Dominant role of GABAB2 and Gbetagamma for GABAB receptor-mediated-ERK1/2/CREB pathway in cerebellar neurons

gamma-aminobutyric acid type B (GABA(B)) receptor is an allosteric complex made of two subunits, GABA(B1) and GABA(B2). GABA(B2) plays a major role in the coupling to G protein whereas GABA(B1) binds GABA. It has been shown that GABA(B) receptor activates ERK(1/2) in neurons of the central nervous system, but the molecular mechanisms underlying this event are poorly characterized. Here, we demonstrate that activation of GABA(B) receptor by either GABA or the selective agonist baclofen induces ERK(1/2) phosphorylation in cultured cerebellar granule neurons. We also show that CGP7930, a positive allosteric regulator specific of GABA(B2), alone can induce the phosphorylation of ERK(1/2). PTX, a G(i/o) inhibitor, abolishes both baclofen and CGP7930-mediated-ERK(1/2) phosphorylation. Moreover, both baclofen and CGP7930 induce ERK-dependent CREB phosphorylation. Furthermore, by using LY294002, a PI-3 kinase inhibitor, and a C-term of GRK-2 that has been reported to sequester Gbetagamma subunits, we demonstrate the role of Gbetagamma in GABA(B) receptor-mediated-ERK(1/2) phosphorylation. In conclusion, the activation of GABA(B) receptor leads to ERK(1/2) phosphorylation via the coupling of GABA(B2) to G(i/o) and by releasing Gbetagamma subunits which in turn induce the activation of CREB. These findings suggest a role of GABA(B) receptor in long-term change in the central nervous system.

💡 Research Summary

This study investigates the intracellular signaling cascade by which the GABA‑B receptor activates the MAPK/ERK pathway and downstream transcription factor CREB in cerebellar granule neurons (CGNs). The GABA‑B receptor is a heterodimer composed of GABA‑B1, which binds the neurotransmitter, and GABA‑B2, which couples the complex to heterotrimeric G proteins. While previous work has shown that GABA‑B activation leads to ERK1/2 phosphorylation, the precise molecular steps remained unclear.

Using primary CGN cultures, the authors first demonstrated that both the endogenous ligand GABA and the selective orthosteric agonist baclofen trigger rapid (5–15 min) phosphorylation of ERK1/2. Importantly, the positive allosteric modulator (PAM) CGP7930, which binds exclusively to the GABA‑B2 subunit, also induced ERK1/2 activation when applied alone, indicating that GABA‑B2 can initiate signaling without ligand‑induced conformational changes in GABA‑B1.

To test the involvement of Gi/o proteins, cultures were pre‑treated with pertussis toxin (PTX), a covalent inhibitor of Gi/o α‑subunits. PTX completely abolished ERK1/2 phosphorylation induced by both baclofen and CGP7930, confirming that Gi/o coupling is essential for the observed MAPK activation.

The downstream relevance of ERK activation was examined by probing CREB phosphorylation. Both baclofen and CGP7930 increased phospho‑CREB levels in an ERK‑dependent manner, linking the GABA‑B‑mediated MAPK cascade to transcriptional regulation.

The role of the Gβγ dimer was dissected using two complementary approaches. First, the PI3‑kinase inhibitor LY294002 reduced baclofen‑ and CGP7930‑induced ERK1/2 phosphorylation, suggesting that Gβγ signals through a PI3K‑Akt axis to activate ERK. Second, overexpression of the C‑terminal fragment of GRK‑2 (βARKct), which sequesters free Gβγ, markedly suppressed ERK activation by both agonist and PAM. These findings demonstrate that liberated Gβγ subunits are required for the full ERK response.

Collectively, the data support a signaling model in which activation of the GABA‑B receptor leads to Gi/o activation via the GABA‑B2 subunit, release of Gβγ dimers, Gβγ‑mediated stimulation of PI3K, and subsequent activation of the ERK1/2 cascade, culminating in CREB phosphorylation. This pathway provides a mechanistic link between GABA‑B receptor activation and long‑term neuronal plasticity.

The authors discuss the broader implications of this mechanism. Because the Gβγ‑PI3K‑ERK‑CREB axis can influence gene expression programs related to neuronal survival, growth, and synaptic remodeling, it may underlie the role of GABA‑B receptors in learning, memory, and adaptive responses to chronic stimuli. Moreover, dysregulation of this cascade could contribute to neuropsychiatric and neurodegenerative conditions where GABA‑B signaling is implicated, such as alcohol dependence, depression, and Huntington’s disease. Targeting specific nodes—Gβγ sequestration, PI3K activity, or downstream ERK/CREB modulation—could therefore represent novel therapeutic strategies.

In summary, this paper provides compelling evidence that GABA‑B2 and the released Gβγ subunits are the dominant drivers of GABA‑B‑induced ERK1/2 activation in cerebellar neurons, and that this signaling culminates in CREB phosphorylation, offering a molecular framework for the long‑term effects of GABA‑B receptor activation in the central nervous system.