Patricia Robledo

Adolescence is characterized by an increased vulnerability to substance abuse, including alcohol consumption. We investigated the effects of a synbiotic intervention on disruptions of the microbiota-gut-brain axis induced by a drinking in the dark model of intermittent alcohol exposure in adolescent mice. We found that alcohol drinking induced specific shifts in gut microbiota, namely it increased Erysipelotrichaceae and reduced fecal butyric and isovaleric acids. In adulthood, other types of gut bacteria were affected such as Rhodospirillales uncultured family and Entrorhabdus uncultured bacterium. Social and nonsocial cognitive impairments were also observed, and disruptions in prefrontal cortex β-hydroxybutyrate and glutamate metabolic profile in the hippocampus were apparent. Importantly, the synbiotic restored gut microbiota alterations and exerted beneficial effects on alcohol-induced behavioral impairments and brain metabolite changes. In correlational studies, we identified two potential functional networks, one relating gut microbiota (Actinobacteria and Lactobacillaceae)-isovaleric acid with prefrontal glutamate metabolism and sociability, and the other relating SCFAs (propionic, butyric, valeric and isovaleric acids) with β-hydroxybutyrate in the hippocampus and reference memory. These results provide correlative data showing that synbiotic supplementation may restore delayed behavioral alterations induced by voluntary sub-binge alcohol drinking during adolescence through microbiota-gut-brain interactions, and might represent a potential therapeutic tool against long-term alcohol induced behavioral and molecular disturbances.

A reduced sense of smell is a common condition in people with cystic fibrosis (CF) that negatively affects their quality of life. While often attributed to nasal mucosa inflammation, the underlying causes of the olfactory loss remain unknown. Here, we characterized gene expression in olfactory epithelium cells from patients with CF using single-nuclei RNA sequencing and found altered expression of olfactory receptors (ORs) and genes related to progenitor cell proliferation. We confirmed these findings in newborn, inflammation-free samples of a CF animal model and further identified ultrastructural alterations in the olfactory epithelium and bulbs of these animals. We established that CFTR, the anion channel whose dysfunction causes CF, is dispensable for odor-evoked signaling in sensory neurons, yet CF animals displayed defective odor-guided behaviors consistent with the morphological and molecular alterations. Our study highlights CF’s major role in modulating epithelial structure and OR expression, shedding light on the mechanisms contributing to olfactory loss in CF.

G protein-coupled receptors (GPCRs) are sophisticated signaling machines able to simultaneously elicit multiple intracellular signaling pathways upon activation. Complete (in)activation of all pathways can be counterproductive for specific therapeutic applications. This is the case for the serotonin 2 A receptor (5-HT_2AR), a prominent target for the treatment of schizophrenia. In this study, we elucidate the complex 5-HT_2AR coupling signature in response to different signaling probes, and its physiological consequences by combining computational modeling, in vitro and in vivo experiments with human postmortem brain studies. We show how chemical modification of the endogenous agonist serotonin dramatically impacts the G protein coupling profile of the 5-HT_2AR and the associated behavioral responses. Importantly, among these responses, we demonstrate that memory deficits are regulated by G_αq protein activation, whereas psychosis-related behavior is modulated through G_αi1 stimulation. These findings emphasize the complexity of GPCR pharmacology and physiology and open the path to designing improved therapeutics for the treatment of stchizophrenia.

The intricate involvement of the serotonin 5-HT_2A receptor (5-HT_2AR) both in schizophrenia and in the activity of antipsychotic drugs is widely acknowledged. The currently marketed antipsychotic drugs, although effective in managing the symptoms of schizophrenia to a certain extent, are not without their repertoire of serious side effects. There is a need for better therapeutics to treat schizophrenia for which understanding the mechanism of action of the current antipsychotic drugs is imperative. With bioluminescence resonance energy transfer (BRET) assays, we trace the signaling signature of six antipsychotic drugs belonging to three generations at the 5-HT_2AR for the entire spectrum of signaling pathways activated by serotonin (5-HT). The antipsychotic drugs display previously unidentified pathway preference at the level of the individual Gα subunits and β-arrestins. In particular, risperidone, clozapine, olanzapine and haloperidol showed G protein-selective inverse agonist activity. In addition, G protein-selective partial agonism was found for aripiprazole and cariprazine. Pathway-specific apparent dissociation constants determined from functional analyses revealed distinct coupling-modulating capacities of the tested antipsychotics at the different 5-HT-activated pathways. Computational analyses of the pharmacological and structural fingerprints support a mechanistically based clustering that recapitulate the clinical classification (typical/first generation, atypical/second generation, third generation) of the antipsychotic drugs. The study provides a new framework to functionally classify antipsychotics that should represent a useful tool for the identification of better and safer neuropsychiatric drugs and allows formulating hypotheses on the links between specific signaling cascades and in the clinical outcomes of the existing drugs.

Chronic alcohol intake has been shown to disrupt gut microbiota homeostasis, but whether microbiota modulation could prevent behavioral alterations associated with chronic alcohol intake remains unknown. We investigated the effects of synbiotic dietary supplementation on the development of alcohol-related addictive behavior in female and male mice and evaluated whether these effects were associated with changes in bacterial species abundance, short-chain fatty acids, tryptophan metabolism, and neurotransmitter levels in the prefrontal cortex and hippocampus. Chronic intermittent exposure to alcohol during 20 days induced escalation of intake in both female and male mice. Following alcohol deprivation, relapse-like behavior was observed in both sexes, but anxiogenic and cognitive deficits were present only in females. Synbiotic treatment reduced escalation and relapse to alcohol intake in females and males. In addition, the anxiogenic-like state and cognitive deficits observed in females following alcohol deprivation were abolished in mice exposed to synbiotic. Alcohol-induced differential alterations in microbial diversity and abundance in both sexes. In females, synbiotic exposure abrogated the alterations provoked by alcohol in Prevotellaceae UCG-001 and Ruminococcaceae UCG-014 abundance. In males, synbiotic exposure restored the changes induced by alcohol in Akkermansia and Muribaculum uncultured bacterium abundance. Following alcohol withdrawal, tryptophan metabolites, noradrenaline, dopamine, and γ-aminobutyric acid concentrations in the prefrontal cortex and the hippocampus were correlated with bacterial abundance and behavioral alterations in a sex-dependent manner. These results suggested that a dietary intervention with a synbiotic to reduce gut dysbiosis during chronic alcohol intake may impact differently the gut-brain-axis in females and males.

A close epidemiological link has been reported between cannabis use and schizophrenia (SCZ). However, biochemical markers in living humans related to the impact of cannabis in this disease are still missing. Olfactory neuroepithelium (ON) cells express neural features and offer a unique advantage to study biomarkers of psychiatric diseases. The aim of our study was to find exclusively deregulated proteins in ON cells of SCZ patients with and without a history of cannabis use. Thus, we compared the proteomic profiles of SCZ non-cannabis users (SCZ/nc) and SCZ cannabis users (SCZ/c) with control subjects non-cannabis users (C/nc) and control cannabis users (C/c). The results revealed that the main cascades affected in SCZ/nc were cell cycle, DNA replication, signal transduction and protein localization. Conversely, cannabis use in SCZ patients induced specific alterations in metabolism of RNA and metabolism of proteins. The levels of targeted proteins in each population were then correlated with cognitive performance and clinical scores. In SCZ/c, the expression levels of 2 proteins involved in the metabolism of RNA (MTREX and ZNF326) correlated with several cognitive markers and clinical signs. Moreover, use duration of cannabis negatively correlated with ZNF326 expression. These findings indicate that RNA-related proteins might be relevant to understand the influence of cannabis use on SCZ.

Schizophrenia (SCZ) has been associated with serotonergic and endocannabinoid systems dysregulation, but difficulty in obtaining in vivo neurological tissue has limited its exploration. We investigated CB₁R-5-HT_2AR heteromer expression and functionality via intracellular pERK and cAMP quantification in olfactory neuroepithelium (ON) cells of SCZ patients non-cannabis users (SCZ/nc), and evaluated whether cannabis modulated these parameters in patients using cannabis (SCZ/c). Results were compared vs healthy controls non-cannabis users (HC/nc) and healthy controls cannabis users (HC/c). Further, antipsychotic effects on heteromer signaling were tested in vitro in HC/nc and HC/c. Results indicated that heteromer expression was enhanced in both SCZ groups vs HC/nc. Additionally, pooling all 4 groups together, heteromer expression correlated with worse attentional performance and more neurological soft signs (NSS), indicating that these changes may be useful markers for neurocognitive impairment. Remarkably, the previously reported signaling properties of CB₁R-5-HT_2AR heteromers in ON cells were absent, specifically in SCZ/nc treated with clozapine. These findings were mimicked in cells from HC/nc exposed to clozapine, suggesting a major role of this antipsychotic in altering the quaternary structure of the CB₁R-5-HT_2AR heteromer in SCZ/nc patients. In contrast, cells from SCZ/c showed enhanced heteromer functionality similar to HC/c. Our data highlight a molecular marker of the interaction between antipsychotic medication and cannabis use in SCZ with relevance for future studies evaluating its association with specific neuropsychiatric alterations.

Cannabis is the third most commonly used psychoactive substance of abuse, yet it also receives considerable attention as a potential therapeutic drug. Therefore, it is essential to fully understand the actions of cannabis in the human brain. The olfactory neuroepithelium (ON) is a peripheral nervous tissue that represents an interesting surrogate model to study the effects of drugs in the brain, since it is closely related to the central nervous system, and sensory olfactory neurons are continually regenerated from populations of stem/progenitor cells that undergo neurogenesis throughout life. In this study, we used ON cells from chronic cannabis users and healthy control subjects to assess alterations in relevant cellular processes, and to identify changes in functional proteomic pathways due to cannabis consumption. The ON cells from cannabis users exhibited alterations in the expression of proteins that were related to the cytoskeleton, cell proliferation and cell death, as well as, changes in proteins implicated in cancer, gastrointestinal and neurodevelopmental pathologies. Subsequent studies showed cannabis provoked an increase in cell size and morphological alterations evident through β-Tubulin III staining, as well as, enhanced beta-actin expression and a decrease in the ability of ON cells to undergo cell attachment, suggesting abnormalities of the cytoskeleton and cell adhesion system. Furthermore, these cells proliferated more and underwent less cell death. Our results indicate that cannabis may alter key processes of the developing brain, some of which are similar to those reported in mental disorders like DiGeorge syndrome, schizophrenia and bipolar disorder.