Alzheimer's disease is a neurodegenerative disorder with the highest prevalence in the population and for which we do not have a cure so far. The aim of this thesis was to test the mediator system of the N-methyl-D-aspartate receptor and nitric oxide in an animal model of sporadic form of Alzheimer's disease (Samaritan Alzheimer's Rat Model; Taconic Pharmaceuticals, USA). Then compare these results with changes in hippocampal cholinergic system and cognitive tests. The Samaritan rat model is based on the unilateral in vivo application of β-amyloid42 and the pro-oxidative substances (ferrous sulfate heptahydrate and L-buthionine-(S,R)-sulfoximine). Neurochemical methods included testing of the NR1/NR2A/NR2B subunits of the N-methyl-D-aspartate receptor and activity of nitric oxide synthases (neuronal, endothelial, inducible) in the cortex, in both cases in the right and left hemisphere separately. Our results show that Samaritan rats exhibited significant changes in expression of NR2A/NR2B subunits of the N-methyl-D-aspartate receptor and activity of inducible nitric oxide synthase in cortex compared to control rats. The results of glutamatergic system are consistent with changes in activity of cholinergic transporter and cognitive tests (Morris water maze and active allothetic place avoidance)....

NMDA receptors belong to the family of ionotropic glutamate receptors, and are involved in synaptic plasticity, learning and memory. However, overactivation by the agonist glutamate can lead to neuronal death - excitotoxicity. Exitotoxicity is a result of excessive calcium influx into the cell through NMDA receptors, and is associated with many cental nervous system (CNS) diseases. Neurosteroids are endogenous compounds capable of NMDA receptor modulation, thus they may have pharmacological potential in the treatment of CNS disorders. The aim of this work was to investigate how pregnanolone sulfate (PA-S) and pregnanolone hemipimelate (PA-hPim) influence somatic calcium and excitotoxicity. We used fluorescence microscopy for recording changes in somatic calcium concentration. We observed that PA-S had no influence on relative somatic calcium concentration. Synthetic analog PA-hPim increased somatic calcium levels slightly. Next, we used oxygen-glucose deprivation (OGD) in vitro to study the influence of neurosteroids on excitotoxicity. Both PA-S and PA-hPim were neuroprotective in the model of acute OGD in vitro. Moreover, PA-S or PA-hPim pretreatment induced ischemic tolerance to a subsequent OGD episode. Our results suggest that neurosteroids PA-S and PA-hPim are potential candidates for the development...

of Ph.D. thesis Molecular mechanisms of regulation of trafficking and function of different subtypes of NMDA receptors in hippocampal neurons Mgr. Kristýna Skřenková N-methyl-D-aspartate (NMDA) receptors are ionotropic glutamate receptors that play a key role in the mammalian central nervous system. Under physiological conditions, these receptors are important for excitatory synaptic transmission and memory formation. However, under pathological conditions, their abnormal regulation or activation may lead to many neurological and psychiatric disorders, such as Alzheimer's disease, Parkinson's disease, Huntington's disease, epilepsy or schizophrenia. Previous studies have shown that the number and type of NMDA receptors on the cell surface are regulated at multiple levels, including their synthesis, folding, internalization or degradation. During the trafficking of NMDA receptors to the cell surface membrane, both the agonist binding and receptor activation are examined. Moreover, NMDA receptors undergo many posttranslational modifications such as palmitoylation, phosphorylation or N-glycosylation. In this thesis, we studied the molecular mechanisms that may affect the trafficking and functional properties of NMDA receptors in mammalian cells and rat hippocampal neurons. Specifically, we studied i)...

Several trials have reported rapid antidepressant response from the anesthetic drug ketamine although the mechanism behind this effect is not fully understood. Research has focused mainly on ketamine’s action in the brain, including its effects on chemical balance, connections between brain cells and networks, and cognition. Trials with psychedelic drugs have had similar antidepressant results as ketamine, and the quality of the subjective psychedelic experience seems to mediate antidepressant action. Ketamine causes similar alterations of consciousness, which have been viewed as side effects. This thesis examines whether ketamine works in a similar way as psychedelics, where the ketamine-induced dissociative-like experience has a relationship to antidepressant response. Leading theories of depression and ketamine’s action in the brain are presented, and eight studies examining the relationship between ketamine-induced subjective experience and antidepressant response are reviewed. Three included studies found a relationship between psychedelic- and dissociative-like symptoms and reduction in depression, while five did not. The supposed relationship between psychedelic- and dissociative-like symptoms and antidepressant action has not been adequately explored and needs further examination in clinical trials.

An increasing amount of research suggests Ketamine in subanaesthetic doses to be an effective antidepressant for Major Depressive Disorder (MDD) and Treatment-Resistant Disorder (TRD). After the finding that NMDA-receptor antagonists may hold antidepressant effect, several studies have suggested Ketamine to have great effect in relief of depressive symptoms. A time lag between biological and behavioural effects have been shown in currently available antidepressants and are not guaranteed to be efficient; only 30% of patients reach adequate response. The aim for this thesis is to systematically review available studies on the efficiency of Ketamine's antidepressant effects in patients with TRD. Scopus, Web of Science, and PubMed were the databases searched for relevant research regarding the subject. Six articles were included in the analysis. A compilation of the results presented a moderate to large effect size for Ketamine compared to placebo at 24 hours through day seven. It is of immense weight that prolonged adverse effects and possible abuse are taken into consideration for future research, as well as how to sustain the dramatic acute antidepressant effect of Ketamine.

Alzheimer¿s disease is associated with aging and is characterised by a progressive cognitive decline. Its onset in women coincides with the abrupt depletion of ovarian steroids prompting the investigation of utilising oestrogen replacement therapy as restoration or a preventative measure. Gonadal steroids have also recently been implicated in other disease states, particularly schizophrenia. In addition to the cognitive decline, sufferers of Alzheimer¿s disease and schizophrenia display anxiety related behaviour which gonadal steroids have also been shown to ameliorate. In this thesis several paradigms were used to investigate the effects of oestradiol benzoate (EB) on cognition and anxiety, utilising the NMDA receptor antagonist PCP, the muscarinic receptor antagonist scopolamine and the dopamine releasing agent amphetamine to induce a cognitive deficit in rats by different pharmacological mechanisms. The thesis also investigated the effects of EB on a delay dependent cognitive deficit model of forgetfulness in natural aging. Results showed that subchronic PCP dosing failed to induce a significant deficit in the novel object recognition task. Locomotor activity tests demonstrated that the PCP treated rats were sensitised to the treatment suggesting that the PCP dosing regimen was successful. There was no significant effect of oestrogen in the reversal learning model or in the plus maze task designed to explore EB¿s effects on anxiety. However, in the latter task there was a trend towards an anxiogenic effect of EB. Results from the delay dependent model of forgetfulness in natural aging demonstrated that EB could enhance recognition memory, but not spatial memory. The results are discussed in the context of the role of gonadal steroids especially oestrogen in combating the cognitive decline seen in schizophrenia, neurodegenerative disease and natural aging.

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<p>The superior olivary complex (SOC) is comprised of nuclei involved in sound localization. To compute interaural sound level differences, lateral superior olive (LSO) neurons integrate converging glutamatergic inputs from the cochlear nucleus with glycinergic inputs from the medial nucleus of the trapezoid body (MNTB). To compute interaural timing differences, the medial superior olive (MSO) integrates converging glutamatergic inputs from the ipsilateral and contralateral cochlear nucleus. The MSO also receives a major inhibitory input from the MNTB. N-methyl-D-aspartate receptors (NMDARs) are thought to play a role in the developmental refinement of these auditory brainstem pathways. The GluN2A and GluN2B NMDAR subunits confer widely different properties on NMDARs, substantially affecting plasticity. We assessed postnatal developmental messenger RNA (mRNA) expression of GluN1, GluN2A and GluN2B subunits in the LSO, MSO and MNTB using quantitative <em>in-situ</em> hybridization in tissue from 10 litters, ages postnatal day 1 to 36 (P1-36).</p> <p>GluN1 mRNA expression in the LSO, MSO and MNTB decreased with age. In all three nuclei, GluN2B mRNA expression was highest during the first postnatal week, dropping to low levels thereafter. In the LSO, GluN2A levels increased, then decreased to moderate levels. In the MNTB, GluN2A levels decreased from initially high levels. In the MSO, GluN2A levels increased to intermediate levels. The GluN2A/2B ratio increased 2-fold between P1 and P8 in the MNTB, whereas the ratio increased 3-fold between P8 and P15 in the LSO and MSO. The changes in GluN2A:GluN2B ratio are consistent with a developmental switch from GluN2B-containing NMDARs to GluN2A-containing NMDARs. These results are consistent with prior electrophysiological experiments that show NMDAR-mediated currents declining with age in the aVCN-MNTB, aVCN-LSO and MNTB-LSO pathways. The GluN2A subunit exhibited different developmental expression patterns in MNTB, LSO and MNTB, which suggests that GluN2A mRNA expression is locally regulated between nuclei, whereas GluN2B may be globally regulated.</p> / Master of Science (MSc)

Schizophrenia is a debilitating neuropsychiatric disorder that affects approximately one percent of the global population. Aberrant N-methyl-D-aspartate receptors and endoplasmic reticulum stress have been implicated in the pathogenesis of schizophrenia. Despite a century of extensive research, outcomes from best-practice treatments remain dismal. Lurasidone hydrochloride is a novel atypical antipsychotic drug with a unique receptor binding profile that can potentially treat the heterogeneous symptomology of schizophrenia. However, discrepancies in experimental design (i.e. animal models used, symptoms assessed etc.) have yielded conflicting results surrounding the procognitive and antidepressant properties of lurasidone. Furthermore, the limited aqueous solubility of lurasidone poses a considerable challenge for improving antipsychotic drug delivery to the brain and limiting the prevalence of adverse side effects. These obstacles coupled with the elusive pathophysiology of schizophrenia and its incurable nature, highlight the importance of investigating novel therapeutic targets and their underlying mechanisms to improve treatment and enhance the quality of life of patients with schizophrenia. This thesis sought to accomplish three primary objectives: (1) validate the behavioural efficacy of lurasidone hydrochloride; (2) investigate the role of mesencephalic astrocyte-derived neurotrophic factor as a potential therapeutic target of lurasidone; and (3) evaluate the therapeutic potential of intranasal lurasidone administration as a novel method for antipsychotic drug delivery. The data presented within this thesis suggest that repeated lurasidone treatment may be effective at treating the positive, negative, and cognitive symptoms of schizophrenia, but not sensorimotor gating deficits. Furthermore, sub-chronic lurasidone treatment in rats significantly increased the relative expression of mesencephalic astrocyte-derived neurotrophic factor in the rat prefrontal cortex, a primary site of impairment observed in schizophrenia. Lastly, we conclude that lurasidone administered via the nasal route using a novel poly(oligo ethylene glycol methacrylate)-based nanogel formulation required four times less drug to achieve a therapeutic response comparable to traditional intraperitoneal routes. The findings presented within this thesis suggest that lurasidone might be a favourable atypical antipsychotic drug that exerts its therapeutic effects through the modulation of neurotrophic factor expression in the brain regions affected by schizophrenia. This thesis offers new insight that can help guide future studies toward improving the prognosis of patients suffering from schizophrenia. / Thesis / Master of Science (MSc)

<p>Obsessive-compulsive disorder (OCD) is a devastating illness that afflicts around 2% of the world's population with recurrent distressing thoughts (obsessions) and repetitive ritualistic behaviors (compulsions). While dysfunction at excitatory glutaminergic excitatory synapses leading to hyperactivity of the orbitofrontal cortex and head of the caudate - brain regions involved in reinforcement learning - are implicated in the pathology of OCD, clinical studies involving patients are unable to dissect the molecular mechanisms underlying this cortico-striatal circuitry defect. Since OCD is highly heritable, recent studies using mutant mouse models have shed light on the cellular pathology mediating OCD symptoms. These studies point toward a crucial role for deltaFosB, a persistent transcription factor that accumulates with chronic neuronal activity and is involved in various diseases of the striatum. Furthermore, elevated deltaFosB levels results in the transcriptional upregulation of Grin2b, which codes GluN2B, an N-methyl-D-aspartate glutamate receptor (NMDAR) subunit required for the formation and maintenance of silent synapses. Taken together, the current evidence indicates that deltaFosB-mediated expression of aberrant silent synapses in caudate medium spiny neurons (MSNs), in particular D1 dopamine-receptor expressing MSNs (D1 MSNs), mediates the defective cortico-striatal synaptic transmission that underlies compulsive behavior in OCD.</p> / Dissertation