Effect of Adenosine and Intracellular GTP on K ATP Channels of Mammalian Skeletal Muscle R. Barrett-Jolley, A. Comtois, N.W. Davies, P.R. Stanfield, N.B. Standen Ion Channel Group, Department of Cell Physiology & Pharmacology, University of Leicester, PO Box 138, Leicester LE1 9HN, UK Received: 29 December 1995/Revised: 22 March 1996 Abstract. We investigated the action of adenosine and GTP on K ATP channels, using inside-out patch clamp recordings from dissociated single fibers of rat flexor digitorum brevis (FDB) skeletal muscle. In excised patches, K ATP channels could be activated by a combi- nation of an extracellular adenosine agonist and intracel- lular Mg 2+ -ATP and GTP or GTP--S. The activation required hydrolyzable ATP and could be partially re- versed with Mg 2+ , suggesting that it may involve a G- protein dependent phosphorylation of K ATP channels. We found that K ATP channels of the rat FDB could not be activated by Mg 2+ -ATP alone or by Mg 2+ -ATP in the presence of extracellular adenosine. Patches whose channel activity had been ‘rundown’ by Ca 2+ could not be recovered by adenosine, GTP or Mg 2+ -ATP. K ATP chan- nels activated by adenosine receptor agonists had a simi- lar ATP sensitivity to those under control conditions; but adenosine appears to be able to switch these K ATP chan- nels from an inactive to an active mode. Key words: Potassium channel — Adenosine — Aden- osine triphosphate (ATP) — Skeletal muscle Introduction ATP-dependent potassium channels (K ATP channels) oc- cur in several tissues including muscle, pancreatic beta cells, some neurones and epithelia (Ashcroft & Ashcroft, 1990). In many of these tissues, it is unlikely that changes in intracellular ATP concentration ([ATP] i ) form the major regulator of channel activity under physi- ological conditions, since [ATP] i often changes little ex- cept under conditions of severe metabolic stress. Rather, ATP binding may set a low background open probability against which other regulators, including signalling sys- tems linked to G-protein coupled receptors, serve to con- trol channel activity. In skeletal muscle, ATP is espe- cially well buffered by creatine phosphate and creatine kinase (Carlson & Siger, 1960), and it seems certain that regulators other than ATP are involved in the physiologi- cal control of K ATP channel activity: one of these may be intracellular pH (Davies, Standen & Stanfield, 1992). It has also been suggested that adenosine released from active muscle fibers under conditions of systemic hyp- oxia opens K ATP channels, to increase K + efflux and cause a rise in extracellular [K + ] and so vasodilation (Marshall, Thomas & Turner, 1993; Comtois et al., 1994). Receptors for adenosine have been shown to be present in the sarcolemmal membrane of skeletal muscle fibers (Challiss, Richards & Budohoski, 1992) and the activation of K ATP channels by adenosine has been dem- onstrated in both cardiac muscle and coronary arterial smooth muscle (Kirsch et al., 1990; Dart & Standen, 1993). In cardiac cells, the link between the receptor and the channel apparently occurs via G i , since the applica- tion of G i -subunits to the cytoplasmic face of the mem- brane activates K ATP channels (Terzic et al., 1994). Regulation of K ATP channels by receptors coupled to G proteins has not so far been demonstrated in skeletal muscle, though GTP--S has been reported to activate K ATP channels reincorporated from t-tubule membrane into lipid bilayers (Parent & Coronado, 1989). We have previously reported that when membrane patches from rat skeletal muscle fibers were exposed to intracellular GTP, in the presence of extracellular adenosine and in- tracellular ATP, a modest increase in K ATP channel ac- tivity was observed (Barrett-Jolley et al., 1995). We have found that in the presence of extracellular adeno- sine, the removal of intracellular ATP elicits a substantial increase in the subsequent K ATP channel activity. A pos- sible explanation for this is that ATP plays a dual role in the regulation of skeletal K ATP channels, so that while Correspondence to: R. Barrett-Jolley J. Membrane Biol. 152, 111–116 (1996) The Journal of Membrane Biology © Springer-Verlag New York Inc. 1996