DIFFERENTIAL EFFECTS OF HYPOXIA ON THE INTRACELLULAR Ca²⁺ CONCENTRATION OF MYOCYTES ISOLATED FROM DIFFERENT REGIONS OF THE RAT PULMONARY ARTERIAL TREE

Abstract

Hypoxic pulmonary vasoconstriction (HPV) may be mediated, in part, by an oxygen-sensing mechanism intrinsic to pulmonary arterial smooth muscle. It has been proposed that hypoxia inhibits a K⁺ conductance, which promotes membrane depolarization, subsequent activation of L-type Ca²⁺ channels and ultimately constriction. We have monitored hypoxia-induced changes in the intracellular Ca²⁺ concentration ([Ca²⁺]_i) of single myocytes isolated from the rat pulmonary arterial tree using microspectrofluorimetry and ratiometric measurement of indo-1 fluorescence. The basal level of [Ca²⁺]_i was in the range 35-80 nM and cells were quiescent at rest exhibiting no spontaneous oscillations in [Ca²⁺]_i. When the extracellular K⁺ concentration ([K⁺]_o) was raised to 20 mM, the [Ca²⁺]_i increased from [similar]60 to [similar]100 nM. This increase was abolished by nifedipine, demonstrating the presence, and need for activation, of functional voltage-gated L-type Ca²⁺ channels. Hypoxia (P_O2 [less than or equal to] 30 mmHg; throughout) had little effect on the resting [Ca²⁺]_i in myocytes isolated from either the main intrapulmonary artery, or its primary, secondary or tertiary branches. However, upon raising the [Ca²⁺]_i by increasing [K⁺]_o to 20 mM, hypoxia was found to lower [Ca²⁺]_i, from [similar]110 to [similar]70 nM, in cells isolated from the main conduit and primary branches of the intrapulmonary artery. In marked contrast, when [Ca²⁺]_i was raised, by increasing [K⁺]_o to 20 mM, in myocytes isolated from secondary and tertiary branches of the intrapulmonary artery, hypoxia induced a further reversible increase in the [Ca²⁺]_i from [similar]160 to [similar]240 nM. Neither hypoxia alone nor in combination with 20 mM K⁺_o induced any increase in the [Ca²⁺]_i in the presence of nifedipine. We conclude that hypoxia may modulate [Ca²⁺]_i in rat pulmonary artery myocytes only following its elevation by a depolarizing stimulus.