Evidence that GIRK Channels Mediate the DREADD-hM4Di Receptor Activation-Induced Reduction in Membrane Excitability of Striatal Medium Spiny Neurons

Abstract

The hM4Di receptor-based chemogenetic DREADD system has been widely used to suppress neuronal activities, which has contributed substantially to the identification of behavior-associated neuronal circuitries including those in the striatum. One major mechanism by which hM4Di receptor activation suppresses neuronal activity is that the activation reduces membrane excitability, which is thought to be mediated by the opening of GIRK channels. However, previous studies have suggested that GIRK channels are barely expressed in the striatum, which naturally raises the question whether the hM4Di receptor activation-induced reduction in membrane excitability found in striatal medium spiny neurons (MSNs, which constitute 95–98% of the striatal neuronal population) is truly mediated by the endogenous GIRK channels in such scarcity. This study aims to answer this question by applying a GIRK channel-selective blocker, tertiapin-Q (TPNQ), to striatal MSNs. This study first verified that application of clozapine (CZP), an hM4Di receptor agonist, to MSNs expressing the hM4Di receptors hyperpolarized the cell membrane, and reduced membrane excitability and input resistance. This study next revealed that TPNQ post-treatment completely canceled the above CZP-induced electrophysiological effects and that TPNQ pretreatment mostly prevented further expression of the above CZP-induced electrophysiological effects. In addition, confocal microscopy imaging also revealed significant above-background GIRK1 immunofluorescence signals in striatal MSNs. These data suggest that the TPNQ-sensitive GIRK channels, despite being expressed at low levels, are likely the major mediator downstream of hM4Di receptor activation to reduce membrane excitability in striatal MSNs. These results imply that the notion held by scientists in the field that GIRK channels are absent in the striatum or their expression level is not significant enough to exert any function might be oversimplified or incorrect.

Contralesional Anodal Transcranial Direct Current Stimulation Promotes Intact Corticospinal Tract Axonal Sprouting and Functional Recovery After Traumatic Brain Injury in Mice

“…The hM4Di receptor-based chemogenetic designer receptors exclusively activated by designer drugs (DREADD) system has been widely used to suppress neuronal activity. 34 We conducted experiments to verify the feasibility of this method, and the results are shown in Figure 6B. We found that pre-infecting unaffected axons with hM4Di-DREADD did not affect behavior following intraperitoneal injection of saline.…”

Validation of DREADD agonists and administration route in a murine model of sleep enhancement

“…A possible concern raised by this mechanism of action is whether the neurons expressing DREADDs also express sufficient numbers of GIRK channels. To address this concern, a recent study has shown that neurons in brain areas with low GIRK expression are still able to respond to hM4Di activation via GIRK channels to reduce membrane excitability ( Shan et al, 2022 ).…”

Understanding electrical and chemical transmission in the brain

“…In addition, extrasynaptic D2 receptors, such as those in A2A-D2 heteroreceptor complexes, further decrease the firing of striatal-pallidal GABA neurons. The inclusion of GIRK channels in the postsynaptic striatal-pallidal GABA neuron was hypothesized based on experimental evidence in striatal medium spiny neurons (MSNs) (Shan et al, 2022(Shan et al, ). 10.3389/fncel.2024 Frontiers in Cellular Neuroscience 05 frontiersin.org…”