Parkinson's disease (PD) is a progressive neurological disorder that affects movement, causing symptoms such as tremors, stiffness, slowness, and balance problems due to the degeneration of dopamine-producing neurons in the brain. Nowadays there is no cure for PD. Alpha synuclein (α-syn) aggregates, which are a hallmark of PD, are known to induce microglial activation, specifically the detrimental M1 microglial phenotype, which contributes to neuroinflammation and disease progression. Cannabinoid receptor 2 (CB2R) activation has been shown to counteract neuroinflammation. CB2R is able to interact with N-methyl-D-aspartate (NMDA) receptors (NMDAR), which has also attracted attention in PD research due to its role in excitotoxicity. Here we aimed to study the interaction between CB2R and NMDAR in a PD context in rat tissue. We observed that α-syn fibrils alter CB2R activation and CB2R-NMDAR heteromerization in a heterologous expression system. Furthermore, activation of CB2R counteracted NMDAR signaling. In microglia, α-syn fibrils decreased CB2R-NMDAR heteromer expression while increasing CB2R signaling. Importantly, CB2R activation counteracted the α-syn fibrils-induced increase in M1-activated microglia, while it favored the polarization of microglia to the beneficial M2 phenotype. These results reinforce the idea of using cannabinoids for treating PD, as they provide not only the anti-inflammatory effects of cannabinoids but also counteract the detrimental increase in NMDAR signaling present in this disease.

G protein-coupled receptors (GPCRs) are integral membrane proteins which closely interact with their plasma membrane lipid microenvironment. Cholesterol is a lipid enriched at the plasma membrane with pivotal roles in the control of membrane fluidity and maintenance of membrane microarchitecture, directly impacting on GPCR stability, dynamics, and function. Cholesterol extraction from pancreatic beta cells has previously been shown to disrupt the internalisation, clustering, and cAMP responses of the glucagon-like peptide-1 receptor (GLP-1R), a class B1 GPCR with key roles in the control of blood glucose levels via the potentiation of insulin secretion in beta cells and weight reduction via the modulation of brain appetite control centres. Here, we unveil the detrimental effect of a high cholesterol diet on GLP-1R-dependent glucoregulation in vivo, and the improvement in GLP-1R function that a reduction in cholesterol synthesis using simvastatin exerts in pancreatic islets. We next identify and map sites of cholesterol high occupancy and residence time on active vs inactive GLP-1Rs using coarse-grained molecular dynamics (cgMD) simulations, followed by a screen of key residues selected from these sites and detailed analyses of the effects of mutating one of these, Val229, to alanine on GLP-1R-cholesterol interactions, plasma membrane behaviours, clustering, trafficking and signalling in INS-1 832/3 rat pancreatic beta cells and primary mouse islets, unveiling an improved insulin secretion profile for the V229A mutant receptor. This study (1) highlights the role of cholesterol in regulating GLP-1R responses in vivo; (2) provides a detailed map of GLP-1R - cholesterol binding sites in model membranes; (3) validates their functional relevance in beta cells; and (4) highlights their potential as locations for the rational design of novel allosteric modulators with the capacity to fine-tune GLP-1R responses.
© 2024, Oqua et al.

Parkinson’s disease (PD) is a progressive neurological disorder that affects movement, causing symptoms such as tremors, stiffness, slowness, and balance problems due to the degeneration of dopamine-producing neurons in the brain. Nowadays there is no cure for PD. Alpha synuclein (α-syn) aggregates, which are a hallmark of PD, are known to induce microglial activation, specifically the detrimental M1 microglial phenotype, which contributes to neuroinflammation and disease progression. Cannabinoid receptor 2 (CB 2 R) activation has been shown to counteract neuroinflammation. CB2R is able to interact with NMDA receptors (NMDAR), which has also attracted attention in PD research due to its role in excitotoxicity. Here we aimed to study the interaction between CB 2 R and NMDAR in a PD context. We observed that α-syn fibrils alter CB2R activation and CB 2 R-NMDAR heteromerization in a heterologous expression system. Furthermore, activation of CB2R counteracted NMDAR signaling. In neurons α-syn fibrils decreased CB2R-NMDAR heteromer expression, while increasing CB 2 R signaling. Importantly, CB 2 R activation counteracted the α-syn fibrils-induced increase in M1 activated microglia, while it favored the polarization of microglia to the beneficial M2 phenotype. These results reinforce the idea of using cannabinoids for treating PD, as they provide not only the anti-inflammatory effects of cannabinoids but also counteract the detrimental increase in NMDAR signaling present in this disease.

The class B1 G protein-coupled receptor (GPCR) subfamily is a class of receptors known for their regulatory roles in metabolism and neuronal activity important as drug targets. Lipids play key functional roles in modulation of GPCR signalling, yet our understanding of the molecular level detail of specific lipid interactions with class B1 GPCRs remains limited. Here we present coarse-grained molecular dynamics (MD) simulations of the active and inactive states of 15 human class B1 family members and use aiida-gromacs to capture full provenance for the set-up of simulations in complex plasma membranes. Receptors exhibit state-dependent lipid interactions with the regulatory sterol cholesterol and phospholipid phosphatidylinositiol-4,5-bisphosphate [PI(4,5)P 2 ] at defined locations on the receptor surface. Global analysis of trends across the subfamily reveals conserved patterns of lipid interaction dynamics. The glycosphingolipid GM3 exerts a modulatory influence on the dynamics of class B1 extracellular domains in both simulations and in vitro time-resolved FRET assays. Teaser Characterisation of regulatory lipid interactions with all class B1 GPCRs.

The μ-opioid receptor (μOR), a prototypical G protein-coupled receptor (GPCR), is the target of opioid analgesics such as morphine and fentanyl. Due to the severe side effects of current opioid drugs, there is considerable interest in developing novel modulators of μOR function. Most GPCR ligands today are small molecules, however biologics, including antibodies and nanobodies, represent alternative therapeutics with clear advantages such as affinity and target selectivity. Here, we describe the nanobody NbE, which selectively binds to the μOR and acts as an antagonist. We functionally characterize NbE as an extracellular and genetically encoded μOR ligand and uncover the molecular basis for μOR antagonism by determining the cryo-EM structure of the NbE-μOR complex. NbE displays a unique ligand binding mode and achieves μOR selectivity by interactions with the orthosteric pocket and extracellular receptor loops. Based on a β-hairpin loop formed by NbE that deeply protrudes into the μOR, we design linear and cyclic peptide analogs that recapitulate NbE's antagonism. The work illustrates the potential of nanobodies to uniquely engage with GPCRs and describes lower molecular weight μOR ligands that can serve as a basis for therapeutic developments.
© 2024. The Author(s).