8-Bromo-cAMP decreases the Ca²⁺sensitivity of airway smooth muscle contraction through a mechanism distinct from inhibition of Rho-kinase

Abstract

Endou, Katsuaki, Kunihiko Iizuka, Akihiro Yoshii, Hideo Tsukagoshi, Tamotsu Ishizuka, Kunio Dobashi, Tsugio Nakazawa, and Masatomo Mori. 8-Bromo-cAMP decreases the Ca 2ϩ sensitivity of airway smooth muscle contraction through a mechanism distinct from inhibition of Rho-kinase. Am J Physiol Lung Cell Mol Physiol 287: L641-L648, 2004. First published April 30, 2004 10.1152/ajplung.00287.2003.-To clarify whether cyclic AMP (cAMP)/cAMP-dependent protein kinase (PKA) activation and Rhokinase inhibition share a common mechanism to decrease the Ca 2ϩ sensitivity of airway smooth muscle contraction, we examined the effects of 8-bromoadenosine 3Ј,5Ј-cyclic monophosphate (8-BrcAMP), a stable cAMP analog, and (ϩ)-(R)-trans-4-(1-aminoethyl)-N-(4-pyridyl) cyclohexane carboxamide dihydrochloride, monohydrate (Y-27632), a Rho-kinase inhibitor, on carbachol (CCh)-, guanosine 5Ј-O-(3-thiotriphosphate) (GTP␥S)-, 4␤-phorbol 12,13-dibutyrate (PDBu)-, and leukotriene D4 (LTD4)-induced Ca 2ϩ sensitization in ␣-toxin-permeabilized rabbit tracheal and human bronchial smooth muscle. In rabbit trachea, CCh-induced smooth muscle contraction was inhibited by 8-BrcAMP and Y-27632 to a similar extent. However, GTP␥S-induced smooth muscle contraction was resistant to 8-BrcAMP. In the presence of a saturating concentration of Y-27632, PDBu-induced smooth muscle contraction was completely reversed by 8-BrcAMP. Conversely, PDBu-induced smooth muscle contraction was resistant to Y-27632. In the presence of a saturating concentration of 8-BrcAMP, GTP␥S-induced Ca 2ϩ sensitization was also reversed by Y-27632. The 8-BrcAMP had no effect on the ATPtriggered contraction of tracheal smooth muscle that had been treated with calyculin A in rigor solutions. The 8-BrcAMP and Y-27632 additively accelerated the relaxation rate of PDBu-and GTP␥S-treated smooth muscle under myosin light chain kinase-inhibited conditions. In human bronchus, LTD4-induced smooth muscle contraction was inhibited by both 8-BrcAMP and Y-27632. We conclude that cAMP/PKA-induced Ca 2ϩ desensitization contains at least two mechanisms: 1) inhibition of the muscarinic receptor signaling upstream from Rho activation and 2) cAMP/PKA's preferential reversal of PKC-mediated Ca 2ϩ sensitization in airway smooth muscle. is the primary regulator of smooth muscle contraction, Ca 2ϩ sensitivity of the contractile apparatus can change in response to agonists. An increase and a decrease in muscle tension at a constant Ca 2ϩ concentration are correspondingly referred to as Ca 2ϩ sensitization and desensitization of smooth muscle contraction. These are believed to be the results of changes in the ratio of kinase and phosphatase activities toward the 20-kDa light chain of myosin (MLC 20 ) (22,25). We have demonstrated that receptor-dependent, G protein-mediated Ca 2ϩ sensitization occurs in canine, rabbit, and human airway smooth muscles (28) and that a small G protein, RhoA, and its target protein, Rho-kinase, play a key role in G protein-mediated Ca 2ϩ sensitization of smooth muscle contraction (2...

Involvement of reduced sensitivity to Ca2+ in β‐adrenergic action on airway smooth muscle

“…All these cAMP‐related agents caused less reduction in [Ca 2+ ] i than SKF‐96365 when an equivalent relaxation was achieved (Figs 1–3), demonstrating that Ca 2+ ‐independent mechanisms are involved in cAMP‐dependent relaxation in ASM. This observation is consistent with previous reports [19, 20].…”

NSC23766, a Widely Used Inhibitor of Rac1 Activation, Additionally Acts as a Competitive Antagonist at Muscarinic Acetylcholine Receptors

“…Cardiac myocyte-specific overexpression of an active form of Rac predisposes the heart to postischemic contractile dysfunction (Talukder et al, 2013). Through activation of NADPH oxidase, Rac1 activation contributes to oxidative stress, which promotes cardiac injury and dysfunction (Maack et al, 2003;Endou et al, 2004;Elnakish et al, 2012;Nagase et al, 2012;Zhang et al, 2012;Ma et al, 2013). Moreover, Rac1 is essential for the hypertrophic response in the heart (Sussman et al, 2000;Satoh et al, 2006;Vettel et al, 2012).…”

Modulation of the Ca2+ sensitivity of airway smooth muscle cells in murine lung slices

“…A similar relaxation of model airways was also observed in response to the cAMPelevating compounds, FSK and IBMX. This effect of cAMP may also act via the activation of MLCP because in previous studies, cAMP-dependent PKA (or cross-activated PKG) was reported to antagonize inhibitors of MLCP, including of RhoA, pre-RhoA signaling molecules (i.e., G␣ q or G␣ 12,13 ), and CPI-17 (12,26,37). Caffeine, presumably acting as a phosphodiesterase inhibitor to elevate cAMP, also served to relax the MCh-contracted airways but this effect required relatively high concentrations of caffeine compared with the effective concentrations of FSK or IBMX.…”