Calcium signaling in airway smooth muscle

Abstract

Contractility of airway smooth muscle requires elevation of intracellular calcium concentration. Under resting conditions, airway smooth muscle cells maintain a relatively low intracellular calcium concentration, and activation of the surface receptors by contractile agonists results in an elevation of intracellular calcium, culminating in contraction of the cell. The pattern of elevation of intracellular calcium brought about by agonists is a dynamic process and involves the coordinated activities of ion channels located in the plasma membrane and the sarcoplasmic reticulum. Among the signaling molecules involved in this dynamic calcium regulation in airway smooth muscle cells are inositol 1,4,5-trisphosphate and cyclic ADP-ribose, which mobilize calcium from the sarcoplasmic reticulum by acting via the inositol 1,4,5-trisphosphate and ryanodine receptors, respectively. In addition, calcium influx from the extracellular space is critical for the repletion of the intracellular calcium stores during activation of the cells by agonists. Calcium influx can occur via voltage- and receptor-gated channels in the plasma membrane, as well as by influx that is triggered by depletion of the intracellular stores (i.e., store-operated calcium entry mechanism). Transient receptor potential proteins appear to mediate the calcium influx via receptor- and store-operated channels. Recent studies have shown that proinflammatory cytokines regulate the expression and activity of the pathways involved in intracellular calcium regulation, thereby contributing to airway smooth muscle cell hyperresponsiveness. In this review, we will discuss the specific roles of cyclic ADP-ribose/ryanodine receptor channels and transient receptor potential channels in the regulation of intracellular calcium in airway smooth muscle cells.

Figure 1.

Mechanisms of cyclic-ADP-ribose (cADPR)-mediated Ca²⁺ mobilization through ryanodine receptor. cADPR interacts directly or indirectly with the ryanodine receptor (RyR) to increase the open probability of the ion channel. FK506-binding protein 12.6 (FKBP12.6) is physically associated with the subunits of RyR and keeps it in a closed state. Interaction of cADPR with the RyR causes dissociation of FKBP12.6 and an increase in the open probability of RyR to release sarcoplasmic reticular (SR) Ca²⁺. Calmodulin (CaM), a Ca²⁺-binding protein, is known to play a critical role in cADPR-mediated Ca²⁺ mobilization in sea urchin egg microsomes. In pancreatic β-cells, the CaM is known to recruit CaM kinase II to phosphorylate the RyR. The phosphorylation increases the open probability of the RyR. cADPR can also act directly on RyR to open the channel.

Figure 2.

An integrative model of differential recruitment of cellular calcium mobilization mechanisms. Acetylcholine acts on M2 and M3 subtypes of muscarinic cholinergic receptors on the ASM cells to elicit a calcium response. M3 subtype of receptor, which is coupled to G_αq protein, recruits phospholipase C-β (PLC-β) and generates inositol 1,4,5-tris-phosphate (IP₃). IP₃ acts on IP₃-receptor (IP₃R) to increase its open probability to release Ca²⁺ from SR. Activation of the M2 subtype of receptor, which is coupled to G_αi protein, recruits CD38/cADPR signaling pathway. The β-NAD is transported out from the cytoplasm of the myocyte by connexin 43 hemichannel proteins (Cx43). cADPR generated by CD38 is transported back into the myocyte cytoplasm either through specialized nucleoside transporters (NT) or via the oligomerized CD38. ADPR, the predominant product of the CD38 enzymatic activity, acts on transient receptor potential (TRP) channels to elicit influx of divalent cations including Ca^2+. Upon interaction of cADPR with RyR (direct or indirect), dissociation of FK506-binding protein 12.6 (FKBP12.6) from RyR results in open probability of the ion channel and releases SR Ca²⁺. SR Ca²⁺-ATPase 2 (SERCA2) pumps the cytosolic Ca²⁺ back into SR to replenish the SR Ca²⁺ stores. The TRP channels also play major role in replenishing the SR Ca²⁺ store through store-operated Ca²⁺ entry (SOCE). EC = extra cellular space; PM = plasma membrane.

Figure 3.

Treatment of cells with 8Br-cADPR results in attenuated [Ca²⁺]_i responses to agonists in control and cytokine-treated human airway smooth muscle (HASM) cells. Fura-2/AM-loaded control and cytokine-treated HASM cells were incubated with 8Br-cADPR and [Ca²⁺]_i responses to agonists were measured. The net [Ca²⁺]_i values in the presence of 8Br-cADPR in control and cytokine-treated cells were compared with those of the corresponding controls (in the absence of 8Br-cADPR). Net [Ca²⁺]_i responses to acetylcholine, bradykinin, and thrombin in control and cytokine-treated HASM cells, in the presence or absence of 8Br-cADPR, are shown. Panels on the right represent % inhibition of the [Ca²⁺]_i responses by 8Br-cADPR. Note significant (P ≤ 0.05) attenuation of [Ca²⁺]_i responses in the presence of 8Br-cADPR in control and cytokine-treated HASM cells. Data are from three different cell preparations. IL-1β = interleukin-1β; TNF-α = tumour necrosis factor-α; IFN-γ = interferon-γ. Reprinted by permission from Reference .