A recent study published in Cell Host & Microbe revealed that gut colonization by proteobacteria modulates cocaine neurobehavioral response in mice.
The human microbiota is composed of bacteria of numerous phyla, such as Firmicutes, Proteobacteria, and Bacteroides. Distinct microbiota compositions impact host physiology, homeostasis, metabolic profile, and disease vulnerability. Likewise, host environmental and genetic factors shape the establishment of microbiota enterotypes. This suggests strong bidirectional crosstalk between the microbiome and the host.
Recent studies have demonstrated that some gut microbes can modulate different host behaviors. The altered gut microbiota composition has adverse neurologic implications in humans and animals. However, it remains unknown whether the observed changes in gut microbiota cause, promote, or enhance disease or are consequences of an unrelated pathology.
Similar scenarios are noted in the role of gut microbiota on substance use disorders (SUDs) and addiction. Psychostimulants cause changes in gut microbiota composition, and microbiota depletion affects cocaine responses. However, the underlying molecular mechanisms of these interactions are less explicit.
The study and findings
In the present study, researchers investigated whether and how cocaine affects intestinal microbial colonization in mice. First, mice were infected with Citrobacter rodentium and treated with cocaine or saline, which showed an increased pathogen burden in cecum tissues in mice treated with cocaine. Cocaine treatment also elevated intestinal disease markers in response to infection.
Further investigation indicated that cocaine exposure was the pro-virulence mediator for Citrobacter rodentium by increasing the levels of norepinephrine in cecal tissues, which activated the bacterial adrenergic receptor (QseC) signaling and promoted the subsequent expression of the locus of the enterocyte effacement (LEE) and pathogen colonization.
Next, the researchers assessed if C. rodentium infection altered behavioral plastic changes observed after repeated/multiple exposure(s) to cocaine. The authors noted that infected mice significantly increased locomotor response to repeated cocaine exposure compared to mock-infected mice. This effect was not observed in infected mice with saline administration. Metagenomic analysis revealed significant changes in the gut microbiota of mice infected with C. rodentium.
Infected mice showed a marked increase in γ-proteobacteria, primarily due to C. rodentium, whereas non-infected animals were free from γ-proteobacteria. The authors confirmed that this bloom in γ-proteobacteria driven by the colonization of C. rodentium was responsible for the alterations in cocaine-induced behavioral plasticity rather than the inflammation triggered by the pathogen.
Next, the research team tested if changes in behavioral responses to cocaine were related to metabolite alterations in mice infected with the avirulent C. rodentium escN mutant or Escherichia coli HS. Liquid chromatography-mass spectrometry was performed for untargeted comparative metabolomics of cerebrospinal fluid (CSF) from antibiotic-treated mice reconstituted with C. rodentium escN mutant, E. coli HS, or fecal microbiota transplant (FMT) and treated with cocaine.
Integrated pathway analysis of CSF metabolites between mice reconstituted with FMT and those reconstituted with γ-proteobacteria revealed several affected pathways; the glycine, serine, and threonine metabolism were one of the most significantly altered. Seven of the 11 hits identified in the pathway analysis were downregulated in mice reconstituted with γ-proteobacteria.
Moreover, glycine levels were significantly reduced in the CSF of mice reconstituted with E. coli HS or C. rodentium escN mutant relative to those reconstituted with FMT. Besides, CSF glycine levels were negatively correlated with cocaine-induced behavioral responses. Glycine levels were decreased in the cecal tissues of mice reconstituted with γ-proteobacteria compared with FMT.
Furthermore, genome-wide transcriptional profiling was performed in a brain area related to cocaine-elicited synaptic plasticity, the nucleus accumbens (NAcc). The transcriptomic analysis of NAcc of the FMT- and γ-proteobacteria-reconstituted mice revealed multiple differentially expressed genes (DEGs) after cocaine exposure. There were substantial alterations in pathways involved in glutamatergic synapse and those regulating dopaminergic synapse between FMT- and γ-proteobacteria-reconstituted mice.
Glycine transport uptake is mediated by a permease encoded by the cycA gene. Therefore, the researchers reasoned if cycA deletion may decrease bacteria-induced glycine depletion. Strikingly, the cocaine-induced locomotor behavioral response was no longer detected in mice reconstituted with E. coli HS with a cycA deletion.
Finally, using a conditioned place preference (CPP), the authors demonstrated that antibiotic-treated mice reconstituted with FMT lacked drug-seeking behavior to cocaine. In contrast, significant drug-seeking behavior was observed in mice infected with E. coli HS or C. rodentium escN mutant. Furthermore, mice infected with E. coli HS cycA mutant that cannot uptake glycine did not exhibit significant drug-seeking behavior.
The findings suggest that γ-proteobacteria sense cocaine-induced alterations in host catecholamine levels. In addition, they also indicate that such a microbiota compositional shift toward γ-proteobacteria could deplete host metabolites from the CSF and gut, thereby influencing cocaine-elicited NAcc plasticity and addiction-like behavior. γ-proteobacteria colonization of the gut caused a significant decrease in glycine levels in the blood, CSF, and the gut leading to an exacerbated response to cocaine.