Pacemaker Channels Produce an Instantaneous Current

Spontaneous rhythmic activity in mammalian heart and brain depends on pacemaker currents (Ih), which are produced by hyperpolarization-activated cyclic nucleotide-gated (HCN) channels. Here, we report that the mouse HCN2 pacemaker channel isoform also produced a large instantaneous current (Iinst(HCN2)) in addition to the well characterized, slowly activating Ih. Iinst(HCN2) was specific to expression of HCN2 on the plasma membrane and its amplitude was correlated with that of Ih. The two currents had similar reversal potentials, and both were modulated by changes in intracellular Cl− and cAMP. A mutation in the S4 domain of HCN2 (S306Q) decreased Ih but did not alter Iinst(HCN2), and instantaneous currents in cells expressing either wild type HCN2 or mutant S306Q channels were insensitive to block by Cs+. Co-expression of HCN2 with the accessory subunit, MiRP1, decreased Ih and increased Iinst(HCN2), suggesting a mechanism for modulation of both currents in vivo. These data suggest that expression of HCN channels may be accompanied by a background conductance in native tissues and are consistent with at least two open states of HCN channels: Iinst(HCN2) is produced by a Cs+-open state; hyperpolarization produces an additional Cs+-sensitive open state, which results in Ih. Spontaneous rhythmic activity in mammalian heart and brain depends on pacemaker currents (Ih), which are produced by hyperpolarization-activated cyclic nucleotide-gated (HCN) channels. Here, we report that the mouse HCN2 pacemaker channel isoform also produced a large instantaneous current (Iinst(HCN2)) in addition to the well characterized, slowly activating Ih. Iinst(HCN2) was specific to expression of HCN2 on the plasma membrane and its amplitude was correlated with that of Ih. The two currents had similar reversal potentials, and both were modulated by changes in intracellular Cl− and cAMP. A mutation in the S4 domain of HCN2 (S306Q) decreased Ih but did not alter Iinst(HCN2), and instantaneous currents in cells expressing either wild type HCN2 or mutant S306Q channels were insensitive to block by Cs+. Co-expression of HCN2 with the accessory subunit, MiRP1, decreased Ih and increased Iinst(HCN2), suggesting a mechanism for modulation of both currents in vivo. These data suggest that expression of HCN channels may be accompanied by a background conductance in native tissues and are consistent with at least two open states of HCN channels: Iinst(HCN2) is produced by a Cs+-open state; hyperpolarization produces an additional Cs+-sensitive open state, which results in Ih. Hyperpolarization-activated “pacemaker” currents in mammalian heart (If) 1The abbreviations used are:Ih or Ifhyperpolarization-activated currentHCN2hyperpolarization-activated cyclic nucleotide-gated channel, isoform 2Iinst(HCN2)instantaneous current in CHO cells expressing HCN2GFPgreen fluorescent proteinIinst(GFP)instantaneous current in CHO cells expressing GFPSAsino-atrialCHOChinese hamster ovaryCNBDcyclic nucleotide-binding domainKvvoltage-gated potassium channelIbsodium-dependent background currentPBSphosphate-buffered saline 1The abbreviations used are:Ih or Ifhyperpolarization-activated currentHCN2hyperpolarization-activated cyclic nucleotide-gated channel, isoform 2Iinst(HCN2)instantaneous current in CHO cells expressing HCN2GFPgreen fluorescent proteinIinst(GFP)instantaneous current in CHO cells expressing GFPSAsino-atrialCHOChinese hamster ovaryCNBDcyclic nucleotide-binding domainKvvoltage-gated potassium channelIbsodium-dependent background currentPBSphosphate-buffered saline and brain (Ih or Iq) contribute to generation and modulation of spontaneous rhythmic activity (1DiFrancesco D. Annu. Rev. Physiol. 1993; 55: 455-472Crossref PubMed Scopus (658) Google Scholar, 2Pape H.C. Annu. Rev. Physiol. 1996; 58: 299-327Crossref PubMed Scopus (969) Google Scholar). These currents are carried by Na+ and K+ and have reversal potentials of about −20 mV in physiological solutions (3Accili E.A. DiFrancesco D. Pflugers Arch. 1996; 431: 757-762Crossref PubMed Scopus (35) Google Scholar). Pacemaker currents, collectively referred to here as Ih, activate slowly upon hyperpolarization negative to a threshold of ∼−45 mV (depending on the tissue). Cyclic AMP (cAMP) shifts the midpoint of Ih activation to more positive potentials through a direct, phosphorylation-independent mechanism that has no effect on the maximal conductance (4DiFrancesco D. Tortora P. Nature. 1991; 351: 145-147Crossref PubMed Scopus (636) Google Scholar). Extracellular Cs+ is a voltage-dependent blocker of Ih (5DiFrancesco D. J. Physiol. (Lond.). 1981; 314: 359-376Crossref Scopus (262) Google Scholar). Ih is not carried by Cl−, but extracellular Cl− is required for production of Ih (6Frace A.M. Maruoka F. Noma A. J. Physiol. 1992; 453: 307-318Crossref PubMed Scopus (41) Google Scholar) and variations in intracellular Cl− have been shown to modify the conductance of Ih (7Lenz R.A. Pitler T.A. Alger B.E. J. Neurosci. 1997; 17: 6133-6141Crossref PubMed Google Scholar). hyperpolarization-activated current hyperpolarization-activated cyclic nucleotide-gated channel, isoform 2 instantaneous current in CHO cells expressing HCN2 green fluorescent protein instantaneous current in CHO cells expressing GFP sino-atrial Chinese hamster ovary cyclic nucleotide-binding domain voltage-gated potassium channel sodium-dependent background current phosphate-buffered saline hyperpolarization-activated current hyperpolarization-activated cyclic nucleotide-gated channel, isoform 2 instantaneous current in CHO cells expressing HCN2 green fluorescent protein instantaneous current in CHO cells expressing GFP sino-atrial Chinese hamster ovary cyclic nucleotide-binding domain voltage-gated potassium channel sodium-dependent background current phosphate-buffered saline The hyperpolarization-activated, cyclic nucleotide-gated (HCN) channels that produce Ih have been recently cloned (8Santoro B. Grant S.G. Bartsch D. Kandel E.R. Proc. Natl. Acad. Sci. U. S. A. 1997; 94: 14815-14820Crossref PubMed Scopus (228) Google Scholar, 9Santoro B. Liu D.T. Yao H. Bartsch D. Kandel E.R. Siegelbaum S.A. Tibbs G.R. Cell. 1998; 93: 717-729Abstract Full Text Full Text PDF PubMed Scopus (576) Google Scholar, 10Ludwig A. Zong X. Jeglitsch M. Hofmann F. Biel M. Nature. 1998; 393: 587-591Crossref PubMed Scopus (778) Google Scholar, 11Ishii T.M. Takano M. Xie L.H. Noma A. Ohmori H. J. Biol. Chem. 1999; 274: 12835-12839Abstract Full Text Full Text PDF PubMed Scopus (232) Google Scholar, 12Gauss R. Seifert R. Kaupp U.B. Nature. 1998; 393: 583-587Crossref PubMed Scopus (374) Google Scholar). Their amino acid sequence predicts a structure similar to that of voltage-gated potassium (Kv) channels and cyclic nucleotide-gated channels. Thus, HCN subunits are thought to assemble as tetramers, with each subunit containing six transmembrane domains. Although HCN channels are activated by hyperpolarization, a positively charged S4 domain appears to act as a voltage sensor (13Chen J. Mitcheson J.S. Lin M. Sanguinetti M.C. J. Biol. Chem. 2000; 275: 36465-36471Abstract Full Text Full Text PDF PubMed Scopus (69) Google Scholar, 14Vaca L. Stieber J. Zong X. Ludwig A. Hofmann F. Biel M. FEBS Lett. 2000; 479: 35-40Crossref PubMed Scopus (41) Google Scholar), as it does in depolarization-activated channels (15Horn R. Biochemistry. 2000; 39: 15653-15658Crossref PubMed Scopus (38) Google Scholar, 16Bezanilla F. Physiol. Rev. 2000; 80: 555-592Crossref PubMed Scopus (711) Google Scholar). Some mutations in the S4 domain of HCN channels reduce Ih but produce large instantaneous currents (13Chen J. Mitcheson J.S. Lin M. Sanguinetti M.C. J. Biol. Chem. 2000; 275: 36465-36471Abstract Full Text Full Text PDF PubMed Scopus (69) Google Scholar, 17Chen J. Mitcheson J.S. Tristani-Firouzi M. Lin M. Sanguinetti M.C. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 11277-11282Crossref PubMed Scopus (132) Google Scholar), and an instantaneous current has been noted for some heterologously expressed wild type HCN channels (11Ishii T.M. Takano M. Xie L.H. Noma A. Ohmori H. J. Biol. Chem. 1999; 274: 12835-12839Abstract Full Text Full Text PDF PubMed Scopus (232) Google Scholar,12Gauss R. Seifert R. Kaupp U.B. Nature. 1998; 393: 583-587Crossref PubMed Scopus (374) Google Scholar). HCN channels also have a COOH-terminal cyclic nucleotide-binding domain (CNBD) that is homologous to those of cyclic nucleotide-gated channels and other cyclic nucleotide-binding proteins (18Shabb J.B. Corbin J.D. J. Biol. Chem. 1992; 267: 5723-5726Abstract Full Text PDF PubMed Google Scholar). The CNBD is thought to inhibit activation of the channels by hyperpolarization; direct binding of cAMP is thought to shift the voltage dependence of the channels by relieving this inhibition (19Wainger B. DeGennaro M. Santoro B. Siegelbaum S. Tibbs G. Nature. 2001; 411: 805-810Crossref PubMed Scopus (388) Google Scholar). As for other voltage-gated channels, HCN channels probably exist as tetramers of different HCN α subunits (20Chen S. Wang J. Siegelbaum S.A. J Gen Physiol. 2001; 117: 491-504Crossref PubMed Scopus (322) Google Scholar, 21Ulens C. Tytgat J. J. Biol. Chem. 2001; 276: 6069-6072Abstract Full Text Full Text PDF PubMed Scopus (173) Google Scholar) complexed with other proteins. For instance, it has recently been shown that association of HCN2 with MinK-related peptide 1 (MiRP1) increases expression levels and activation rate (22Yu H., Wu, J. Potapova I. Wymore R.T. Holmes B. Zuckerman J. Pan Z. Wang H. Shi W. Robinson R.B., El- Maghrabi M.R. Benjamin W. Dixon J. McKinnon D. Cohen I.S. Wymore R. Circ. Res. 2001; 88: E84-87Crossref PubMed Google Scholar). MiRP1 is a member of the MinK family of single transmembrane-spanning proteins which have been shown to associate with an ever-growing list of voltage-gated channels, including KCNQ1 (23Barhanin J. Lesage F. Guillemare E. Fink M. Lazdunski M. Romey G. Nature. 1996; 384: 78-80Crossref PubMed Scopus (1368) Google Scholar, 24Sanguinetti M.C. Curran M.E. Spector P.S. Keating M.T. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 2208-2212Crossref PubMed Scopus (389) Google Scholar), HERG (25McDonald T.V., Yu, Z. Ming Z. Palma E. Meyers M.B. Wang K.W. Goldstein S.A. Fishman G.I. Nature. 1997; 388: 289-292Crossref PubMed Scopus Google Scholar), and and M. M. Circ. Res. 2001; 88: PubMed Scopus Google Scholar). association with a channels a voltage-gated to a S. M. Lazdunski M. J. J. 2000; PubMed Scopus Google Scholar, S. S. M. R. R. Nature. 2000; PubMed Scopus Google Scholar, S. M.C. Goldstein S.A. Cell. 2001; Full Text Full Text PDF PubMed Scopus Google Scholar). currents are a but of A is in the sino-atrial of the heart a background current H. H. S. J. Physiol. (Lond.). 1992; Scopus Google Scholar) has been to contribute to the of the H. W. Physiol. Rev. 1993; PubMed Scopus Google D. DiFrancesco D. Proc. R. Biol. Sci. 1992; PubMed Scopus Google Scholar). The of this background current has not been the of the of HCN2 channels expressed in Chinese hamster ovary we a large instantaneous current (Iinst(HCN2)) in addition to J. 2000; Scholar). here that Iinst(HCN2) was with expression of and the amplitude of Iinst(HCN2) was correlated with that of Ih. Iinst(HCN2) had in with both were modulated by were but not by intracellular and had similar reversal A mutation in the S4 domain (S306Q) that Ih had effect on the instantaneous current Iinst(HCN2) was by Cs+. These data suggest that Iinst(HCN2) through a of the HCN2 Co-expression of HCN2 with MiRP1 Ih but increased Iinst(HCN2), suggesting a mechanism for modulation of the two currents in vivo. The mutant HCN2 channels and S306Q were a mouse HCN2 A. Zong X. Jeglitsch M. Hofmann F. Biel M. Nature. 1998; 393: 587-591Crossref PubMed Scopus (778) Google Scholar) the mutations were by of cells were in with and and at with were in mammalian expression wild type or mutant HCN2 channels, the cells MiRP1 were the cells with a green fluorescent protein the expressing the proteins were by green expressing on were by binding of to 2 a of with cells was to a and with a K+ extracellular of the the was to a as in the The were with either a Cl− or which was with 1 cAMP in some as The of solutions used for is in I. currents were the with which had of with the intracellular were at were an and were at 2 and were and were at was not currents were as the current the for cells expressing were for cells in which a current was also of the green fluorescent The voltage dependence of activation was currents at mV were to of HCN2 channels. current were as a of and were with a for K+ extracellular in and solutions were to in a in and solutions were to 1 to the midpoint of activation and The single used in some were by a to mV to the channels, and the current was to to its the of were an in were or an with either a or for (depending on the was the was are as and the of cells which were a of for each in wild type and S306Q mutant channels is insensitive to Cs+. current produced by a expressing S306Q and current in cells expressing S306Q or HCN2 in to voltage to Ih the in current the and of the and a in with cells expressing were in the K+ extracellular and were with the Cl− intracellular of a CHO with a of instantaneous current in cells expressing S306Q or HCN2 in to voltage to currents were cells were in a K+ extracellular The Cs+ were the cells 1 to a K+ extracellular containing 2 Cs+. The was with a Cl− for to cells on were with and in in for The cells were with and with with containing the cells were with a to HCN2 at a of in with for at The was and cells were with were with a with at a of in with for 1 at in the was cells were in and the with cells were on were and For were in a and an of 1 A current CHO cells expressing either the GFP or GFP with the mouse HCN2 pacemaker channel isoform were with a K+ extracellular to current and were with the Cl− intracellular of solutions used for is in cells expressing HCN2 produced not the current but also a large instantaneous current a instantaneous current was in the GFP cells for and Iinst(HCN2) were to and are as a of voltage in 1 B. and Iinst(HCN2) had a voltage dependence with a of negative to about the which was the of the was for Iinst(HCN2) for and The reversal potentials of the instantaneous currents also mV and a different for with Ih and Iinst(HCN2) were we for a in Iinst(HCN2) in to voltage to mV was as a of Ih in the cells 1 A a positive These that the production of Ih and Iinst(HCN2), and that both currents may through the HCN2 to that Iinst(HCN2) was not a Cl− current Cl− currents in CHO cells X. S.A. J. Biol. Chem. 2000; 275: Full Text Full Text PDF PubMed Scopus Google Scholar) have been by HCN2 Thus, we and reversal potentials for Iinst(HCN2) and in intracellular solutions containing a or of Cl− was for Cl− in the for The and Cl− solutions reversal potentials for Cl− of and which were of the extracellular that the conductance of Iinst(HCN2) that of the Cl− intracellular and as it had been in the Cl− The reversal for and Iinst(HCN2) were the of instantaneous as those in B. The reversal for Ih was the activated in which the maximal Ih current was at potentials as the in current to mV HCN channels are and to mV the channels are These are as a of in 2 A. The reversal potentials for Iinst(HCN2) and Ih did not as in 2 for the extracellular either a Cl− or Cl− intracellular in reversal was we used the K+ extracellular the reversal potentials with Cl− in the were mV and mV for Iinst(HCN2) and Ih, These are similar to those for Ih. (3Accili E.A. DiFrancesco D. Pflugers Arch. 1996; 431: 757-762Crossref PubMed Scopus (35) Google Scholar). These that Iinst(HCN2) and Ih have similar consistent with the that through the the reversal potentials for Iinst(HCN2) and Ih were both more positive intracellular Cl− was intracellular Cl− was the reversal for did not the two solutions 2 that Ih Iinst(HCN2) was carried by Cl− the reversal for a have been more negative intracellular Cl− was no was in the reversal of in the a in for the more positive reversal potentials of Ih and Iinst(HCN2) in the Cl− Iinst(HCN2) was specific to expression of HCN2 on the plasma membrane not a of of membrane we expressed the which of the to the transmembrane domain the expressing on were by binding of in addition to green current in to to mV did not in cells expressing with those expressing and in both the instantaneous current was Iinst(HCN2) 2 the similar reversal potentials of and the effect of intracellular Cl− on Iinst(HCN2) and Ih are consistent with the that both currents through the HCN2 channel suggest that the is by intracellular to cAMP is a of HCN channels. Iinst(HCN2) was produced by the HCN2 channel, we Ih, was modulated by cAMP. we Ih and Iinst(HCN2) with or 1 cAMP in the the voltage dependence of Ih activation currents at mV and that it was more positive 1 cAMP was in the mV the we also a in the conductance of Iinst(HCN2) in the of this did not currents not with expression of HCN2 of the Iinst(HCN2) 1 and a effect of we used a more to a of Iinst(HCN2) by cAMP in to mV in cells expressing either HCN2 or GFP and with the the was to the containing and which cAMP in CHO cells J. H. A. I. H., M. F. J. Biol. Chem. 1999; 274: Full Text Full Text PDF PubMed Scopus Google Scholar). These of and produced a and in Iinst(HCN2) but not as for cells in C. 1 in the of Iinst(HCN2) in cells was with its cAMP no was in the of The effect of cAMP on Iinst(HCN2) was the was to the K+ extracellular A similar in Iinst(HCN2) was the was to containing 1 not that the of and were to increased intracellular cAMP. These results that Iinst(HCN2) is modulated by cAMP and the that HCN2 produces both Ih and an to the CNBD of HCN2 the cAMP of Iinst(HCN2), we a mutant channel in which the CNBD and of the were this mutant channel did not produce either Iinst(HCN2) or Ih in CHO cells The amplitude of the instantaneous current produced by did not and was Iinst(HCN2) a HCN2 a in on the of cells expressing with those expressing wild type HCN2 The and data suggest that the mutation expression of HCN2 by the of channel protein on the plasma has shown that expression of some mutant channels is by of mammalian cells at Z. J. Biol. Chem. 1999; 274: Full Text Full Text PDF PubMed Scopus Google Scholar). was not the for of the cells at for did not expression of Iinst(HCN2) or cells expressing data not with the that Iinst(HCN2) is specific to expression of HCN2 2 data suggest that the HCN2 protein be on the plasma membrane in for Iinst(HCN2) to be produced and a in which Iinst(HCN2) through the HCN2 channel A report that a mutation in the S4 of HCN2 (S306Q) Ih and produced a large instantaneous current (13Chen J. Mitcheson J.S. Lin M. Sanguinetti M.C. J. Biol. Chem. 2000; 275: 36465-36471Abstract Full Text Full Text PDF PubMed Scopus (69) Google Scholar). expressed the S306Q mutant in CHO cells and its to those of that Ih was by the S306Q mutation and as (13Chen J. Mitcheson J.S. Lin M. Sanguinetti M.C. J. Biol. Chem. 2000; 275: 36465-36471Abstract Full Text Full Text PDF PubMed Scopus (69) Google Scholar). in probably did not a decreased expression of the S306Q mutant channel in cells expressing S306Q was similar that in cells expressing the wild type HCN2 channel with The amplitude of the instantaneous current in cells expressing S306Q did not that of Iinst(HCN2) and and was by cAMP in the as Iinst(HCN2) not was produced by an of the mutant channels, as has been for some HCN2 S4 mutations J. Mitcheson J.S. Tristani-Firouzi M. Lin M. Sanguinetti M.C. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 11277-11282Crossref PubMed Scopus (132) Google Scholar), we 2 a voltage-dependent blocker of HCN2 A. Zong X. Jeglitsch M. Hofmann F. Biel M. Nature. 1998; 393: 587-591Crossref PubMed Scopus (778) Google Scholar, A. A. C. C. J. M. DiFrancesco D. Pflugers Arch. 2000; PubMed Scopus Google Scholar), to the extracellular that Cs+ Ih at mV not but had no effect on either or the instantaneous current 1 with a K+ extracellular containing 2 Cs+ did not those in the of Cs+ These data are consistent with a in which HCN2 has at least two open Iinst(HCN2) is produced by a state, Ih is produced a hyperpolarization-activated, Cs+-sensitive open The S306Q mutation has no effect on the instantaneous conductance but the hyperpolarization-activated The MinK family of single transmembrane-spanning proteins has been shown to instantaneous currents and currents with voltage-dependent of the family S. M. Lazdunski M. J. J. 2000; PubMed Scopus Google Scholar). member of this MiRP1 has been recently shown to associate with and the expression of HCN2 expressed in (22Yu H., Wu, J. Potapova I. Wymore R.T. Holmes B. Zuckerman J. Pan Z. Wang H. Shi W. Robinson R.B., El- Maghrabi M.R. Benjamin W. Dixon J. McKinnon D. Cohen I.S. Wymore R. Circ. Res. 2001; 88: E84-87Crossref PubMed Google Scholar). MiRP1 with HCN2 to Iinst(HCN2) and Ih in CHO that MiRP1 decreased Ih and increased Iinst(HCN2) of MiRP1 did not alter the instantaneous current with and no current was produced by either MiRP1 or GFP A and These suggest a physiological mechanism for modulation of Iinst(HCN2) and Ih by this we have shown that expression of the HCN2 pacemaker channel isoform in CHO cells produced a large instantaneous current (Iinst(HCN2)) in addition to the well current Iinst(HCN2) was shown to be specific to expression of HCN2 its amplitude was correlated with that of Ih and of the did not produce in instantaneous expression of the HCN2 protein on the plasma membrane was shown to be required for Iinst(HCN2) a HCN2 channel, which did not on the plasma did not produce either Ih or The of an channel is a for it more that the two currents are produced by the channel have both currents have similar and both are by changes in intracellular Cl− and by of cAMP. be in to the of Ih and Iinst(HCN2) the of Iinst(HCN2) by of in different cells this in the of a specific blocker for Iinst(HCN2) The S306Q mutation for the that Iinst(HCN2) through the HCN2 channel has that S4 as produce large instantaneous currents (13Chen J. Mitcheson J.S. Lin M. Sanguinetti M.C. J. Biol. Chem. 2000; 275: 36465-36471Abstract Full Text Full Text PDF PubMed Scopus (69) Google Scholar, 17Chen J. Mitcheson J.S. Tristani-Firouzi M. Lin M. Sanguinetti M.C. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 11277-11282Crossref PubMed Scopus (132) Google Scholar). have shown here that does not Iinst(HCN2), that both through the HCN2 channel the data here are consistent with a in which HCN2 channels have at least two open 2 Iinst(HCN2) through a open of Ih through the hyperpolarization-activated open The is insensitive to and the to is insensitive to The of Cs+ and by cAMP of both and Iinst(HCN2) that the two open states are and that may different of the to this the S306Q mutation does not the state, but the of the channel to the is consistent with the of (13Chen J. Mitcheson J.S. Lin M. Sanguinetti M.C. J. Biol. Chem. 2000; 275: 36465-36471Abstract Full Text Full Text PDF PubMed Scopus (69) Google Scholar) that is for with Cs+ and and the that the of and Iinst(HCN2) did not in CHO to that the mutant channels are in the the state; the channels are the S306Q the also appears to the of HCN2 channels to the be required to the of S306Q and MiRP1 are the The block of Ih and in Iinst(HCN2) produced MiRP1 is with HCN2 are similar to its with it current and increases instantaneous current S. M. Lazdunski M. J. J. 2000; PubMed Scopus Google Scholar). MiRP1 is expressed in the heart and a direct MiRP1 and HCN2 has been shown (22Yu H., Wu, J. Potapova I. Wymore R.T. Holmes B. Zuckerman J. Pan Z. Wang H. Shi W. Robinson R.B., El- Maghrabi M.R. Benjamin W. Dixon J. McKinnon D. Cohen I.S. Wymore R. Circ. Res. 2001; 88: E84-87Crossref PubMed Google Scholar). is to be for the in Iinst(HCN2) and the in have in CHO These were not the two proteins were in (22Yu H., Wu, J. Potapova I. Wymore R.T. Holmes B. Zuckerman J. Pan Z. Wang H. Shi W. Robinson R.B., El- Maghrabi M.R. Benjamin W. Dixon J. McKinnon D. Cohen I.S. Wymore R. Circ. Res. 2001; 88: E84-87Crossref PubMed Google Scholar), but this may be to the in expression A similar of expression was for the of KCNQ1 to a open channel by MiRP1, which was in mammalian cells and not in S. M. Lazdunski M. J. J. 2000; PubMed Scopus Google Scholar, F. I. M.E. K.W. Keating M.T. Goldstein S.A. Cell. 1999; Full Text Full Text PDF PubMed Scopus Google Scholar). H., Wu, J. Potapova I. Wymore R.T. Holmes B. Zuckerman J. Pan Z. Wang H. Shi W. Robinson R.B., El- Maghrabi M.R. Benjamin W. Dixon J. McKinnon D. Cohen I.S. Wymore R. Circ. 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Cell. 2000; Google Scholar) that suggest the potassium channels have both an instantaneous and a current the that the of both current is a to or voltage-gated potassium channels. Ludwig for the and Goldstein for the MiRP1

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