The aims of the thesis were to examine i) whether the endogenous opioid system (EOS) is lateralized in human brain areas involved in processing of emotions and pain; ii) whether EOS responses to unilateral brain injury depend on side of lesion, and iii) whether in human alcoholics, this system is involved in molecular adaptations in brain areas relevant for cognitive control of addictive behavior and habit formation. The main findings were that (1) opioid peptides but not opioid receptors and classic neurotransmitters are markedly lateralized in the anterior cingulate cortex involved in processing of  positive and negative emotions and affective component of pain. The region-specific lateralization of neuronal networks expressing opioid peptides may underlie in part lateralization of higher functions in the human brain including emotions and pain. (2) Analysis of the effects of traumatic brain injury (TBI) demonstrated predominant alteration of dynorphin levels in the hippocampus ipsilateral to the injury, while injury to the right hemisphere affected dynorphin levels in the striatum and frontal cortex to a greater extent than that to the left hemisphere. Thus, trauma reveals a lateralization in the mechanisms mediating the response of dynorphin expressing neuronal networks in the brain. These networks may differentially mediate effects of left or right brain injury on lateralized brain functions. (3) In human alcoholics, the enkephalin and dynorphin systems were found to be downregulated in the caudate nucleus and / or putamen that may underlie in part changes in goal directed behavior and formation of a compulsive habit in alcoholics. In contrast to downregulation in these areas, PDYN mRNA and dynorphins in dorsolateral prefrontal cortex, k-opioid receptor mRNA in orbitofrontal cortex, and dynorphins in hippocampus were upregulated in alcoholics. Activation of the k-opioid receptor by upregulated dynorphins may underlie in part neurocognitive dysfunctions relevant for addiction and disrupted inhibitory control. We conclude that the EOS exhibits region-specific lateralization in human brain and brain-area specific lateralized response after unilateral TBI in mice; and that the EOS is involved in adaptive processes associated with specific aspects of alcohol dependence.

The dorsal anterior cingulate cortex (dACC) has been implicated in a broad range of cognitive and emotional functions, including the processing of negative emotion. Furthermore, abnormalities in dACC activity have been associated with anxiety and depression, disorders in which negative emotion is dysregulated. Thus, a better understanding of the precise contributions of the dACC to negative emotion could give us important insights into the neurobiological mechanisms underlying these debilitating neuropsychiatric disorders. However, despite extensive study of the dACC, its precise role in negative emotion is unclear. Instead there is mounting evidence that rather than being one functionally homogeneous region, subregions of the dACC may have distinct functional roles. This evidence is largely correlational, and interventional studies in experimental animals are required to address this. Accordingly, the work in this thesis causally assessed the contributions of two spatially distinct subregions of the dACC (rostral and caudal) to the regulation of the behavioural and cardiovascular correlates of negative emotion in the common marmoset (Callithrix jacchus). These dACC subregions were targeted with indwelling cannulae to enable pharmacological manipulations to be carried out in a range of tasks, used to assess distinct components of negative emotion, such as conditioned fear and anxiety. The findings suggest that the rostral dACC and the caudal dACC do indeed have distinct contributions to the expression of negative emotion and the regulation of anxiety, respectively. Furthermore, an assessment of the anterograde projections of these subregions provides anatomical support for the observed functional differences.

The overall goal of this dissertation is to elucidate the cellular and circuit mechanisms underlying flexible behavior in the prefrontal cortex. We are often faced with situations in which the appropriate behavior in one context is inappropriate in others. If these situations are familiar, we can perform the appropriate behavior without relearning how the context relates to the behavior — an important hallmark of intelligence. Neuroimaging and lesion studies have shown that this dynamic, flexible process of remapping context to behavior (task switching) is dependent on prefrontal cortex, but the precise contributions and interactions of prefrontal subdivisions are still unknown. This dissertation investigates two prefrontal areas that are thought to be involved in distinct, but complementary executive roles in task switching — the dorsolateral prefrontal cortex (dlPFC) and the anterior cingulate cortex (ACC). Using electrophysiological recordings from macaque monkeys, I show that synchronous network oscillations in the dlPFC provide a mechanism to flexibly coordinate context representations (rules) between groups of neurons during task switching. Then, I show that, wheras the ACC neurons can represent rules at the cellular level, they do not play a significant role in switching between contexts — rather they seem to be more related to errors and motivational drive. Finally, I develop a set of web-enabled interactive visualization tools designed to provide a multi-dimensional integrated view of electrophysiological datasets. Taken together, these results contribute to our understanding of task switching by investigating new mechanisms for coordination of neurons in prefrontal cortex, clarifying the roles of prefrontal subdivisions during task switching, and providing visualization tools that enhance exploration and understanding of large, complex and multi-scale electrophysiological data.

How is the brain capable of mediating pain relief via the mind alone? Placebo analgesia is just such a case, wherein an inert substance yields relief from a number of pain inducing stimuli. Scholars typically separate several factors thought to contribute to the placebo effect into psychological and neurobiological influences. Psychological mechanisms include expectation and conditioning of analgesic effects, while neurobiological mechanisms implicate the opioidergic descending pain system. The current paper proposes an integrative model in which the rostral anterior cingulate cortex (rACC), implicated in cognitive-affective modulation, receives goal-directed input (i.e., expected pain relief) from the prefrontal cortex. As the rACC processes the cognitive difference between expected and actual pain, it recruits a critical descending pain pathway by means of modulating the periaqueductal gray area (PAG). The PAG is a key relay station that connects to other endogenous subsystems of opioidergic pain relief. Whether the rACC and its connection to the PAG are necessary for the placebo effect is a question future research will have to address.

Cognitive dissonance has traditionally been defined as the negative affective state which accompanies inconsistent cognitions and motivates one to make the cognitions consistent. This thesis critically evaluates two theories about cognitive dissonance. The action-based model of dissonance argues that inconsistent cognitions have the potential to interfere with effective and unconflicted action. The new look model of dissonance, contradicting the traditional definition of dissonance, argues that it is aversive consequences rather than inconsistent cognitions that cause dissonance. Recent studies investigating the neural correlates of dissonance show that parts of anterior cingulate cortex and prefrontal cortex seem to be involved in the dissonance process. One of the major predictions of the new look model of dissonance has been undermined by recent evidence. In contrast, the action-based model of dissonance is supported by recent studies.

Vocal plasticity is presumed to be a key element underlying the evolution of human speech and language, but the mechanisms and neuroanatomical basis for this plasticity remain largely unknown. The Mexican free-tailed bat, Tadarida brasiliensis, presents a unique opportunity to advance our understanding of the evolution and neurobiology of mammalian vocal communication because this animal displays elements of vocal complexity and plasticity that are more sophisticated than any mammal other than humans, including non-human primates. Current models of vocal control in mammals do not account for the vocal complexity of free-tailed bats. The purpose of this dissertation is to fill that gap in knowledge by identifying a possible neuronal basis for vocal complexity in free-tailed bats. This will be achieved by 1) providing a detailed analysis of the free-tailed bat’s vocal behaviors, 2) mapping the distribution of neurotransmitter receptor types suspected of involvement in vocal control, 3) identifying brain regions that exhibit increased neuronal activity during vocalizing, and 4) pharmacologically manipulating putative vocal control regions to confirm and characterize their function in vocalizing. Analysis of Tadarida’s vocal behavior indicated that they have a vast vocal repertoire, including many different call types, context-dependent sensory-feedback driven vocal plasticity, and syntactically-organized stereotyped songs. Their vocal behavior changed seasonally, so I mapped the distribution of melatonin binding sites in the brain, finding high densities in the striatum, similar to dopamine receptor distribution. I then used immunohistochemical labeling of the immediate early gene cfos to map neuronal activation in brains of highly vocal bats to find ROIs activated by vocal production. This technique not only identified all previously known regions of the mammalian vocal motor pathway but also revealed activity in novel brain regions that could potentially account for vocal plasticity, including a localized region of the basal ganglia, the dorsolateral caudate nucleus, and the anterior cingulate region of the frontal cortex. Pharmacological excitation of these regions evoked complex vocal sequences similar to the songs recorded in the field and lab. These results support the hypothesis that the mammalian basal ganglia may play a crucial role in the plasticity and complexity of mammalian vocal behaviors.

How is the brain capable of mediating pain relief via the mind alone? Placebo analgesia is just such a case, wherein an inert substance yields relief from a number of pain inducing stimuli. Scholars typically separate several factors thought to contribute to the placebo effect into psychological and neurobiological influences. Psychological mechanisms include expectation and conditioning of analgesic effects, while neurobiological mechanisms implicate the opioidergic descending pain system. The current paper proposes an integrative model in which the rostral anterior cingulate cortex (rACC), implicated in cognitive-affective modulation, receives goal-directed input (i.e., expected pain relief) from the prefrontal cortex. As the rACC processes the cognitive difference between expected and actual pain, it recruits a critical descending pain pathway by means of modulating the periaqueductal gray area (PAG). The PAG is a key relay station that connects to other endogenous subsystems of opioidergic pain relief. Whether the rACC and its connection to the PAG are necessary for the placebo effect is a question future research will have to address.

Introduction: The aim of the project was to investigate the effects of antidepressants on brain networks whilst at rest. My hypothesis was that antidepressants work by reversing persistent activity in the brain’s default mode network (DMN). The DMN is implicated in self-reflection and rumination in MDD. The methodologies and results of studies of resting state networks in MDD and the effects of antidepressants are reviewed in the thesis. Increasing evidence implicates glutamate in the action of antidepressant drugs. Whether there are illness related changes in glutamate function is unresolved, largely because of the lack of techniques for assessing it. Ketamine and other NMDA antagonists have improved MDD symptoms within 24 hours though the effects are short lasting. The molecular neural networks involved in ketamine’s putative antidepressant effects are unclear. The thesis reviews the evidence. Much evidence implicates ACC as a site of action of antidepressant effects but whether this is through its regulation of the DMN or other networks is not known. This thesis compares the effect of ketamine and citalopram on ACC-related systems. Method: The thesis combines two systematic reviews of the effects of MDD and antidepressant drugs on i) resting state networks (53 studies) and ii) glutamate neurotransmission (45 studies of clinical efficacy of ketamine). There are two experimental chapters. The first describes investigation into two rapid acting antidepressant drugs acting via glutamate mechanisms. 54 unmedicated cMDD were scanned across two centres on 3T MRI scanners while being infused with placebo (0.5% saline), 0.5mg/kg ketamine or 100mg AZD6765 over 1 hour. fMRI resting state data between drug treatments was compared for the final 25 minutes of the drug infusion and for a 25 minute resting state scan a day later. The second experimental chapter examines whether these effects were shared by citalopram, a standard antidepressant. 67 unmedicated cMDD, rMDD and HC were administered citalopram 7.5mg i.v. and scanned on a 1.5T MRI scanner. In a second study 63 cMDD and HC were administered i.v. citalopram 7.5mg or placebo (0.5% saline). fMRI resting state data for the final 12 ½ minutes following drug infusion was compared. Independent Component Analysis was performed using the Group ICA for fMRI toolbox. The resting component with the highest spatial correlation to the ACC was used. Brain maps of the intensity of the selected component were constructed for each individual. Group averages were calculated and compared using SPM. Regional analysis was performed using Marseille Boite a Regions d'interet. Results: On day 1 AZD6765 significantly increased mean intensity of ACC resting component in the right insula, right IPL and left cingulate gyrus greater than ketamine or placebo. Ketamine increased mean intensity of ACC resting component greater than placebo in the right lentiform nucleus and left mFG. Significantly decreased mean intensity of ACC resting component in the left insula in the AZD6765 group compared to placebo was noted. On day 2 AZD6765 increased mean intensity of ACC resting component greater than ketamine and placebo in the left and right lentiform nuclei. AZD6765 reduced mean intensity of the ACC resting component in the left and right MFG. The first citalopram study revealed reduced mean intensity of ACC resting component in cMDD compared to rMDD and HC in PCC. rMDD had reduced mean intensity of ACC resting component in the precuneus compared to HC. In the second study, citalopram had no effect in HC but normalised precuneus activity in cMDD producing a significant drug x group interaction. Conclusions: The acute antidepressant effects of citalopram are modulated by changes in the bilateral precuneus. The precuneus is central to connectivity with other regions in MDD. It has a prominent role in the DMN and is linked to rumination. The mechanism of the antidepressant effects of AZD6765 is different from those of ketamine and citalopram. The insula, IPL, MFG, cingulate gyrus and lentiform nuclei are all regions implicated in MDD suggesting antidepressant effects. The rapid antidepressant effects of AZD6765 are possibly due to a resetting of the interface between DMN and salience networks.

Autism spectrum disorder (ASD) is associated with functional abnormalities of the anterior cingulate cortex (ACC), a brain area that mediates social behavior. Given evidence of a role of inflammation in ASD, markers of pro-inflammatory and anti-inflammatory microglia were studied using postmortem ACC tissues from ASD and age-matched typically developed control donors. Gene expression levels of pro-inflammatory (CD68, HLA-DRA, IL1B, NOS2, PTGS2) and anti-inflammatory (ARG1, IGF1, MRC1, PPARG) microglial genes were measured using quantitative real-time PCR. Additionally, brain sections were immunohistochemically stained for a microglial marker. Expression levels of IGF1 were modestly higher, while the expression of MRC1 was modestly lower in ASD donors when compared to control donors. No other differences in gene expression levels between the two groups of donors were observed. Statistical significance for changes in expression levels IGF1 and MRC1 did not survive correction for multiple comparisons. Further research on anti-inflammatory microglial involvement in ASD is warranted.

Generalized anxiety disorder (GAD) is a prevalent anxiety disorder that is characterized by persistent excessive worrying that is often difficult to control. The pathology of GAD has been associated with abnormal neural activity and functional connectivity. This systematic review has examined the central neural correlates of GAD which are the prefrontal cortex (PFC), the anterior cingulate cortex (ACC) and the amygdala and how activation and functional connectivity in these brain areas differ between patients with GAD and healthy controls. This review also investigated how abnormal functional connectivity and activation in these brain regions relates to worry which is the most prominent psychological symptom in patients with GAD. A systematic review was conducted and seven original functional magnetic resonance imaging (fMRI) studies were included after a literature search on PubMed, Scopus and, Web of Science. The main findings from this review revealed decreased activation in the PFC and ACC and enhanced activation in the amygdala during the viewing of negative stimuli in patients with GAD. Identifying the neural correlates of GAD and how it relates to worry may provide improved treatment in the future such as developing more effective psychotropic drugs or improved psychotherapy. GAD has been associated with lower well-being and life satisfaction and may even be a risk factor for suicidal thoughts. One of the limitations from this review is that several of the included studies recruited patients with comorbidities and for that reason results from these studies cannot be generalized and applied to individuals with GAD.