Bimodal Control of a Ca 2 1 - activated Cl 2 Channel by Different Ca 2 1 Signals

Ca 2 1 -activated Cl 2 channels play important roles in a variety of physiological processes, including epithelial secretion, maintenance of smooth muscle tone, and repolarization of the cardiac action potential. It remains unclear, however, exactly how these channels are controlled by Ca 2 1 and voltage. Excised inside-out patches containing many Ca 2 1 -activated Cl 2 channels from Xenopus oocytes were used to study channel regulation. The currents were mediated by a single type of Cl 2 channel that exhibited an anionic selectivity of I 2 . Br 2 . Cl 2 (3.6:1.9:1.0), irrespective of the direction of the current flow or [Ca 2 1 ]. However, depending on the amplitude of the Ca 2 1 signal, this channel exhibited qualitatively different behaviors. At [Ca 2 1 ] , 1 m M, the currents activated slowly upon depolarization and deactivated upon hyperpolarization and the steady state current–voltage relationship was strongly outwardly rectifying. At higher [Ca 2 1 ], the currents did not rectify and were time independent. This difference in behavior at different [Ca 2 1 ] was explained by an apparent voltage-dependent Ca 2 1 sensitivity of the channel. At 1 120 mV, the EC 50 for channel activation by Ca 2 1 was approximately fourfold less than at 2 120 mV (0.9 vs. 4 m M). Thus, at [Ca 2 1 ] , 1 m M, inward current was smaller than outward current and the currents were time dependent as a consequence of voltage-dependent changes in Ca 2 1 binding. The voltage-dependent Ca 2 1 sensitivity was explained by a kinetic gating scheme in which channel activation was Ca 2 1 dependent and channel closing was voltage sensitive. This scheme was supported by the observation that deactivation time constants of currents produced by rapid Ca 2 1 concentration jumps were voltage sensitive, but that the activation time constants were Ca 2 1 sensitive. The deactivation time constants increased linearly with the log of membrane potential. The qualitatively different behaviors of this channel in response to different Ca 2 1 concentrations adds a new dimension to Ca 2 1 signaling: the same channel can mediate either excitatory or inhibitory responses, depending on the amplitude of the cellular Ca 2 1 signal. key words: ion channels • electrophysiology • ion channel gating • calcium signaling • ion transport I N T R O D U C T I O N Ca 2 1 -activated Cl 2 channels play fundamental roles in physiological processes in many tissues, including secretion in airway epithelium (Wagner et al., 1991; Gray et al., 1995), repolarization of the cardiac action potential (Zygmunt, 1994; Kawano et al., 1995; Wang et al., 1995; Collier et al., 1996), regulation of vascular tone (Nelson et al., 1997; Yuan, 1997; Nilius et al., 1997a,b), modulation of photoreceptor light responses (Barnes and Deschenes, 1992), olfactory transduction (Kurahashi and Yau, 1994), neuronal excitability (DeCastro et al., 1997), regulation of platelet cell volume (Fine et al., 1994), and fast block to polyspermy in oocytes (Jaffe and Cross, 1986). Ca 2 1 -activated Cl 2 channels may be involved in several human diseases, including cystic fibrosis and cardiac arrhythmias. Although defects in the CFTR Cl 2 channel cause cystic fibrosis, upregulation of a Ca 2 1 -activated Cl 2 current in the airway of CFTR knockout mice can compensate for the lack of CFTR and ameliorate the pathology in this mouse model (Clarke et al., 1994; Grubb et al., 1994). Furthermore, overexpression of CFTR in cultured airway epithelial cells from CF patients results in a decrease in Ca 2 1 -activated Cl 2 current (Johnson et al., 1995). Ca 2 1 -activated Cl 2 channels also play a role in the repolarization of the cardiac action potential and contribute to the transient outward current (I to ) (Zygmunt and Gibbons, 1991, 1992; Zygmunt, 1994; Zygmunt et al., 1998.). Recently, it has been shown that dogs which are genetically prone to cardiac sudden death have an abnormal I to (Freeman et al., 1997), implying that Ca 2 1 -activated Cl 2 channels might play a role in cardiac sudden death. Ca 2 1 -activated Cl 2 channels also contribute to the transient inward current (Han and Ferrier, 1992, 1996), which is believed to trigger cardiac arrhythmias during Ca 2 1 overload (Berlin et al., 1989). Despite the importance of Ca 2 1 -activated Cl 2 channels in cell physiology, our understanding of the mechanisms of regulation and gating of these channels remains rudimentary. In different studies and cell types, Ca 2 1 -activated Cl 2 currents behave differently. Often, these currents are voltage sensitive: they activate slowly on depolarization and deactivate on hyperpolarization (e.g., Cliff and Frizzell, 1990; Arreola et al., 1996; Nilius et al., 1997b; Anderson and Welsh, 1998), but in other studies the currents appear time and voltage independent (e.g., Xie et al., 1996; Collier et al., 1996). It is not Address correspondence to H. Criss Hartzell, 1648 Pierce Dr., Department of Cell Biology, Emory University School of Medicine, Atlanta, GA 30322-3030. Fax: 404-727-6256; E-mail: [email protected] on Jne 1, 2017 D ow nladed fom Published December 28, 1999