Membrane depolarization causes a direct activation of G protein-coupled receptors leading to local Ca ²⁺ release in smooth muscle

Abstract

Membrane depolarization activates voltage-dependent Ca 2+ channels (VDCCs) inducing Ca 2+ release via ryanodine receptors (RyRs), which is obligatory for skeletal and cardiac muscle contraction and other physiological responses. However, depolarization-induced Ca 2+ release and its functional importance as well as underlying signaling mechanisms in smooth muscle cells (SMCs) are largely unknown. Here we report that membrane depolarization can induce RyR-mediated local Ca 2+ release, leading to a significant increase in the activity of Ca 2+ sparks and contraction in airway SMCs. The increased Ca 2+ sparks are independent of VDCCs and the associated extracellular Ca 2+ influx. This format of local Ca 2+ release results from a direct activation of G protein-coupled, M 3 muscarinic receptors in the absence of exogenous agonists, which causes activation of Gq proteins and phospholipase C, and generation of inositol 1,4,5-triphosphate (IP 3 ), inducing initial Ca 2+ release through IP 3 receptors and then further Ca 2+ release via RyR2 due to a local Ca 2+ -induced Ca 2+ release process. These findings demonstrate an important mechanism for Ca 2+ signaling and attendant physiological function in SMCs.

Calcineurin upregulates local Ca2+signaling through ryanodine receptor-1 in airway smooth muscle cells

“…However, the frequency of Ca 2ϩ sparks in RyR3 Ϫ/Ϫ cerebral artery myocytes is not significantly increased. Furthermore, we have shown that resting and depolarization-induced Ca 2ϩ sparks are not prevented in RyR3 Ϫ/Ϫ ASMCs (25) and that the downregulation of local Ca 2ϩ signaling by PKC⑀ is not blocked in RyR3 Ϫ/Ϫ cells either (24). These findings, taken together, clearly point out that CaN regulates local Ca 2ϩ signaling specifically through RyR1, but not RyR2 or RyR3.…”

“…1E), unlike in cardiac myocytes where PP2A was shown to increase Ca 2ϩ sparks (40). RyRs are essential for spark formation; however, IP 3 Rs cross talk with RyRs, promoting Ca 2ϩ spark formation through a local CICR process (25,26). In the present study, we have revealed that CaN is able to modulate Ca 2ϩ sparks in the absence of functional IP 3 Rs (Fig.…”

“…Because we have just outlined the role for the serine/threonine protein phosphatase CaN in regulating local Ca 2ϩ signaling, we questioned if it works through RyR1. First, RyR1 heterozygous gene deletion (RyR1 Ϫ/ϩ ) mice were used because RyR1 homozygous knockout mice (RyR1 Ϫ/Ϫ ) die before or at birth; second, local Ca 2ϩ signaling is significantly different in embryonic vs. adult ASMCs; and third, the effects of PKC⑀ are fully blocked in RyR1 Ϫ/ϩ ASMCs (24,25). Here, we found that treatment with CAIP was without effect on local Ca 2ϩ signals in RyR1 (Fig.…”

“…Nevertheless, previous reports have shown that ASMC contraction is not significantly affected by LTCC blockers in isolated animal ASMCs and tissues, and clinical studies indicate that LTCC blockers are ineffective in the treatment of airway smooth muscle contraction and asthma (9, 14, 38). Our recent research demonstrates that, under physiological conditions, membrane depolarization causes a direct activation of G q protein-coupled M 3 muscarinic receptors, leading to intracellular Ca 2ϩ release and contraction without the involvement of LTCCs in ASMCs (25), signifying the especially important role of intracellular Ca 2ϩ release for Recent work from our laboratory has shown that PKC⑀ downregulates local Ca 2ϩ signaling through its specific effect on RyR1 as well as inhibiting the contractile response of ASMCs to MCh (23). Because a kinase regulates the activity of its substrate in coordination with one or more phosphatases by causing phosphorylation and dephosphorylation, we wondered whether and which phosphatase is involved in the regulation of local Ca 2ϩ signals in ASMCs.…”

“…We have reported that a direct activation of native M 3 muscarinic receptors following exposure of the receptor agonist methacholine (MCh) or membrane depolarization results in activation of a G q protein-coupled receptor-mediated signaling cascade, initiating the phospholipase C (PLC)-dependent breakdown of phosphatidylinositol 4,5-bisphosphate into diacylgycerol (DAG) and inositol 1,4,5-trisphosphate (IP 3 ); IP 3 activates IP 3 receptors (IP 3 Rs), inducing intracellular Ca 2ϩ release; this activates nearby RyR2 via a local Ca 2ϩ -induced Ca 2ϩ -release (CICR) mechanism, amplifying agonist-evoked, IP 3 R-mediated Ca 2ϩ release and contraction in ASMCs (25). As a result of the MCh exposure or membrane depolarization, PLC also produces DAG, which activates protein kinase C (PKC).…”