Toll, L. | Search bioRxiv

bioRxiv2026

Whole Brain Cellular Activation After Mu and NOP Receptor Agonism Identifies Differential Regional and Network Consequences

Mu opioid agonists, the most widely used opioids, produce analgesia, respiratory depression, constipation, and reward leading to substance abuse. Like all opioid receptor family members, mu receptors are Gi/o-coupled and inhibitory, yet mu-mediated disinhibition of GABAergic neurons propagates downstream activation that differs between naive and dependent animals and across receptor subtypes. Here we used TRAP2/Ai9 reporter mice, which deposit tdTomato in active neurons via the c-Fos locus, with tissue clearing and light sheet microscopy to map whole-brain neuronal activation after acute morphine (10 mg/kg), an escalating twice-daily morphine regimen producing dependence, or the NOP receptor agonist Ro 64-6198. Despite Gi/o coupling, all treatments raised global activation relative to vehicle, though this whole-brain increase reached significance only in males, and acute morphine engaged the largest number of regions, all increased with none decreased. Sex differences were prominent: males were more sensitive than females across treatments, with morphine engaging mainly pain- and reward-related midbrain circuitry in males and hypothalamic circuitry in females. In dependent mice, activation was lower across several regions, including a cluster of cerebellar regions that trended toward being turned off relative to acute morphine. NOP receptor activation produced a far weaker profile. Although chronic morphine produced less activation than acute morphine, it produced more widespread co-activation, and while hub composition differed across conditions, global network organization was preserved. These results reveal whole-brain changes in neuronal activation and establish a TRAP2/Ai9 model that renders activated regions genetically accessible for future site-directed investigation.

neuroscience↗

bioRxiv2023

Rare GPR37L1 variants reveal potential roles in anxiety and migraine disorders

GPR37L1 is an orphan receptor that couples through heterotrimeric G-proteins to regulate physiological functions. Since its role in humans is not fully defined, we used an unbiased computational approach to assess the clinical significance of rare GPR37L1 genetic variants found among 51,289 whole exome sequences from the DiscovEHR cohort. Briefly, rare GPR37L1 coding variants were binned according to predicted pathogenicity, and analyzed by Sequence Kernel Association testing to reveal significant associations with disease diagnostic codes for epilepsy and migraine, among others. Since associations do not prove causality, rare GPR37L1 variants were then functionally analyzed in SK-N-MC cells to evaluate potential signaling differences and pathogenicity. Notably, receptor variants exhibited varying abilities to reduce cAMP levels, activate MAPK signaling, and/or upregulate receptor expression in response to the agonist prosaptide (TX14(A)), as compared to the wild-type receptor. In addition to signaling changes, knockout of GPR37L1 or expression of certain rare variants altered cellular cholesterol levels, which were also acutely regulated by administration of the agonist TX14(A) via activation of the MAPK pathway. Finally, to simulate the impact of rare nonsense variants found in the large patient cohort, a knockout (KO) mouse line lacking Gpr37L1 was generated, revealing loss of this receptor produced sex-specific changes implicated in migraine-related disorders. Collectively, these observations define the existence of rare GPR37L1 variants in the human population that are associated with neuropsychiatric conditions and identify the underlying signaling changes that are implicated in the in vivo actions of this receptor in pathological processes leading to anxiety and migraine. SIGNIFICANCE STATEMENTG-protein coupled receptors (GPCRs) represent a diverse group of membrane receptors that contribute to a wide range of diseases and serve as effective drug targets. However, a number of these receptors have no identified ligands or functions, i.e., orphan receptors. Over the past decade, advances have been made, but there is a need for identifying new strategies to reveal their roles in health and disease. Our results highlight the utility of rare variant analyses of orphan receptors for identifying human disease associations, coupled with functional analyses in relevant cellular and animal systems, to ultimately reveal their roles as novel drug targets for treatment of neurological disorders that lack wide-spread efficacy.

neuroscience↗