Calcium compartments in vascular smooth muscle cells as detected by aequorin signal.
1. To examine whether cytosolic Ca2+ in smooth muscle cells distributes evenly, cytosolic Ca2+ levels were measured with two different Ca2+ indicators in the ferret isolated portal vein; a fluorescent indicator, fura-PE3, that shows the average Ca2+ level, and a photoprotein, aequorin, that preferentially shows a high Ca2+ compartment. 2. A noradrenaline (10 microM)-induced sustained contraction was associated with a sustained increase in the fura-PE3 signal, or a transient increase followed by small sustained increase in the aequorin signal. A high K(+)-induced contraction was associated with a sustained increase in both the fura-PE3 and aequorin signals. 3. A second application of noradrenaline or high K+ induced reproducible contractions and fura-PE3 signals. In contrast, the aequorin signal resulting from a second application of noradrenaline or high K+ was much smaller than the first signal. 4. Following a 13 h but not a 3 h resting period, the aequorin signal stimulated by noradrenaline or high K+ recovered, without any change in the contractile response. 5. In Ca(2+)-free solution, high K+ was ineffective, whereas noradrenaline induced only a small aequorin signal and contraction compared to those obtained in the presence of external Ca2+. After the addition of Ca2+, the first application of noradrenaline induced a large aequorin signal and a large contraction, although a second application induced a much smaller aequorin signal accompanied by a large contraction. 6. These results suggest that high K+ and noradrenaline increase Ca2+ in at least two cytosolic compartments; a compartment that is coupled to the contractile mechanism ('contractile' Ca2+ compartment; major portion of cytoplasm containing contractile elements) and a compartment that is not coupled to contractile mechanisms ('non-contractile' Ca2+ compartment; small sub-membrane area that does not contain contractile elements). On stimulation, the Ca2+ level in the 'contractile' compartment may increase to a level high enough to stimulate myosin light chain kinase but not so high as to consume aequorin rapidly. In contrast, the Ca2+ level in the 'non-contractile' compartment may increase so greatly that aequorin in this compartment is rapidly consumed. These two compartments may be separated by a diffusion barrier and, during a resting period, aequorin may slowly diffuse from the 'contractile' compartment to the 'non-contractile' compartment and thus restore the full aequorin signal. An increase in Ca2+ in the 'non-contractile' compartment seems to be dependent mainly on Ca2+ influx and partly on Ca2+ release.
Abe
F
F
Department of Pharmacology, Yamagata University School of Medicine, Japan.
Mitsui
M
M
Karaki
H
H
Endoh
M
M
D000331:Aequorin; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002451:Cell Compartmentation; D005260:Female; D005289:Ferrets; D005456:Fluorescent Dyes; D016257:Fura-2 / Q000031:analogs & derivatives; D066298:In Vitro Techniques; D008297:Male; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000378:metabolism*; D009638:Norepinephrine / Q000494:pharmacology; D011169:Portal Vein / Q000166:cytology / Q000378:metabolism; D011188:Potassium / Q000494:pharmacology; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*; D014662:Vasoconstrictor Agents / Q000494:pharmacology
D005456:Fluorescent Dyes; D014662:Vasoconstrictor Agents; C098106:fura-PE3; D000331:Aequorin; D011188:Potassium; D002118:Calcium; D016257:Fura-2; D009638:Norepinephrine