Monovalent cation and L‐type Ca2+ channels participate in calcium paradox‐like phenomenon in rabbit aortic smooth muscle cells

Removal of extracellular Ca2+ followed by Ca2+ readmission produces pathological changes in a variety of mammalian cells and organs. This phenomenon was first described in the heart and called the ‘calcium paradox’ (Zimmerman & Hulsmann, 1966). It is thought (for review see Chapman & Tunstall, 1987) that during exposure of cardiomyocytes to Ca2+-free media, Na+ enters the cell significantly increasing the intracellular Na+ concentration. As a result, following extracellular Ca2+ readmission, there is a massive Ca2+ influx into the cell carried by the Na+-Ca2+ exchanger working in the reverse mode. In spite of many studies, the role and mechanism for Na+ influx remain controversial (Van Echteld et al. 1998). Na+ current through L-type Ca2+ channels is thought to be the major pathway; however, inhibition of these channels is known to decrease but not eliminate calcium paradox. Recently, Mubagwa et al. (1997) showed that a monovalent cation (MC) current develops in cardiomyocytes in the absence of extracellular divalent cations which conduct Na+ and produce depolarization which can activate voltage-gated Ca2+ channels. They proposed that the MC current plays an important role in triggering calcium paradox in the heart. A similar MC current sensitive to extracellular divalent cations has also been found in amphibian epithelial cells (Van Driessche & Zeiske, 1985), chick embryo ectoderm (Li et al. 1994; Sabovik et al. 1996), and in Xenopus oocytes (Arellano et al. 1995). The physiological significance of MC current remains uncertain, but it can be one of the major mechanisms underlying the pathological calcium paradox in a variety of cells. A calcium paradox-like phenomenon (similar to the one described in cardiomyocytes and the heart) has also been observed in smooth muscle cells (SMCs) and blood vessels (Kobayashi et al. 1985; Kutsky & Hester, 1986; Lindner & Heinle, 1987), but the mechanisms of its development are not understood. Contraction of blood vessels under conditions which induce calcium paradox in the heart is of particular interest because of the importance of coronary blood flow to cardiac function. In this respect, it is important to determine the distinct pathways which in the absence of extracellular divalent cations are responsible for pathological Na+ influx in SMCs leading to vessel contraction upon Ca2+ readmission. The present study is the first attempt to determine the ion channel-mediated mechanisms of Na+ influx in experimental calcium paradox conditions in individual SMCs, and to determine their effects on the behaviour of the whole blood vessel. Using the perforated patch-clamp technique in isolated SMCs from rabbit aorta, we describe a nifedipine-insensitive monovalent cation (MC) current which develops in Ca2+,Mg2+-free solution. When all the residual extracellular Ca2+ was chelated by EGTA, in addition to MC current, a nifedipine-sensitive Na+ current through L-type Ca2+ channels develops in SMCs (Mironneau et al. 1982; for review also see Tsien et al. 1987). The results in rabbit aortic rings indicated that under calcium paradox conditions, nifedipine-insensitive MC current alone can produce Na+ influx and depolarization of SMCs sufficient for contraction of the vessel upon Ca2+ readmission. Activation of Na+ influx through L-type Ca2+ channels can be triggered by MC-induced depolarization and occurred only in the presence of EGTA which greatly potentiated vessel contraction. Thus, Na+ influx through both MC and L-type Ca2+ channels could precede the development of SMC contraction under experimental calcium paradox conditions. A preliminary report of this work has appeared as an abstract (Zakharov et al. 1998).