title: Pharmacological evidence for capacitative Ca(2+) entry in cannulated and pressurized skeletal muscle arterioles.
abstract: Arteriolar myogenic tone shows a marked dependency on extracellular Ca(2+). The contribution played by mechanisms such as intracellular Ca(2+) release and capacitative entry, however, are less certain. The present studies aimed to demonstrate functional evidence for involvement of such mechanisms in myogenic tone and reactivity. Single cremaster arterioles were denuded of endothelium, pressurized under no-flow conditions and loaded with fura 2-AM for measurement of changes in intracellular Ca(2+) [Ca(2+)](i). The cell permeable, putative, IP(3) receptor antagonist 2APB (2 aminoethoxydiphenyl borate) was used to determine the possible role of IP(3) receptor-mediated mechanisms in arteriolar myogenic tone and reactivity. Arterioles dilated in response to increasing concentrations of 2APB (1 - 300 microM) without a concomitant change in global [Ca(2+)](i). Also 2APB (50 microM) completely inhibited the myogenic constriction in response to a step change in luminal pressure (50 - 120 mmHg) with no apparent effect on pressure-mediated increases in [Ca(2+)](i). 2APB markedly attenuated the constrictor response and [Ca(2+)](i) increase stimulated by phenylephrine but not KCl. Capacitative Ca(2+) influx in arterioles was demonstrated either by re-addition of extracellular [Ca(2+)] following pre-treatment with 1 or 10 microM nifedipine in Ca(2+) free buffer or exposure of vessels to thapsigargin (1 microM) to induce store depletion. In both cases 2APB inhibited the increase in [Ca(2+)](i). Capacitative Ca(2+) entry showed an inverse relationship with intraluminal pressure over the range 10 - 120 mmHg. Consistent with an effect on a Ca(2+) entry pathway, 2APB had no effect on intracellular (caffeine releasable) Ca(2+) stores while decreasing the rate of Mn(2+) quench of fura 2 fluorescence. The results provide functional evidence for capacitative Ca(2+) entry in intact arteriolar smooth muscle. The effectiveness of 2APB in inhibiting both non-voltage gated Ca(2+) entry and responsiveness to an acute pressure step is consistent with the involvement of an axis involving IP(3)-mediated and or capacitative Ca(2+) entry mechanisms in myogenic reactivity. Given the lack of effect of 2APB on pressure-induced changes in global [Ca(2+)](i) it is suggested that such mechanisms participate on a localized level to couple the myogenic stimulus to contraction.
authors.affiliation: Microvascular Biology Group, School of Medical Sciences, Division of Biosciences, RMIT University, Plenty Road, Bundoora, Victoria 3083, Australia.
authors.forename: S J
authors.initials: SJ
authors.lastname: Potocnik
authors.forename: M A
authors.initials: MA
authors.lastname: Hill
mesh_terms: D000199:Actins / Q000187:drug effects / Q000378:metabolism; D000818:Animals; D001160:Arterioles / Q000187:drug effects* / Q000502:physiology; D001896:Boron Compounds / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002121:Calcium Channel Blockers / Q000494:pharmacology; D003599:Cytoskeleton / Q000187:drug effects / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004730:Endothelium, Vascular / Q000502:physiology; D066298:In Vitro Techniques; D008297:Male; D008345:Manganese / Q000494:pharmacology; D018482:Muscle, Skeletal / Q000098:blood supply*; D009131:Muscle, Smooth, Vascular / Q000187:drug effects / Q000378:metabolism; D009543:Nifedipine / Q000494:pharmacology; D010656:Phenylephrine / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D019284:Thapsigargin / Q000494:pharmacology; D014661:Vasoconstriction / Q000187:drug effects*; D014662:Vasoconstrictor Agents / Q000494:pharmacology; D014665:Vasodilator Agents / Q000494:pharmacology
chemical_list: D000199:Actins; D001896:Boron Compounds; D002121:Calcium Channel Blockers; D014662:Vasoconstrictor Agents; D014665:Vasodilator Agents; D010656:Phenylephrine; D008345:Manganese; D019284:Thapsigargin; C109986:2-aminoethoxydiphenyl borate; D009543:Nifedipine; D002118:Calcium