Surface charge and properties of cardiac ATP-sensitive K+ channels.

Abstract

ATP-sensitive K + (KATP) channels are present in a wide variety of tissues. The sensitivity of these channels to closure by cytosolic ATP (ATPi) varies significantly among different tissues and even within the same tissue. The purpose of this study was to test the hypothesis that negative surface charges modulate the sensitivity of the KATe channels to ATPi by influencing surface potential in the vicinity of the ATP-binding site(s) of the channel. Unitary currents through KATe channels were measured in inside-out membrane patches excised from rabbit ventricular myocytes using the patch-clamp technique. Agents known to be effective at screening negative surface charges were applied to the cytosolic surface of the patches, and their effects on ATP sensitivity were examined. These agents included Mg 2+ (2-15 mM), Ba 2+ (2-10 mM), and the polycations protamine (0.01-10 ~,M), poly-L-lysine (500 p,M), and poly-L-arginine (0.5 I~M). The divalent cations and the various polycations all dramatically reduced the concentration of ATP i required to half-maximally suppress current through KATe channels (Kd), from ~ 100 IxM in the absence of these agents to 1.6-8 ~M in their presence. The effects were dose dependent. Protamine also reduced the sensitivity of KA~ channels to block by cytosolic ADP. The sensitivity of KATV channels to block by ATP was independent of membrane potential, suggesting that the ATP-binding site is not located within the transmembrane voltage field. The effects of the polycation poly-L-lysine on ATP sensitivity were also independent of membrane potential or the direction (inward or outward) of current through KATe channels. In addition to increasing ATP sensitivity, Mg 2+, Ba 2+, and the polycations all caused dose-dependent block of inward and outward currents through KAav channels over similar concentration ranges as their effects on ATP sensitivity. The block of inward current by polycations was not associated with reduction of single-channel conductance or evidence of fast open channel block. However, the polycations did cause a modest reduction in singlechannel conductance of outward current. These results are consistent with the presence of negative surface charges that reduce the local ATP concentration at the ATP-binding site(s) on the channel, relative to the bulk cytosolic ATP concentration. Screening these negative surface charges with divalent cations or polycations on May 12, 2018 jgp.rupress.org Downloaded from http://doi.org/10.1085/jgp.104.4.773 Published Online: 1 October, 1994 | Supp Info: 774THE JOURNAL OF GENERAL PHYSIOLOGY 9 VOLUME 104 9 1994 decreases the local ATP gradient, resulting in a decrease in the apparent Kd for ATP. Applying surface potential theory to the Mg 2+ data, a rough calculation of the surface charge density necessary to account for these effects yielded a value of 1 negative charge per 199/~2, corresponding to surface potentials in the range of -50 to -70 inV. The putative negative surface charges are most likely located on the KATe channel protein it...

Sulfonylurea and K+-Channel Opener Sensitivity of KATP Channels

“…Treatment with polycations can reverse the stimulatory actions of PIP 2 on open probability and ATP sensitivity (Deutsch et al 1994; Shyng and Nichols 1998), probably by shielding the negative charges introduced by PIP 2 . As shown in Fig.…”

Isoproterenol Opens K+ ATPChannels via a β2-Adrenoceptor-linked Mechanism in Sertoli Cells from Immature Rats

“…This means that the K + ATP channels are also modulated by ionic interactions. Polyca− tions such as polylysine, polyamines, neomycin, and LaCl 3 , which are known to bind to phospholipids and neutralise their negative charges, abolish the effect of PIPs on ATP inhibition [22].…”

Alteration of the membrane lipid environment by L -palmitoylcarnitine modulates K ATP channels in guinea-pig ventricular myocytes

“…Thus, L-PC potentially inhibited the K ATP channels by interacting with Kir6.2. Deutch et al have demonstrated that polycations can screen surface negative charges, thereby increasing the sensitivity of K ATP channels in rabbit ventricular myocytes to ATP [4]. Since L-PC also interacts with membrane negative charges and thus has surface negative charge-screening effects [11] we examined whether L-PC also modifies the sensitivity of K ATP channels to ATP.…”