Differential effects of diltiazem and nitroglycerin on cytosolic Ca2+ concentration and on force in the bovine ophthalmic artery.
OBJECTIVE
To determine the mechanisms of inhibition by diltiazem (Dil) and nitroglycerin (NG) of the contraction induced by serotonin (5-HT) in the ophthalmic artery.


METHODS
Using front-surface fluorometry of fura-2 and the medial strips of the bovine ophthalmic artery, [Ca2+]i and force were monitored simultaneously. Changes in the force at a constant [Ca2+]i were determined by use of receptor-coupled membrane permeabilization with alpha-toxin.


RESULTS
In the presence of extracellular Ca2+, 5-HT (10(-5) M) induced an initial transient and subsequently lower steady state elevation of [Ca2+]i. The transient elevation of [Ca2+]i was dependent on both intracellular and extracellular [Ca2+]i, whereas the steady state elevation was dependent on only extracellular Ca2+. For a given level of elevation of [Ca2+]i, 5-HT produced a greater force than the depolarization with high external K+ (118 mM) solution. In the permeabilized ophthalmic artery smooth muscle, 5-HT enhanced the contractile response to constant cytosolic Ca2+ (pCa 6.5) in the presence of guanosine triphosphate (GTP, 10 microM), but not in its absence. Therefore, 5-HT induces [Ca2+]i elevation, depending on both extracellular (Ca2+ influx) and intracellular Ca2+ (Ca2+ release), and it potentiates the Ca2+ sensitivity of the contractile apparatus through the activation of G-proteins. 5-HT-induced release of Ca2+ from the store was inhibited by NG, but not by Dil, in a concentration-dependent manner. However, neither NG nor Dil inhibited caffeine (20 mM)-induced release of Ca2+ from the store. Dil (10 microM) and NG (10 microM) inhibited in a concentration-dependent manner the steady state elevations of [Ca2+]i (Ca2+ influx) and force induced by 5-HT (10 microM) in the presence of extracellular Ca2+. Dil equally inhibited the steady state elevations of [Ca2+]i and force induced by 5-HT, whereas NG inhibited the force to a greater extent than expected from the reduction in [Ca2+]i. In the permeabilized ophthalmic artery smooth muscle, NG (10 microM), but not Dil (10 microM), decreased the force development induced by GTP (10 microM) and 5-HT (10 microM) at constant [Ca2+]i (pCa 6.5). These results indicate that NG, but not Dil, decreases the Ca2+ sensitivity of contractile apparatus.


CONCLUSIONS
The authors found that 5-HT contracts the ophthalmic artery smooth muscle by the elevation of [Ca2+]i mediated by the release of intracellular Ca2+ and the influx of extracellular Ca2+, as well as by an increase in the Ca2+ sensitivity of the contractile apparatus through the activation of G-proteins, and that Dil relaxes 5-HT-mediated contraction of ophthalmic artery primarily by inhibiting the Ca2+ influx and, hence, by decreasing [Ca2+]i without having any effect on the Ca2+ sensitivity of the contractile apparatus. Nitroglycerin relaxes the ophthalmic artery not only by decreasing [Ca2+]i (inhibition of both the Ca2+ release and Ca2+ influx) but also by decreasing the Ca2+ sensitivity of the contractile apparatus.
Hiroishi
G
G
Division of Molecular Cardiology, Faculty of Medicine, Kyushu University, Fukuoha, Japan.
Kobayashi
S
S
Nishimura
J
J
Inomata
H
H
Kanaide
H
H
D000818:Animals; D002110:Caffeine / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism* / Q000494:pharmacology; D002121:Calcium Channel Blockers / Q000494:pharmacology*; D002417:Cattle; D003600:Cytosol / Q000378:metabolism; D004110:Diltiazem / Q000494:pharmacology*; D005260:Female; D016257:Fura-2 / Q000378:metabolism; D008297:Male; D009119:Muscle Contraction / Q000187:drug effects; D009131:Muscle, Smooth, Vascular / Q000187:drug effects / Q000502:physiology*; D005996:Nitroglycerin / Q000494:pharmacology*; D009880:Ophthalmic Artery / Q000187:drug effects / Q000502:physiology*; D010726:Phosphodiesterase Inhibitors / Q000494:pharmacology; D012701:Serotonin / Q000494:pharmacology; D014665:Vasodilator Agents / Q000494:pharmacology*
D002121:Calcium Channel Blockers; D010726:Phosphodiesterase Inhibitors; D014665:Vasodilator Agents; D012701:Serotonin; D002110:Caffeine; D004110:Diltiazem; D005996:Nitroglycerin; D002118:Calcium; D016257:Fura-2