Florey Department of Neuroscience and Mental Health - Theses

Permanent URI for this collection

Search Results

Now showing 1 - 5 of 5
  • Item
    Thumbnail Image
    The effects of chronic methamphetamine exposure during adolescence on brain and behaviour
    Luikinga, Sophia Johanna ( 2018)
    Methamphetamine is a highly addictive psychostimulant that is used world-wide. The age of initial methamphetamine use typically occurs during adolescence, which is a particularly vulnerable period to the development of addiction. Therefore, this thesis aimed to elucidate the effects of methamphetamine during adolescence compared to adulthood on brain and behaviour. My first study examined the effects of either experimenter-injected binge exposure or self-administration of methamphetamine on subsequent fear related behaviours because the cycle of anxiety following methamphetamine use and withdrawal may be different in adolescence, which may contribute towards their methamphetamine use. In the binge model, adolescent and adult rats were injected with high and increasing doses of methamphetamine followed by fear conditioning. Extinction recall was impaired due to methamphetamine in adults but not adolescents. Methamphetamine self-administration did not differ between adults and adolescents, but it caused a deficit in the acquisition of conditioned fear in adults but not adolescents. In summary, prior methamphetamine exposure had effects on fear conditioning and extinction only in adults, but not in adolescents. My second study examined methamphetamine-cue extinction and cue-induced reinstatement following methamphetamine self-administration. While cue extinction reduced cue-induced reinstatement in adults and adolescents, adolescents showed higher cue-induced reinstatement compared to adults following 2 sessions of cue extinction. This chapter further showed that while adolescent and adult rats acquire methamphetamine self-administration similarly when the dose starts at 0.03 or 0.01 mg/kg/infusion, adolescents increase their methamphetamine intake when dose is increased from acquisition. This suggests that adolescents may be more vulnerable to escalate their methamphetamine intake if the dose is increased. My final study investigated the potential neuroadaptations induced by methamphetamine self-administration in adult and adolescent rats. Genome wide transcriptome analysis of the dorsal striatum identified 30 potential candidate genes with significant RNA expression changes due to methamphetamine. Based on ingenuity pathway analysis, 6 genes were followed up for quantitative real-time polymerase chain reaction validation. The most notable finding was that methamphetamine self-administration caused a decrease in solute-carrier family 18 member a1 (slc18a1) in adolescents but not in adults. The changes in the level of protein encoded by this gene, vesicular monoamine transporter 1 (VMAT1), were further validated by western blot. This chapter identifies factors that may explain adolescent vulnerability to addiction, such as their resistance to cue extinction and escalation of intake with dose increase. Age specific changes in gene expression following methamphetamine self-administration have been observed, which may explain the age-differences in methamphetamine-taking and seeking. Ultimately, these discoveries may provide novel ways to treat methamphetamine addiction depending on the age of onset of methamphetamine use.
  • Item
    Thumbnail Image
    Prefrontal dopaminergic mechanisms of adolescent cue extinction learning
    Zbukvic, Isabel ( 2016)
    Addiction and anxiety disorders represent the most prevalent mental illnesses in young people worldwide. Unfortunately, adolescents attain poorer outcomes following extinction-based treatment for these disorders compared to adults. Cue extinction learning involves dopamine signaling via the dopamine 1 receptor (D1R) and dopamine 2 receptor (D2R) in the medial prefrontal cortex. In particular, the infralimbic cortex, a subregion of the medial prefrontal cortex, has been implicated in extinction learning in both adolescent and adult rodents. The prefrontal dopamine system changes dramatically during adolescence. However, the role of prefrontal dopamine in adolescent cue extinction learning is poorly understood. Therefore, this thesis aimed to elucidate the role of prefrontal dopamine in adolescent cue extinction, using cocaine self-administration and fear conditioning in rats. My first study examined cocaine self-administration and cocaine-associated cue extinction in adolescent versus adult rats. Adolescents displayed a deficit in cocaine-cue extinction learning compared to adults (postnatal day [P]53 and P88 on cue extinction day, respectively). A single infusion of the full D2R agonist quinpirole into the infralimbic cortex prior to extinction enhanced adolescent cue extinction to reduce relapse-like behavior the next day. This effect was recapitulated by a systemic injection of the partial D2R agonist aripiprazole, an FDA-approved drug for the treatment of psychosis with strong translational potential. My second study examined fear conditioning and extinction in adolescent and adult rats. I first aimed to optimize behavior in late adolescent (P53) and adult (P88) rats during the dark phase of their 12-hour light-dark cycle, to remain consistent with conditions of the previous chapter. However, this produced unreliable behavioral results. In contrast, adolescent rats (P35) consistently display a deficit in long-term fear extinction compared to adults (P88) during the light phase. Infusion of the D1R agonist SKF-81297 into the infralimbic cortex prior to fear extinction had no effect for either age group. However, infusion of quinpirole into the infralimbic cortex significantly enhanced long-term fear extinction in adolescents, whereas it delayed within-session extinction in adults. Interestingly, an acute systemic injection of aripiprazole improved long-term fear extinction in adults. My final experiments measured prefrontal gene expression for D1R, D2R, and D1R relative to D2R (D1R/D2R ratio) in naïve rats across adolescent development, or following cocaine-cue, or fear extinction. There were no significant differences in prefrontal dopamine receptor gene expression across naïve rats age P35, P53, and P88. Following cocaine-cue extinction, prefrontal D1R gene expression was upregulated in adults but not adolescents. By comparison, following fear conditioning, adolescents showed higher D1R and D1R/D2R ratio gene expression compared to adults. D1R/D2R ratio was modulated in opposite directions following fear extinction learning during adolescence versus adulthood. These findings show that adolescents are impaired in extinction of emotionally salient cues across both appetitive (drug) and aversive (fear) learning domains. Functional and molecular data provide novel evidence for divergent involvement of prefrontal dopamine in cue extinction learning across adolescent development. Results not only extend understandings of extinction learning in general, but represent an exciting step towards finding new therapeutic targets to facilitate exposure-based therapy in the clinic.
  • Item
    Thumbnail Image
    The role of CRFR1 in addiction and anxiety disorders
    Chen, Nicola Alexandra ( 2016)
    Addiction and anxiety disorders are highly co-morbid, and represent a huge burden on society. The central role of stress-reactivity in the pathogenesis and maintenance of both of these diseases has led to the identification of corticotropin-releasing factor (CRF) signalling as a key factor in these effects. The focus of this thesis was the ventral tegmental area (VTA), as it is a site where reward- and fear- related circuitry converge and can be modulated by CRF. The broad aims of this thesis were to examine the role of VTA CRF receptor 1 (CRFR1) in animal models of reward-seeking and conditioned fear to understand how these systems can become dysregulated in addiction and anxiety disorders. To this end, chapter 3 of this thesis validated a technique for the viral-mediated downregulation of CRFR1 within the VTA, and chapter 4 established a novel model of stress-induced reinstatement of cocaine-seeking in mice. These techniques were then implemented to examine the effects of VTA CRFR1 knockdown on the acquisition, extinction, and reinstatement behaviours. Chapters 5 and 6 are two separate publications demonstrating that VTA CRFR1 signalling is differentially involved in various components of cocaine-seeking and conditioned fear. In chapter 5, knockdown of CRFR1 in the VTA blocked stress-induced reinstatement of cocaine-seeking and attenuated cued cocaine-seeking, without any effects on self-administration or extinction responding. This was a specific effect on drug-related behaviours, as there were no changes to operant responding for sucrose rewards. In chapter 6, VTA CRFR1 knockdown enhanced the expression of conditioned fear, but had no effects on fear extinction or reinstatement. This evidence suggests that CRFR1 participates in distinct subcircuits within the VTA which mediate fear and reward-seeking.
  • Item
    Thumbnail Image
    Relaxin-3/RXFP3 system in alcohol self-administration and relapse
    Kastman, Hanna Erika ( 2016)
    Alcoholism is a chronic relapsing disorder, accounting for 10% of disability-adjusted life years lost in industrialized countries. Our understanding of the neurobiology of addiction is far from complete and due to high relapse rates, there is a need to identify new therapeutic targets to assist the development of better treatments. In early studies, the neuropeptide relaxin-3 was implicated in the regulation of stress responses and arousal/motivational behaviours such as feeding, as well as spatial memory. Relaxin-3 is primarily expressed in large GABAergic neurons of the nucleus incertus (NI) in the hindbrain that project topographically to forebrain regions containing neurons expressing the native relaxin-3 G-protein-coupled receptor, RXFP3, several of which are implicated in the control of drug-seeking behaviour. Therefore, the present study directly investigated the role of the central relaxin-3/RXFP3 signalling system in alcohol- and sucrose-seeking in alcohol-preferring (iP) and Wistar rats. Firstly, the effect of central antagonism of RXFP3 using a receptor-selective relaxin-3 analogue peptide on self-administration of alcohol and sucrose was investigated; and then the effect of the same antagonist treatment on cue- and stress-induced reinstatement of both sucrose- and alcohol-seeking was studied. Thereafter, the impact of RXFP3 antagonism within the stress-related bed nucleus of stria terminalis (BNST) on stress-induced reinstatement of alcohol-seeking was also examined. Central antagonism of RXFP3 reduced cue- and stress-induced reinstatement of alcohol-seeking, but did not markedly alter sucrose-seeking in either paradigm, suggesting a specific effect on alcohol-related behaviour(s). Alcohol consumption and stress-induced reinstatement were also both attenuated by local RXFP3 antagonism within the BNST. These data are the first identifying a role for relaxin-3/RXFP3 signalling in alcohol use and seeking. In light of these initial findings and the enrichment of RXFP3 in other stress-related brain areas, the ability of RXFP3 antagonism within the central amygdala (CeA) to modulate alcohol self-administration and stress-induced reinstatement of alcohol-seeking was also examined. Indeed, RXFP3 antagonism in CeA also reduced alcohol self-administration and stress-induced reinstatement of alcohol-seeking, further indicating an involvement of native relaxin-3/RXFP3 signalling in stress-induced reinstatement and potential interactions with other major peptide transmitter systems implicated in these behaviours, such as corticotropin-releasing factor (CRF) and orexin. NI relaxin-3 neurons express the CRF type 1 receptor (CRF1) and are activated by CRF; likewise there is anatomical evidence for the expression of orexin-1 (OX1) and orexin-2 (OX2) receptors in the NI. Accordingly, the effect of bilateral NI infusions of the CRF1receptor antagonist (CP376395), the OX1 receptor antagonist (SB-334867) and the OX2 receptor antagonist (TCS-OX2-29) on stress-induced reinstatement of alcohol-seeking was examined. CP376395 and TCS-OX2-29 reduced stress-induced reinstatement of alcohol-seeking, whereas SB-334867 was ineffective. These data suggest that CRF- and orexin-mediated activation of NI neurons occurs during stress-induced reinstatement, via CRF1 and OX2 receptor signalling events. Together, these data demonstrate that the relaxin-3/RXFP3 system can modulate stress-induced reinstatement of alcohol-seeking via both forebrain sites and a potential CRF- and orexin-primed activation of the NI during stress exposure. These findings have added significantly to our knowledge of the neurocircuitry that underpins stress-induced relapse-like behaviour and to our general understanding of addictive behaviours, with implications for the development of better treatment strategies and the identification of potential drug targets for treating alcoholism and other stress-related addictions.
  • Item
    Thumbnail Image
    The role of the metabotropic glutamate 5 and adenosine 2A receptors in methamphetamine addiction
    CHESWORTH, ROSE ( 2015)
    Methamphetamine (METH) is a highly addictive psychostimulant for which there are no pharmacotherapies. Current theories of drug addiction suggest a dysregulation of dopamine and glutamate systems in the development and maintenance of addition. Two receptors which modulate dopamine and glutamate transmission, and which have been implicated in animal models of drug-taking behaviour for other drugs of abuse (e.g. alcohol, cocaine, opiates) are the metabotropic glutamate 5 (mGlu5) and the adenosine 2A (A2A) receptors. This project used germline KO mice to identify the role of these receptors in METH-induced behaviour, and determine a neural locus where these receptors might act to mediate this behaviour. Germline deletion of mGlu5 resulted in a deficit in extinction learning for METH in an operant self-administration paradigm, and an increased propensity to reinstate to drug-associated cues. mGlu5 KO mice also demonstrated enhanced locomotor activity when re-exposed to a drug- associated context compared to wildtype (WT) littermates, suggesting mGlu5 may modulate the contextual salience of drug-associated cues and contexts. In contrast, A2A KO mice exhibited abolished conditioned place preference (CPP) and a reduction in the motivation to self- administer METH under high response requirements. There was also a reduction in sucrose self-administration under higher reinforcement schedules in A2A KO mice, suggesting this receptor is involved in the rewarding and motivational properties of both METH and sucrose. c-Fos immunohistochemistry was used to determine a locus where A2A could mediate the rewarding properties of METH, as assessed by CPP. Initially, Fos-immunoreactivity (IR) was examined following the expression of METH CPP in A2A WT and KO mice; however, there was a global reduction in Fos-IR throughout the forebrain in A2A KO mice, preventing the identification of a potential locus. A second experiment was conducted in A2AloxP/loxP mice, examining Fos-IR following the expression vs. non-expression of CPP. This experiment identified the nucleus accumbens (NAcc) shell and the infralimbic cortex as regions activated following the expression of METH CPP. From this, it was hypothesised that A2A activity in the NAcc shell might mediate METH reward. This hypothesis was addressed using viral mediated knockdown of A2A. Adeno-associated virus encoding Cre-recombinase (AAV-Cre) or mCherry (a control fluorophore) were microinjected into the rostral medial NAcc shell of A2AloxP/loxP mice. This resulted in a deletion of approximately 20% of A2A in the rostral medial NAcc shell. There was no effect of AAV-Cre mediated deletion on the expression of METH reward or METH-induced locomotor behaviour. Furthermore, there was no correlation between the degree of knockdown and CPP, supporting the conclusion that a ~20% knockdown of A2A in the rostral medial NAcc shell had no effect on METH reward. In summary, the findings of this thesis implicate A2A in reward and motivated behaviour for METH, but also in these behaviours for natural reinforcers such as sucrose. Although neural correlates suggested increased activity in the NAcc shell during the expression of METH reward- context associations, I was unable to confirm the involvement of A2A in this behaviour using region specific receptor knockdown. In contrast, mGlu5 appears involved in cognitive processes associated with recognition of drug-associated stimuli and the extinction of drug-taking behaviour.