Biology subjects

Basbaum, A.

Publications and source records attributed to Basbaum, A..

9 recordsLinked to original sources

bioRxiv2025

Structure based discovery of antipsychotic-like TAAR1 agonists

Schizophrenia is a severe mental illness whose current treatments primarily target dopamine and serotonin receptors. These drugs often cause side effects and vary in effectiveness across patients. The trace amine-associated receptor 1 (TAAR1), which modulates monoamine signaling, has emerged as a promising alternative target. To discover new TAAR1 ligands, we computationally docked 65 million molecules against the active state of TAAR1 and experimentally tested 55 of those highly ranked. Fourteen molecules active against TAAR1 with potencies ranging from mid-nanomolar to micromolar, all as agonists. This high functional selectivity may reflect the compact conformation adopted by the activated TAAR1 orthosteric site. While this was favorable for agonist prioritization, simulations suggest that it can be over-optimized for initial hit rates at the expense of subsequent affinity maturation. Here, hit optimization yielded nanomolar agonists whose docking-predicted poses were confirmed by cryo-EM. Three agonists had high brain exposure and potencies apparently better than the investigational drug ulotaront and sufficient for behavioral studies. All three potently normalized amphetamine-induced pre-pulse inhibition in mice, a model for schizophrenia, without catalepsy, a common side effect of traditional antipsychotics.

biochemistry↗

bioRxiv2025

Photothrombotic Ischemic Thalamic Stroke in Mice Recapitulates Spontaneous Pain Features of Central Post-Stroke Pain in Humans

Central post-stroke pain (CPSP) is a highly distressing condition that develops in 50% of people who suffer a thalamic stroke, and is typically unresponsive to current clinical treatments. Hypoxic damage to the ventral posterolateral (VPL) and ventral posteromedial (VPM) sensory thalamic nuclei, in particular, precipitates CPSP. One barrier to developing treatments for CPSP is the lack of preclinical models of thalamic ischemic stroke. In this study, we present a novel mouse model of CPSP induced through targeted photothrombotic ischemia. After eliciting hypoxia in the sensory thalamus of male mice, we assessed pain behaviors over a four-week period. Stroke-affected mice exhibited a persistent spontaneous facial grimace from day four to week four post-stroke, indicative of pain. Hind-paw mechanical hypersensitivity indicative of altered nociception, characteristic of VPL and VPM hemorrhagic CPSP models, was not detected in our model. Immunofluorescence analysis revealed increased activated microglia (Iba1) and reactive astrocytes (GFAP). Iba1 fluorescence intensity in the VPL thalamus-but not the VPM thalamus-correlated with the severity of facial grimace at four weeks post-stroke. Clustering based on behavioral phenotypes identified a subpopulation of mice in which grimace pain spontaneously resolved, by four weeks post-stroke, relative to sham controls, suggesting that this model can be used to understand how stroke recovery may influence pain chronification. This model provides a valuable tool to investigate the cellular and circuit mechanisms underlying CPSP after an ischemic thalamic stroke. Significance StatementResearch into central post-stroke pain (CPSP) is hindered by the lack of preclinical models that match the clinical presentation of the condition. Although 87% of clinical strokes are ischemic, current CPSP preclinical models induce hemorrhagic strokes. To our knowledge, our mouse model is the first to induce CPSP by photothrombosis in the ventral posterolateral and posteromedial thalamus. Moreover, our study quantifies spontaneous pain, using a facial grimace score, in addition to nociception, whereas previous CPSP studies have relied exclusively on nociceptive tests. Finally, our model recapitulates some of the variability observed in clinical CPSP. We anticipate that this model will help dissect the roots of chronic pain after a thalamic stroke, and facilitate development of novel therapies for CPSP.

neuroscience↗

bioRxiv2023

Structure-Based Discovery of a NPFF1R Antagonist with Analgesic Activity

While opioid drugs remain among the most effective analgesics for pain management, adverse effects limit their use. Molecules that synergize with opioids, increasing analgesia without increasing side effects, could prove beneficial. A potential way to do so is via the RF-amide receptor system, as NPFFR1 agonists reduce {micro}- opioid receptor ({micro}OR)-based analgesia while antagonists increase it. These inferences are, however, clouded by the lack of selectivity of most NPFF1R ligands. Seeking selective antagonists of the NPFF1R, we screened a large virtual library against a homology model of NPFF1R. From 26 high-ranking molecules that were synthesized and tested, one antagonized NPFF1R with a Ki of 319 nM. Structure-based optimization led to a 22 nM antagonist of NPFF1R, compound 56, with selectivity against a large panel of GPCRs. When administered alone, 56 has no activity in mouse tail-flick nociception assays. However, coadministration of compound 56 and morphine produced significantly greater antinociception than did morphine alone, consistent with the notion that NPFF1R nociceptive activity occurs via modulation of {micro}OR signaling. Surprisingly, in the hot-plate assays 56 was analgesic by itself, suggesting that NPFF1R alone can also confer analgesia. At equi-analgesic doses, combinations of 56 with morphine reduced the common constipation side effect of morphine versus using morphine alone. The high selectivity of 56 and its activity in cooperation with morphine supports further analgesic development against NPFF1R and against the RF-amide family of receptors more generally.

pharmacology and toxicology↗

bioRxiv2023

Discovery of a functionally selective serotonin 5-HT1A receptor agonist for the treatment of pain

The G protein-coupled serotonin receptor 5-HT1AR mediates antinociception and may serve as a valuable target for the treatment of pain. Starting from a chemical library, ST171, a bitopic chemotype activating 5-HT1AR was evolved. In vitro pharmacological investigations of ST171 revealed potent and selective Gi activation (EC50 = 0.3 nM), with marginal Gs and {beta}-arrestin recruitment. Preclinical studies in mice showed that ST171 was effective in acute and chronic (inflammatory and neuropathic) pain models, without causing sedation. Comparison of cryo-EM structures of a 5-HT1AR-Gi complex bound to the functionally biased agonist ST171, with a structure bound to the functionally balanced agonist befiradol, showed that both ligands bind to the same orthosteric site, but address different exo-sites. The individual poses are associated with ligand-specific helical dispositions and rearrangements of microdomains. Complementation of these studies with molecular dynamics simulations allowed us to derive structural features associated with ST171s functional selectivity, a phenomenon that may be crucial to the discovery of more effective and safe GPCR drugs.

pharmacology and toxicology↗

bioRxiv2023

Tetracycline transactivator overexpression in keratinocytes triggers a TRPV1 primary sensory neuron-dependent neuropathic itch

Mouse models that combine tetracycline-controlled gene expression systems and conditional genetic activation can tightly regulate transgene expression in discrete cell types and tissues. However, the commonly used Tet-Off variant, tetracycline transactivator (tTA), when overexpressed and fully active, can lead to developmental lethality, disease, or more subtle behavioral phenotypes. Here we describe a profound itch phenotype in mice expressing a genetically encoded tTA that is conditionally activated within the Phox2a lineage. Phox2a; tTA mice develop intense, localized scratching and regional skin lesions that can be controlled by the tTA inhibitor, doxycycline. As gabapentin, but not morphine, relieved the scratching, we consider this phenotype to result from chronic neuropathic itch, not pain. In contrast to the Phox2a lineage, mice with tTA activated within the Phox2b lineage, which has many similar areas of recombination within the nervous system, did not recapitulate the scratching phenotype. In Phox2a-Cre mice, but not Phox2b-Cre, intense Cre-dependent reporter expression was found in skin keratinocytes which formed the area at which skin lesions developed. Most interestingly, repeated topical application of the DREADD agonist, CNO, which chronically induced Gi signaling in Phox2a-keratinocytes, completely reversed the localized scratching and skin lesions. Furthermore, ablation of TRPV1-expressing, primary afferent neurons reduced the scratching with a time course comparable to that produced by Gi-DREADD inhibition. These temporal properties suggest that the neuropathic itch condition arises not only from localized keratinocyte activation of peripheral nerves but also from a persistent, gabapentin-sensitive state of central sensitization.

neuroscience↗

bioRxiv2023

Long-term optical imaging of the spinal cord in awake, behaving animals

Advances in optical imaging approaches and fluorescent biosensors have enabled an understanding of the spatiotemporal and long-term neural dynamics in the brain of awake animals. However, methodological difficulties and the persistence of post-laminectomy fibrosis have greatly limited similar advances in the spinal cord. To overcome these technical obstacles, we combined in vivo application of fluoropolymer membranes that inhibit fibrosis; a redesigned, cost-effective implantable spinal imaging chamber; and improved motion correction methods that together permit imaging of the spinal cord in awake, behaving mice, for months to over a year. We also demonstrate a robust ability to monitor axons, identify a spinal cord somatotopic map, conduct Ca2+ imaging of neural dynamics in behaving animals responding to pain-provoking stimuli, and observe persistent microglial changes after nerve injury. The ability to couple neural activity and behavior at the spinal cord level will drive insights not previously possible at a key location for somatosensory transmission to the brain.

neuroscience↗

bioRxiv2022

Structure-based Discovery of Conformationally Selective Inhibitors of the Serotonin Transporter

The serotonin transporter (SERT) removes synaptic serotonin and is the target of anti-depressant drugs. SERT adopts three conformations: outward-open, occluded, and inward-open. All known inhibitors target the outward-open state except ibogaine, which has an unusual anti-depressant profile and stabilizes the inward-open conformation. Unfortunately, ibogaine is promiscuous and cardiotoxic, limiting understanding of inward-open state ligands. We computationally docked over 200 million small molecules against the ibogaine stabilized inward-open state of SERT. Thirty-six top-ranking compounds were synthesized and thirteen inhibited with potencies ranging from 29 to 5000 nM. Structure-based optimization led to two novel inhibitors with Ki values down to 3 nM. The new molecules stabilized an outward-closed state of the transporter and had little activity against off-targets. A cryo-EM structure of one of these bound to SERT confirmed the predicted geometry. In mouse behavioral assays, both had anxiolytic and anti-depressant activity, with potencies up to 200 better than fluoxetine.

biochemistry↗

bioRxiv2021

Structure-based design of a chemical probe set for the 5-HT5A serotonin receptor

The 5-HT5A receptor (5-HT5AR), for which no selective agonists and only a few antagonists exist, remains the least understood serotonin (5-HT) receptor. A single commercial antagonist (SB-699551) has been widely used to investigate central nervous system (CNS) 5-HT5AR function in neurological disorders, including pain. However, because SB-699551 has affinity for many 5-HTRs, lacks inactive property-matched controls, and has assay interference concerns, it has liabilities as a chemical probe. To better illuminate 5-HT5AR function, we developed a probe set through iterative rounds of molecular docking, pharmacological testing, and optimization. Docking over six million lead-like molecules against a 5-HT5AR homology model identified five mid-M ligand starting points with unique scaffolds. Over multiple rounds of structure-based design and testing, a new quinoline scaffold with high affinity and enhanced selectivity for the 5-HT5AR was developed, leading to UCSF678, a 42 nM arrestin-biased partial agonist at the 5-HT5AR with a much more restricted off-target profile and decreased assay liabilities vs. SB-699551. Site-directed mutagenesis supported the docked pose of UCSF678, which was also consistent with recent published 5-HTR structures. Surprisingly, property-matched analogs of UCSF678 that were either inactive across 5-HTRs or retained affinity for UCSF678s off-targets revealed that 5-HT5AR engagement is nonessential for alleviating pain in a mouse model, contrary to previous studies using less-selective ligands. Relative to SB-699551, these molecules constitute a well-characterized and more selective probe set with which to study the function of the 5-HT5A receptor. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=113 SRC="FIGDIR/small/471449v1_ufig1.gif" ALT="Figure 1"> View larger version (28K): org.highwire.dtl.DTLVardef@8fb24dorg.highwire.dtl.DTLVardef@2dd84org.highwire.dtl.DTLVardef@1d3346org.highwire.dtl.DTLVardef@1117331_HPS_FORMAT_FIGEXP M_FIG Table of Contents Graphic C_FIG

biophysics↗

bioRxiv2019

Convergent neural representations of acute nociceptive pain in healthy volunteers: A large-scale fMRI meta-analysis

Characterizing a reliable, pain-related neural signature is critical for translational applications. Many prior fMRI studies have examined acute pain-related brain activation in healthy participants. However, synthesizing these data to identify convergent patterns of activation can be challenging due to the heterogeneity of experimental designs and samples. To address this challenge, we conducted a comprehensive meta-analysis of fMRI studies of stimulus-induced pain in healthy participants. Following pre-registration, two independent reviewers evaluated 4,927 abstracts returned from a search of 8 databases, with 222 fMRI experiments meeting inclusion criteria. We analyzed these experiments using Activation Likelihood Estimation with rigorous type I error control (voxel height p < 0.001, cluster p < 0.05 FWE-corrected) and found a convergent, largely bilateral pattern of pain-related activation in the secondary somatosensory cortex, insula, midcingulate cortex, and thalamus. Notably, these regions were consistently recruited regardless of stimulation technique, location of induction, and participant sex. These findings suggest a highly-conserved core set of pain-related brain areas, encouraging applications as a biomarker for novel therapeutics targeting acute pain.\n\nHIGHLIGHTSO_LIPain stimulation recruits a core set of pain-related brain regions.\nC_LIO_LIThis core set includes thalamus, SII, insula and mid-cingulate cortex.\nC_LIO_LIThese regions were recruited regardless of stimulus modality and stimulus location.\nC_LI

neuroscience↗