Disabling the Gβγ-SNARE interaction disrupts GPCR-mediated presynaptic inhibition, leading to physiological and behavioral phenotypes

Abstract

G protein–coupled receptors (GPCRs) that couple to Gi/o proteins modulate neurotransmission presynaptically by inhibiting exocytosis. Release of Gβγ subunits from activated G proteins decreases the activity of voltage-gated Ca2+ channels (VGCCs), decreasing excitability. A less understood Gβγ-mediated mechanism downstream of Ca2+ entry is the binding of Gβγ to SNARE complexes, which facilitate the fusion of vesicles with the cell plasma membrane in exocytosis. Here, we generated mice expressing a form of the SNARE protein SNAP25 with premature truncation of the C terminus and that were therefore partially deficient in this interaction. SNAP25Δ3 homozygote mice exhibited normal presynaptic inhibition by GABAB receptors, which inhibit VGCCs, but defective presynaptic inhibition by receptors that work directly on the SNARE complex, such as 5-hydroxytryptamine (serotonin) 5-HT1b receptors and adrenergic α2a receptors. Simultaneously stimulating receptors that act through both mechanisms showed synergistic inhibitory effects. SNAP25Δ3 homozygote mice had various behavioral phenotypes, including increased stress-induced hyperthermia, defective spatial learning, impaired gait, and supraspinal nociception. These data suggest that the inhibition of exocytosis by Gi/o-coupled GPCRs through the Gβγ-SNARE interaction is a crucial component of numerous physiological and behavioral processes.

Removing the Gβγ-SNAP25 brake on exocytosis enhances insulin action, promotes adipocyte browning, and protects against diet-induced obesity

“…Our prior studies suggested that SNAP25 Δ3/Δ3 had an altered behavioral response to acute physiological stressors (5,6). We performed calorimetry studies where we assessed energy expenditure and feeding behavior in chow fed male SNAP25 +/+ and SNAP25 Δ3/Δ3 mice at standard housing temperature (22C).…”

“…In our prior work we observed that SNAP25 Δ3/Δ3 have behavioral deficits that impact learning, memory, and the response to stressors (5,6). We asked whether similar deficits could impact energy homeostasis.…”

“…Mice used for these experiments were generated from heterozygous breeding of SNAP25 +/Δ3 mice on a C57BL/6 background. The strategy for the generation of these mice was previously described (5). Mice were maintained ad libitum on chow, Laboratory Rodent Diet 5001 (LabDiet), unless otherwise stated.…”

Developmental Time Course of SNAP-25 Isoforms Regulate Hippocampal Long-Term Synaptic Plasticity and Hippocampus-Dependent Learning

“…We have previously shown that mutations in the C-terminus of SNAP-25 in a gene targeted mouse model enhances the magnitude of LTP at Schaffer collateral-CA1 synapses [42]. This is because of the reduced interaction between SNAP-25 and the inhibitory G i/o proteins [43]. The absence of SNAP-25b did not alter the amplitude of mGluRII-dependent LTD measured at Schaffer collateral-CA1 synapses when DCG-IV was bath applied, or NMDAR-dependent LTD when NMDA was bath applied to hippocampal slices from such mice compared to four month old littermates.…”

Specificity of Gβ and γ subunits to the SNARE complex both at rest and after α2aadrenergic receptor stimulation

“…Previously, we identified G binding sites on both amino-and carboxy-terminal regions of SNAP25 (41,56,57), far enough apart to not both be occupied by a single G. Using SNAP25∆3 mice with a truncated c-terminus SNAP25, we see a two-fold reduction in G-SNARE interaction, as well as an increased in stress-induced hyperthermia, defective spatial learning, impaired gait, supraspinal nociception (58), and enhancement of long-term potentiation at Schaffer collateral-CA1 synapse (59). However, the physiological and biochemical importance of these interaction sites still remains unclear.…”