ACTIONS OF 2ND MESSENGERS SYNTHESIZED BY VARIOUS SPASMOGENIC AGENTS AND THEIR RELATION TO MECHANICAL RESPONSES IN DOG TRACHEAL SMOOTH-MUSCLE

1. The effects of the spasmogenic agents, carbachol (CCh), histamine, 5-hydroxytryptamine (5-HT) and 9,11-epithio-11,12-methano-thromboxane A2 (STA2) were investigated on smooth muscle tissues of the dog trachea. 2. CCh (10 μM) produced a larger contraction than high K (128 mM), 10 μM histamine, 5-HT or STA2. Histamine and 5-HT produced the same amplitude of contraction as each other. In Ca-free solution containing 0.2 mM EGTA, only a phasic contraction was evoked by the above agents (except for K which induced no contraction at all). 3. In skinned muscle tissues, the maximum amplitude of contraction that could be induced by Ca (10 μM) was slightly larger than the maximum CCh-induced contraction (also at 10 μM) evoked in intact muscle tissues. Caffeine and inositol 1,4,5-trisphosphate (IP3) both produced contraction. 4. CCh, histamine and 5-HT (10 μM) produced a sustained contraction for over 30 min and also increased phosphorylation of the 20 kD protein of myosin light chain (MLC20) for over 30 min with no attenuation. Greater concentrations of the above agents caused more phosphorylation of MLC20. 5. CCh (above 1 nM), histamine (above 10 nM) and 5-HT (above 100 nM) increased the amount of IP3, in a concentration-dependent manner. Synthesis of IP3 induced by the above agents reached its peak value within 30s and lasted for about 3 min. The potencies for the synthesis of IP3 were in the following order: CCh > histamine > 5-HT > STA2. 6. Isoprenaline (10 μM) markedly enhanced but CCh (10 μM) slightly reduced the amount of cyclic AMP. 5-HT (10 μM) and STA2 (10 μM) reduced, but histamine (10 μM) and CCh (10 μM) increased the amount of cyclic GMP. 7. Using fura 2, cytosolic Ca was measured by monitoring the ratio of the fluorescent signal excited at 340 and 380 nm wavelengths in the presence of extracellular Ca. CCh (10 μM) increased the Ca transient from 182 nM to 1.42 μM. When the CCh-induced peak Ca transient (10 μM) was normalised, 10 μM histamine, 5-HT and STA2 showed smaller values such as 0.49, 0.53 and 0.04 times the control, respectively, and these values corresponded well with the amplitudes of contraction evoked by each of the stimulants. 8. The results can be summarized as follows: stronger spasmogenic responses occur on application of CCh than on application of 5-HT or histamine, and STA2 may have a minor role as a spasmogenic agent. The maximum amplitudes (peaks) of contraction evoked by the above spasmogenic agents are closely related to the maximum increase in cytosolic Ca, but sustained contraction and increased phosphorylation of myosin cannot be explained by the increased amount of Ca. In the case of 5-HT and histamine, synthesized cyclic nucleotides may interact with the action of IP3 for the regulation of contraction in a positive or negative manner, respectively.