Effects of superoxide anion on intracellular Ca²⁺ concentration in rabbit pulmonary arterial smooth muscle cells

AIM: To study the effect of superoxide anion on the Ca²⁺ homeostasis in smooth muscle cells isolated from the rabbit pulmonary artery. METHODS: Intracellular Ca²⁺ concentration ([Ca²⁺](i)) was investigated using cell suspension of freshly isolated smooth muscle cells from rabbit pulmonary artery (PASMC). Fura-2 fluorescent ratio obtained at 340 nm and 380 nm wave lengths was measured as an indicator of [Ca²⁺](i). RESULTS: ATP 30 μmol·L^-1 induced a transient increase in the ratio (Ca²⁺ transient). Thapsigargin, an inhibitor of sarcoplasmic Ca²⁺ ATPase, induced a phasic increase in the ratio due to Ca²⁺ leak from intracellular store sites, but not the sustained increase, thereby suggesting the absence of Ca²⁺ release- activated Ca²⁺ entry (CRAC) mechanism in PASMC. When PASMC were exposed to superoxide anion by the pretreatment with xanthine and xanthine oxidase (X/XO) for 30 min, sustained component of ATP-induced Ca²⁺ transient was elevated. The ratios at 5 and 10 min after ATP application (Δratio₅ (min) and Δratio₁₀(min)) were increased from 0.091 ± 0.022 to 0.149 ± 0.048 (P < 0.05) and from 0.021 ± 0.020 to 0.117 ± 0.047 (P < 0.01), respectively. But, thapsigargin-induced [Ca²⁺](i) transient was not affected by X/XO. CONCLUSION: Superoxide anion makes ATP induced Ca²⁺ transient sluggish, and does not affect Ca²⁺ leak pathway in PASMC.