Agonist-bound muscarinic receptors open atrial K ؉ channels through a GTP-dependent pathway mediated by the G protein G k . However, nucleotides other than GTP are also able to support channel activity, even in the absence of agonists. This process was proposed to be mediated by nucleoside-diphosphate (NDP) kinase, which would transfer phosphate from nucleotide triphosphates to the GDP bound to G k , producing G k -GTP without the need for receptor-induced GDP-GTP exchange. We examined the

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... t of antibodies to NDP kinase on the ATP-supported activity of atrial muscarinic K ؉ channels and the corresponding GIRK1/CIR channels expressed in HEK 293 cells. Inhibitory antibodies reduced ATP-induced channel openings, but this effect displayed an absolute requirement for agonist and was also seen with antibodies that do not inhibit the enzyme. Both types of antibodies also reduced agonistdependent channel activity in the presence of GTP, ruling out a role for NDP kinase in GDP rephosphorylation. Channel activity was not affected by the antibodies in preparations where ATP-induced muscarinic channels are not under tight receptor control, namely pertussis toxin-treated atrial patches and membranes from cells expressing K ACh channel subunits. Thus, participation of NDP kinase in this pathway requires activated receptors and has a function distinct from phosphate transfer between nucleotides.