GTP AND NORADRENALINE-INDUCED FORCE IN ISOLATED TOXIN-PERMEABILIZED RAT ANOCOCCYGEUS AND GUINEA-PIG PORTAL-VEIN

1. Strips of smooth muscle from rat anococcygeus and guinea-pig portal vein were treated with solutions containing crude alpha-toxin from the bacterium Staphylococcus aureus. This rendered the surface membrane permeable to small molecular weight substances, but left functional sarcolemmal adrenoceptors. Tension measurements from these preparations were used to investigate the effects of guanosine-5'-triphosphate (GTP) on the noradrenaline-induced Ca2+ release from the sarcoplasmic reticulum (SR) of the smooth muscle of rat anococcygeus and guinea-pig portal vein. 2. Under conditions of low Ca2+ buffering (0.2 mM-EGTA), applying a maximal dose of noradrenaline (30-mu-M) to a toxin-permeabilized strip of anococcygeus muscle and longitudinal muscle of guinea-pig portal vein caused a transient contracture. Subsequent exposures to noradrenaline resulted in progressively smaller contractures. However, the rate of decline in the size of the noradrenaline-induced contracture was greater in rat anococcygeus muscle than in guinea-pig portal vein preparations. The decline in the size of the contracture in toxin-permeabilized anococcygeus muscle was not due to a fall in the Ca2+ content of the SR or a reduced Ca2+ release from the SR in response to myo-inositol 1,4,5-trisphosphate (IP3). 3. The tension transients due to noradrenaline were enhanced and maintained in the presence of 100-mu-M-GTP in toxin-permeabilized guinea-pig portal vein. Addition of 100-mu-M-GTP caused a transient contracture in permeabilized rat anococcygeus muscle and only promoted the next noradrenaline response, thereafter the amplitude of the contractures decayed to zero. 4. Addition of guanosine-5'-O-(2 thiodiphosphate) (GDP-beta-S, 100-mu-M) would be expected to cause a reversible reduction of the noradrenaline response by binding to the intermediary G-protein. This was observed in toxin-permeabilized portal vein, but in rat anococcygeus muscle, GDP-beta-S caused slowing of the response to noradrenaline, thereafter the response to noradrenaline was absent. The noradrenaline response did not recover when GDP-beta-S was removed. 5. The non-metabolizable form of GTP, guanosine-5'-O-(3-thiotriphosphate) (GTP-gamma-S, 100-mu-M), caused a transient contracture in both toxin-permeabilized rat anococcygeus muscle and guinea-pig portal vein. In both these tissues, the addition of GTP-gamma-S resulted in the irreversible inhibition of the response to noradrenaline. 6. In the presence of a high concentration (10 mM) of the Ca2+ buffer EGTA, GTP (100-mu-M) and noradrenaline (30-mu-M) increased Ca2+-activated force in both tissues. In guinea-pig portal vein preparations, the response due to GTP was small in comparison to the response due to noradrenaline. In rat anococcygeus muscle, the GTP response was dominant, the noradrenaline response being small in comparison. 7. In conclusion, marked differences exist between the responses of toxin-permeabilized smooth muscle from guinea-pig portal vein and rat anococcygeus to noradrenaline and GTP. Those effects observed in 0.2 mM-EGTA on guinea-pig portal vein may be interpreted as resulting from a G-protein-mediated alpha-1-adrenergic response. However, in rat anococcygeus preparations the alpha-1-adrenergic response was altered by additional factors that eventually abolished responses to noradrenaline.