Mechanisms underlying the activation of G-protein–gated inwardly rectifying K⁺(GIRK) channels by the novel anxiolytic drug, ML297

Abstract

ML297 was recently identified as a potent and selective small molecule agonist of G-protein-gated inwardly rectifying K + (GIRK/Kir3) channels. Here, we show ML297 selectively activates recombinant neuronal GIRK channels containing the GIRK1 subunit in a manner that requires phosphatidylinositol-4,5-bisphosphate (PIP 2 ), but is otherwise distinct from receptor-induced, G-protein-dependent channel activation. Two amino acids unique to the pore helix (F137) and second membrane-spanning (D173) domain of GIRK1 were identified as necessary and sufficient for the selective activation of GIRK channels by ML297. Further investigation into the behavioral effects of ML297 revealed that in addition to its known antiseizure efficacy, ML297 decreases anxiety-related behavior without sedative or addictive liabilities. Importantly, the anxiolytic effect of ML297 was lost in mice lacking GIRK1. Thus, activation of GIRK1-containing channels by ML297 or derivatives may represent a new approach to the treatment of seizure and/or anxiety disorders.electrophysiology | structure-activity relationship S ignal transduction involving inhibitory (G i/o ) G proteins titrates the excitability of neurons, cardiac myocytes, and endocrine cells, influencing behavior, cardiac output, and energy homeostasis (1). G-protein-gated inwardly rectifying potassium (K + ) (GIRK/Kir3) channels are a common effector for G i/odependent signaling pathways in the heart and nervous system (2, 3). Polymorphisms and mutations in human GIRK channels have been linked to arrhythmias, hyperaldosteronism (and associated hypertension), schizophrenia, sensitivity to analgesics, and alcohol dependence (1).GIRK channels are activated by binding of the G protein Gβγ subunit (1-3). Gβγ binding strengthens channel affinity for phosphatidylinositol-4,5-bisphosphate (PIP 2 ), a necessary cofactor for channel gating (4, 5). GIRK channels are also activated in a G-protein-independent manner by ethanol (6, 7), volatile anesthetics (8, 9), and naringin (10). Many psychoactive and clinically relevant compounds with other primary molecular targets inhibit GIRK channels, albeit at relatively high doses (1, 11). The lack of selective GIRK channel modulators, and in particular, drugs that discriminate among GIRK channel subtypes, has hampered investigation into their physiological relevance and therapeutic potential.GIRK channels are homo-and heterotetramers formed by GIRK1, GIRK2, GIRK3, and GIRK4 subunits (2, 3). GIRK subunits exhibit overlapping but distinct cellular expression patterns, potentially yielding multiple channel subtypes (1). Although it cannot form functional homomers (12), GIRK1 is an integral subunit of the cardiac GIRK channel and most neuronal GIRK channels (13,14). GIRK1 confers robust basal and receptordependent activity to GIRK heteromers, attributable in part to unique residues in the pore and second transmembrane domain (15-17). The intracellular C-terminal domain also contributes to the potentiating influence of GIRK1 on channel activity, likely due to th...

GIRK3 gates activation of the mesolimbic dopaminergic pathway by ethanol

“…17). In particular, ML297, a compound that selectively activates GIRK1-containing heteromers, shows promising antiepileptic and anxiolytic properties while being devoid of sedative and rewarding effects (59,60). Our present results and previous findings (15,16) indicate that a molecule selectively targeting GIRK2/3 heteromers could be effective at blunting the reinforcing effects of ethanol.…”

Selective Ablation of GIRK Channels in Dopamine Neurons Alters Behavioral Effects of Cocaine in Mice

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