Adenosine Triphosphate Activates a Noninactivating K 1 Current in Adrenal Cortical Cells through Nonhydrolytic Binding

Bovine adrenal zona fasciculata (AZF) cells express a noninactivating K 1 current (I AC ) that is inhibited by adrenocorticotropic hormone and angiotensin II at subnanomolar concentrations. Since I AC appears to set the membrane potential of AZF cells, these channels may function critically in coupling peptide receptors to membrane depolarization, Ca 2 1 entry, and cortisol secretion. I AC channel activity may be tightly linked to the metabolic state of the cell. In whole cell patch clamp recordings, MgATP applied intracellularly through the patch electrode at concentrations above 1 mM dramatically enhanced the expression of I AC K 1 current. The maximum I AC current density varied from a low of 8.45 6 2.74 pA/pF ( n 5 17) to a high of 109.2 6 26.3 pA/pF ( n 5 6) at pipette MgATP concentrations of 0.1 and 10 mM, respectively. In the presence of 5 mM MgATP, I AC K 1 channels were tonically active over a wide range of membrane potentials, and voltage-dependent open probability increased by only z 30% between 2 40 and 1 40 mV. ATP (5 mM) in the absence of Mg 2 1 and the nonhydrolyzable ATP analog AMP-PNP (5 mM) were also effective at enhancing the expression of I AC , from a control value of 3.7 6 0.1 pA/pF ( n 5 3) to maximum values of 48.5 6 9.8 pA/pF ( n 5 11) and 67.3 6 23.2 pA/pF ( n 5 6), respectively. At the single channel level, the unitary I AC current amplitude did not vary with the ATP concentration or substitution with AMP-PNP. In addition to ATP and AMP-PNP, a number of other nucleotides including GTP, UTP, GDP, and UDP all increased the outwardly rectifying I AC current with an apparent order of effectiveness: MgATP . ATP 5 AMP-PNP . GTP 5 UTP . ADP .. GDP . AMP and ATPg -S. Although ATP, GTP, and UTP all enhanced I AC amplitude with similar effectiveness, inhibition of I AC by ACTH (200 pM) occurred only in the presence of ATP. As little as 50 m M MgATP restored complete inhibition of I AC , which had been activated by 5 mM UTP. Although the opening of I AC channels may require only ATP binding, its inhibition by ACTH appears to involve a mechanism other than hydrolysis of this nucleotide. These findings describe a novel form of K 1 channel modulation by which I AC channels are activated through the nonhydrolytic binding of ATP. Because they are activated rather than inhibited by ATP binding, I AC K 1 channels may represent a distinctive new variety of K 1 channel. The combined features of I AC channels that allow it to sense and respond to changing ATP levels and to set the resting potential of AZF cells, suggest a mechanism where membrane potential, Ca 2 1 entry, and cortisol secretion could be tightly coupled to the metabolic state of the cell through the activity of I AC K 1 channels. key words: adenosine triphosphate • potassium channel • adrenocorticotropic hormone • nucleotide i n t r o d u c t i o n I AC is a novel noninactivating K 1 current that may set the resting potential of bovine adrenal zona fasciculata (AZF) 1 cells. Angiotensin II (AII) and adrenocorticotropic hormone (ACTH) inhibit I AC and depolarize AZF cells at concentrations identical to those that stimulate cortisol production (Mlinar et al., 1993 a ). This K 1 channel appears to act pivotally in coupling these peptide hormone receptors to depolarization-dependent Ca 2 1 entry and corticosteroid production (Enyeart et al., 1993). I AC K 1 channel activity may be regulated by the complex interaction of biochemical factors and membrane voltage. In whole-cell patch clamp recordings from AZF cells, we found that I AC K 1 current measured during depolarizing voltage steps increases dramatically (10–100fold) over a period of many minutes (Mlinar et al., 1993 a ). Inhibitory factors present in the cytoplasm may be diluted during dialysis of the cell by pipette solution, allowing the functional expression of the I AC current. In this regard, the time-dependent growth of I AC is suppressed by including the nonhydrolyzable GTP analog GTPg -S in the recording pipette, indicating the presence of an inhibitory mechanism requiring a GTPbinding protein (Mlinar et al., 1993 a ). Accordingly, inhibition of I AC by both ACTH and AII require G-proAddress correspondence to Dr. John J. Enyeart, Department of Pharmacology, The Ohio State University, College of Medicine, 5188 Graves Hall, 333 W. 10th Avenue, Columbus, OH 43210-1239. Fax: 614-292-7232; E-mail: [email protected] 1 Abbreviations used in this paper: ACTH, adrenocorticotropic hormone; AII, angiotensin II; AZF, bovine adrenal fasciculata; CFTR, cystic fibrosis transmembrane conductance regulator; IV, current-voltage relationship. on Jne 1, 2017 D ow nladed fom Published December 1, 1997