Heterogeneity of the humoral immune response following Staphylococcus aureus bacteremia

Expanding knowledge on the humoral immune response in Staphylococcus aureus-infected patients is a mandatory step in the development of vaccines and immunotherapies. Here, we present novel insights into the antibody responses following S. aureus bacteremia. Fifteen bacteremic patients were followed extensively from diagnosis onwards (median 29 days, range 9-74). S. aureus strains (median 3, range 1-6) and serial serum samples (median 16, range 6-27) were collected. Strains were genotyped by pulsed-field gel electrophoresis (PFGE) and genes encoding 19 staphylococcal proteins were detected by polymerase chain reaction (PCR). The levels of IgG, IgA, and IgM directed to these proteins were determined using bead-based flow cytometry. All strains isolated from individual patients were PFGE-identical. The genes encoding clumping factor (Clf) A, ClfB, and iron-responsive surface-determinant (Isd) A were detected in all isolates. Antigen-specific IgG levels increased more frequently than IgA or IgM levels. In individual patients, different proteins induced an immune response and the dynamics clearly differed. Anti-ClfB, anti-IsdH, and anti-fibronectin-binding protein A IgG levels increased in 7 of 13 adult patients (p < 0.05). The anti-IsdA IgG level increased in 12 patients (initial to peak level: 1.13-10.72 fold; p < 0.01). Peak level was reached 7-37 days after diagnosis. In a bacteremic 5-day-old newborn, antistaphylococcal IgG levels declined from diagnosis onwards. In conclusion, each bacteremic patient develops a unique immune response directed to different staphylococcal proteins. Therefore, vaccines should be based on multiple components. IsdA is immunogenic and, therefore, produced in nearly all bacteremic patients.

Probing the role of dopamine at the intersection of addiction, decision making, and sex

Addiction is an escalating, compulsive, and relapsing psychiatric disease that affects millions worldwide and exacts immense socioeconomic cost. As for potential pharmacotherapeutic targets, the dopamine system is of most interest, but it is unclear as to whether increasing or decreasing dopamine is the best approach. There is consensus that the cues associated with cocaine use and gambling (e.g., a crack pipe or flashing casino lights) are critical in driving disordered behaviour. It is also clear that the responsivity to drug and gambling cues is governed by dopamine and that there is considerable comorbidity between cocaine use and gambling disorder. Biological sex also plays a critical role in addiction but is confounded by the socioeconomic construct of gender in human studies, necessitating animal models. In the following thesis, we preclinically modeled the complex intersection of cocaine use and gambling-like behaviour by combining operant cocaine self-administration with the cued rat gambling task. While female and male rats gambled and took cocaine, we used chemogenetics to bidirectionally modulate the dopaminergic neurons projecting from the ventral tegmental area. We showed that chemogenetic inhibition of dopamine neurons decreased numerous addiction-like behaviours (e.g., risk-taking, impulsivity) in males, but surprisingly increased risk-taking in females. In both sexes, stimulation of the dopamine system had generally deleterious effects. Both inhibition and stimulation caused females and males to take more cocaine, but paradoxically prevented the increased risk-taking that results from cocaine self-administration. Drawing on the reward deficiency and incentive sensitization theories of addiction, this thesis furthermore proposes a biobehavioural framework through which the present findings may be understood.Medicine, Faculty ofGraduat

Contributions of early life adversity, sex, and traits towards compulsive opioid self-administration

Many people try addictive drugs, but only a small percentage transition from situational/recreational use to the escalating, compulsive, and relapsing disorder of addiction. It is therefore important to characterize the individual differences that render individuals vulnerable to addiction. Substantial evidence suggests that individual differences in early life experience, biological sex, personality traits, and neurobiology of the reward system are all associated with the development of addiction. A dimensional model of how all these factors relate to addiction is necessary, but simultaneously and experimentally probing all these processes in humans is impossible. I therefore conducted a large-scale study to explore these phenomena in rats. Throughout ‘adolescence’ (PND 21-35), I exposed a group of rats to a variety of stressors [i.e., forced-swim, restraint, predator odour, food-restriction, and social isolation; termed ‘early life adversity’ (ELA)]. As adults (PND 60-140), I assessed them on numerous addiction-relevant behavioural traits (i.e., impulsivity, anxiety, novelty-preference, and attraction to reward cues). After trait assessment, I observed the rats’ propensity to self-administer the opiate remifentanil. I then killed a subset of rats and quantified DA receptor mRNA in the mPFC. ELA decreased impulsivity and decreased the rate of acquisition of opiate self-administration in male rats only. Compared to males, female rats exhibited greater anxiety-like behaviour and potentiated opioid self-administration. Anxiety-like behaviour and attraction to reward cues predicted several addiction-relevant behaviours

shinyRGT: An R-Shiny application for extraction and visualization of Rat Gambling Task data

The Rat Gambling Task (RGT) is a well validated rodent model of addiction-like behaviour. It is based on the Iowa Gambling Task (IGT) - a commonly used clinical assay to measure gambling-like behaviour. Rats choose between 4 options to earn as many sugar pellets as possible within 30 min. Each option is associated with dif- ferent reward sizes, but also the probability and duration of the time out punishment. The task is designed such that the optimal strategy for earning sugar pellets is to favour the low risk, low reward options. Consistently selecting the high risk, high reward options results in longer and more frequent time-out penalties. Currently, there is no specialized graphical user interface (GUI) designed to extract, clean, and process RGT data. The installation and use of existing tools are challenging for users lacking coding experience and can be extremely time consuming. To address these issues, we developed a free and open source R-Shiny application called shinyRGT, as a GUI for RGT data extraction, wrangling, and visualization. Clean and usable data can be easily extracted. As well, publication ready plots can be readily generated and annotated from user input. All generated tables can be downloaded as CSV files and generated graphs can be saved to local machines. shinyRGT is deployed at https://andrewcli.shinyapps.io/shinyRGT/for online use. The repository is available at https://github.com/andr3wli/ shinyapps

Pan-striatal reduction in the expression of the astrocytic dopamine transporter precedes the development of dorsolateral striatum dopamine-dependent incentive heroin seeking habits.

The emergence of compulsive drug-seeking habits, a hallmark feature of substance use disorder, has been shown to be predicated on the engagement of dorsolateral striatal control over behaviour. This process involves the dopamine-dependent functional coupling of the anterior dorsolateral striatum (aDLS) with the nucleus accumbens core, but the mechanisms by which this coupling occurs have not been fully elucidated. The striatum is tiled by a syncytium of astrocytes that express the dopamine transporter (DAT), the level of which is altered in individuals with heroin use disorder. Astrocytes are therefore uniquely placed functionally to bridge dopamine-dependent mechanisms across the striatum. Here we tested the hypothesis that exposure to heroin influences the expression of DAT in striatal astrocytes across the striatum before the development of DLS-dependent incentive heroin seeking habits. Using Western-blot, qPCR, and RNAscope™, we measured DAT protein and mRNA levels in whole tissue, culture and in situ astrocytes from striatal territories of rats with a well-established cue-controlled heroin seeking habit and rats trained to respond for heroin or food under continuous reinforcement. Incentive heroin seeking habits were associated with a reduction in DAT protein levels in the anterior aDLS that was preceded by a heroin-induced reduction in DAT mRNA and protein in astrocytes across the striatum. Striatal astrocytes were also shown to be susceptible to direct dopamine- and opioid-induced downregulation of DAT expression. These results suggest that astrocytes may critically regulate the striatal dopaminergic adaptations that lead to the development of incentive heroin seeking habits

Pan‐striatal reduction in the expression of the astrocytic dopamine transporter precedes the development of dorsolateral striatum dopamine‐dependent incentive heroin seeking habits

Publication status: PublishedAbstractThe emergence of compulsive drug‐seeking habits, a hallmark feature of substance use disorder, has been shown to be predicated on the engagement of dorsolateral striatal control over behaviour. This process involves the dopamine‐dependent functional coupling of the anterior dorsolateral striatum (aDLS) with the nucleus accumbens core, but the mechanisms by which this coupling occurs have not been fully elucidated. The striatum is tiled by a syncytium of astrocytes that express the dopamine transporter (DAT), the level of which is altered in individuals with heroin use disorder. Astrocytes are therefore uniquely placed functionally to bridge dopamine‐dependent mechanisms across the striatum. Here we tested the hypothesis that exposure to heroin influences the expression of DAT in striatal astrocytes across the striatum before the development of DLS‐dependent incentive heroin seeking habits. Using Western‐blot, qPCR, and RNAscope™, we measured DAT protein and mRNA levels in whole tissue, culture and in situ astrocytes from striatal territories of rats with a well‐established cue‐controlled heroin seeking habit and rats trained to respond for heroin or food under continuous reinforcement. Incentive heroin seeking habits were associated with a reduction in DAT protein levels in the anterior aDLS that was preceded by a heroin‐induced reduction in DAT mRNA and protein in astrocytes across the striatum. Striatal astrocytes were also shown to be susceptible to direct dopamine‐ and opioid‐induced downregulation of DAT expression. These results suggest that astrocytes may critically regulate the striatal dopaminergic adaptations that lead to the development of incentive heroin seeking habits.</jats:p

Noradrenergic regulation of cue-guided decision making and impulsivity is doubly dissociable across frontal brain regions

Rationale: Win-paired stimuli can promote risk taking in experimental gambling paradigms in both rats and humans. We previously demonstrated that atomoxetine, a noradrenaline reuptake inhibitor, and guanfacine, a selective α2A adrenergic receptor agonist, reduced risk taking on the cued rat gambling task (crGT), a rodent assay of risky choice in which wins are accompanied by salient cues. Both compounds also decreased impulsive premature responding. Objective: The key neural loci mediating these effects were unknown. The lateral orbitofrontal cortex (lOFC) and the medial prefrontal cortex (mPFC), which are highly implicated in risk assessment, action selection and impulse control, receive dense noradrenergic innervation. We therefore infused atomoxetine and guanfacine directly into either the lOFC or prelimbic (PrL) mPFC prior to task performance. Results: When infused into the lOFC, atomoxetine improved decision making score and adaptive lose-shift behaviour in males, but not in females, without altering motor impulsivity. Conversely, intra-PrL atomoxetine improved impulse control in risk preferring animals of both sexes, but did not alter decision making. Guanfacine administered into the PrL, but not lOFC, also altered motor impulsivity in all subjects, though in the opposite direction to atomoxetine. Conclusions: These data highlight a double dissociation between the behavioural effects of noradrenergic signaling across frontal regions with respect to risky choice and impulsive action. Given that the influence of noradrenergic manipulations on motor impulsivity could depend on baseline risk preference, these data also suggest that the noradrenaline system may function differently in subjects that are susceptible to the risk-promoting lure of win-associated cues.This work was supported by a Canadian Institutes of Health Research project grant awarded to CAW (PJT-162312). CSC was supported by a Canadian Graduate Scholarship- Master’s level, TJH was supported by a Marshall Graduate Scholarship, and JDS was supported by a UBC Four Year Doctoral Fellowship

Noradrenergic regulation of cue-guided decision making and impulsivity is doubly dissociable across frontal brain regions.

Acknowledgements: This work was supported by a Canadian Institutes of Health Research project grant awarded to CAW (PJT-162312). CSC was supported by a Canadian Graduate Scholarship- Master’s level, TJH was supported by a Marshall Graduate Scholarship, and JDS was supported by a UBC Four Year Doctoral Fellowship. This work took place at a UBC campus situated on the traditional, ancestral, and unceded land of the xʷməθkʷəy̓əm (Musqueam), sə̓lílwətaʔɬSelilwitulh (Tsleil-Waututh) and Sḵwx̱wú7mesh (Squamish) Peoples. We acknowledge and are grateful for their stewardship of this land for thousands of years.Funder: IMH Marshall Fellows ProgramRATIONALE: Win-paired stimuli can promote risk taking in experimental gambling paradigms in both rats and humans. We previously demonstrated that atomoxetine, a noradrenaline reuptake inhibitor, and guanfacine, a selective α2A adrenergic receptor agonist, reduced risk taking on the cued rat gambling task (crGT), a rodent assay of risky choice in which wins are accompanied by salient cues. Both compounds also decreased impulsive premature responding. OBJECTIVE: The key neural loci mediating these effects were unknown. The lateral orbitofrontal cortex (lOFC) and the medial prefrontal cortex (mPFC), which are highly implicated in risk assessment, action selection, and impulse control, receive dense noradrenergic innervation. We therefore infused atomoxetine and guanfacine directly into either the lOFC or prelimbic (PrL) mPFC prior to task performance. RESULTS: When infused into the lOFC, atomoxetine improved decision making score and adaptive lose-shift behaviour in males, but not in females, without altering motor impulsivity. Conversely, intra-PrL atomoxetine improved impulse control in risk preferring animals of both sexes, but did not alter decision making. Guanfacine administered into the PrL, but not lOFC, also altered motor impulsivity in all subjects, though in the opposite direction to atomoxetine. CONCLUSIONS: These data highlight a double dissociation between the behavioural effects of noradrenergic signaling across frontal regions with respect to risky choice and impulsive action. Given that the influence of noradrenergic manipulations on motor impulsivity could depend on baseline risk preference, these data also suggest that the noradrenaline system may function differently in subjects that are susceptible to the risk-promoting lure of win-associated cues.This work was supported by a Canadian Institutes of Health Research project grant awarded to CAW (PJT-162312). CSC was supported by a Canadian Graduate Scholarship- Master’s level, TJH was supported by a Marshall Graduate Scholarship, and JDS was supported by a UBC Four Year Doctoral Fellowship