Receptor-dependent G protein-mediated Ca 2+ sensitization in canine airway smooth muscle

Abstract

To determine the mechanisms of receptor-dependent Ca 2+ sensitization in airway smooth muscle, canine tracheal smooth muscle (CTSM) was permeabilized with α-toxin or β-escin. Although the effects of 5-hydroxytryptamine (100 [LM), histamine (100 μM), and the thromboxane A 2 analogue U-46619 (100 μM) were negligible, carbachol (100 μM) and endothelin-1 (ET-1, 1 μM) evoked additional contractions of 47.0 ± 5.90% and 25.0 ± 5.37% ( n = 6) at pCa 6.7 with GTP (3 μM) (normalized to the maximum contraction at pCa 4.5) in α-toxinpermeabilized CTSM. GDP-β-S (1 μM) reversed the carbachol and ET-1 responses completely. GTP-γ-S (30 μM) and 4β-phorbol 12,13-dibutyrate (PDBu, 3 μM) increased the Ca 2+ sensitivity (median effective pCa) of contraction by 1.8- and 4.4-fold, respectively ( n = 4–11, P < 0.05). The effects of saturating concentrations of GTP-γ-S and PDBu were additive. A synthetic peptide (T 2) corresponding to the actin-binding site of calponin caused a dose-dependent contraction of β-escin permeabilized CTSM, with the peak effect (25 ± 4%, n = 4) at 1200 μM. PDBu (3 μM) caused contraction of the T 2 peptide-treated CTSM. In conclusion, Ca 2+ sensitization of CTSM depends on receptor type and is mediated by G proteins and protein kinase C whose effects are additive, with a partial contribution by calponin.