ Channels in Rat Odontoblasts

Abstract

Odontoblasts play a crucial role in dentin formation and sensory transduction following the application of stimuli to the dentin surface. Various exogenous and endogenous stimuli elicit an increase in the intracellular free calcium concentration ([Ca2+]i) in odontoblasts, which is mediated by Ca2+ release from intracellular Ca2+ stores and/or Ca2+ influx from the extracellular medium. In a previous study, we demonstrated that the depletion of Ca2+ stores in odontoblasts activated store-operated Ca2+ entry (SOCE), a Ca2+ influx pathway. However, the precise biophysical and pharmacological properties of SOCE in odontoblasts have remained unclear. In the present study, we examined the functional expression and pharmacological properties of Ca2+ release-activated Ca2+ (CRAC) channels that mediate SOCE and evaluated the alkali sensitivity of SOCE in rat odontoblasts. In the absence of extracellular Ca2+, treatment with thapsigargin (TG), a sarco/endoplasmic reticulum Ca2+-ATPase inhibitor, induced an increase in [Ca2+]i. After [Ca2+]i returned to near-resting levels, the subsequent application of 2.5 mM extracellular Ca2+ resulted in an increase in [Ca2+]i which is a typical of SOCE activation. Additionally, application of 2-methylthioadenosine diphosphate trisodium salt (2-MeSADP), a P2Y1,12,13 receptor agonist, or carbachol (CCh), a muscarinic cholinergic receptor agonist, in the absence of extracellular Ca2+, induced a transient increase in [Ca2+]i. The subsequent addition of extracellular Ca2+ resulted in significantly higher [Ca2+]i in 2-MeSADP- or CCh-treated odontoblasts than in untreated cells. SOCE, that is activated by addition of extracellular Ca2+ in the TG pretreated odontoblasts was then suppressed by Synta66, BTP2, or lanthanum, which are CRAC channel inhibitors. Treatment with an alkaline solution enhanced SOCE, while treatment with HC030031, a TRPA1 channel antagonist, inhibited it. The amplitude of SOCE at pH 9 in the presence of HC030031 was higher than that at pH 7.4 in the absence of HC030031. These findings indicate that CRAC channel-mediated alkali-sensitive SOCE occurs in odontoblasts. SOCE is mediated by P2Y and muscarinic-cholinergic receptors, which are activated by endogenous ligands in odontoblasts.

Large-Conductance Calcium-Activated Potassium Channels and Voltage-Dependent Sodium Channels in Human Cementoblasts

“…Cementoblasts express cation (such as Ca 2+ ) permeable transient receptor potential ankyrin subfamily member 1 (TRPA1) channels, which are known to be sensitive to alkaline extracellular conditions (Tsumura et al, 2013;Kimura et al, 2016) to mediate proliferation and cementum mineralization (Muramatsu et al, 2019). Although confirmation is needed, cementoblast K Ca channel activation may play a particularly important role in cell proliferation and differentiation, and cementum formation, via crosstalk with intracellular Ca 2+ signaling pathways mediated by high pH-sensitive TRP channel subfamily members (Kimura et al, 2018). TRPA1 channels also mediate mechanosensitive Ca 2+ signaling (Sato et al, 2013(Sato et al, , 2018Tsumura et al, 2013;Shibukawa et al, 2015;Kimura et al, 2016).…”

Plasma Membrane Ca2+–ATPase in Rat and Human Odontoblasts Mediates Dentin Mineralization

“…To induce membrane stretching [3,29], a hypotonic solution (200 mOsm/L) was prepared by decreasing the concentration of mannitol to 64 mM. For the high-pH (pH 8) extracellular solution, 12 mM NaHCO 3 in Krebs solution was changed to 8 mM (pH 8) NaOH [6,7]; this change had no effect on the extracellular free Ca 2+ concentration in the test solution. Caloxin 1b1 was obtained from Karebay Biochem, Inc. (Monmouth Junction, NJ, USA).…”

“…Odontoblasts play critical roles in the generation of dentinal sensitivity (known as the odontoblast hydrodynamic receptor model [1]) and dentin formation (dentinogenesis) in both physiological and pathological settings. We previously reported that odontoblasts are sensory receptor cells capable of detecting multiple stimuli applied to the dentin surface [1][2][3][4][5][6][7][8]. These stimuli at the surface are transformed into dentinal fluid movements, which elicit intracellular Ca 2+ signaling by increasing the concentration of intracellular free Ca 2+ ([Ca 2+ ] i ) through Ca 2+ influx activated by mechanosensitive ion channels, transient receptor potential (TRP) channel subtypes, and Piezo1 channels [1,5].…”

Distinct and Overlapping Expression Patterns of the Homer Family of Scaffolding Proteins and Their Encoding Genes in Developing Murine Cephalic Tissues

“…In odontoblasts Ca 2+ signaling and Ca 2+ transport mechanism are involved in sensory transduction upon exposure of dentin to diverse stimuli and in dentin mineralization [81][82][83][84][85][86][87][88][89][90]. It has been shown that odontoblasts express Orai1 [45,91] as well as Stim1 and Stim2 [49] and that SOCE is activated upon intracellular Ca 2+ store depletion in odontoblasts [85,90]. Ca 2+ influx into odontoblasts is also mediated by voltage-gated L-type channels [81,83,92] and odontoblasts express L-type Ca v 1.2 channels [93].…”