Indiana University-Purdue University Indianapolis (IUPUI) / The opioid crisis has resulted in an unprecedented number of neonates born with prenatal opioid exposure; however, the long-term effects of opioid exposure on offspring behavior and neurodevelopment remain relatively unknown. I developed a translational mouse model of prenatal methadone exposure (PME) that resembles the typical pattern of opioid use by pregnant women who first use oxycodone then switch to methadone maintenance pharmacotherapy, and subsequently become pregnant while maintained on methadone. PME produced substantial impairments in offspring growth, sensorimotor milestone acquisition, and activity in an open field. Furthermore, these behavioral alterations were associated with significant disruptions in the primary motor cortex (M1). Notably, layer 5 pyramidal neurons of the M1 displayed significantly increased voltage sag which is primarily mediated by HCN1 channels. Interestingly, the α2-adrenergic receptor, a known modulator of HCN1 channels, displayed significantly increased expression in the M1 of PME animals. The locomotor activity in an open field was significantly reduced following in vivo pharmacological activation of the α2-adrenergic receptor with clonidine in PME offspring suggesting this may be therapeutic target for the hyperactivity associated with prenatal exposure to opioids. Previous work has also described an association between prenatal opioid exposure and alterations in opioid reward-related behavior; however, the effect of PME on alcohol reward remains undetermined. Given the widespread accessibility and usage, alcohol represents the most likely addictive substance the growing population of opioid exposed neonates will encounter as they age. I discovered that PME disrupts conditioned preference for alcohol, enhances the locomotor stimulating effects of alcohol, and increases alcohol consumption in a sex-dependent manner. This alcohol-reward phenotype in PME offspring was associated with altered excitatory neurotransmission and disrupted cannabinoid-mediated long-term depression (CB-LTD) in the dorsolateral striatum, an important substrate involved in compulsive drug use. Further work is required to determine the specific inputs at which CB-LTD is disrupted and if restoring this form of plasticity in PME animals prevents the enhanced alcohol addiction phenotype. / 2023-03-02

Autism spectrum disorder (ASD) refers to a highly prevalent neuropsychiatric disorder, currently affecting one in every 68 children. ASD is understood as a heterogeneous disorder, and individuals with this condition vary considerably in terms of symptom presentation. This heterogeneity contributes to the difficulty faced by researchers and clinicians in trying to determine the precise underlying mechanisms and treatment for this condition. Furthermore, it remains unknown whether the variations in symptom manifestation are attributed to differences in underlying etiologies of the disorder or other factors as yet to be identified. Currently, it is believed that ASD is likely due to the interaction between different genetic and environmental factors. The maternal immune system is one example of where the environment may act upon genetic predispositions and lead to altered fetal brain development. Considering the importance of the immune environment during fetal development, maternal antibodies (Abs) directed against fetal proteins have been considered as potentially playing a critical function in the pathology of ASD. This thesis examines the literature focused on the role of maternal Abs in fetal development and their impact on the neuropathology of ASD. Studies have collected samples from mothers of children diagnosed with ASD and examined the reactivity patterns of the maternal Abs against fetal proteins. Through review and inspection of methodologies and results, this thesis highlights the important insights obtained as well as proposes possible reasons for the disparity in findings. Lastly, this thesis proposes future directions and therapeutic implications of identifying the maternal Abs that could be involved in at least a subset of ASD cases.

ABSTRACT Paediatric HIV remains one of the most significant challenges to face children, their families and their health care providers in South Africa. The prevalence rate of paediatric HIV infection in South Africa is set to remain high until such time as universal access to antiretrovirals for prevention of mother to child transmission is achieved, and the mother to child transmission rates of HIV start to come down. HIV is neurotrophic and is known to invade the developing central nervous system and cause widespread damage. The result of this is a well described encephalopathy which has the potential to affect all facets of development. Children in South Africa who are infected with HIV are vulnerable to a number of factors which may cause developmental delay. Poverty and malnutrition are likely to exacerbate the developmental delay caused by HIV encephalopathy. Physiotherapists in South Africa have not become involved in the long term management of children infected with HIV and paediatric HIV clinics do not routinely offer any rehabilitation services. The prevalence and extent of developmental delay in HIV infected children in South Africa has not been established. Despite the fact that a number of studies have highlighted the prevalence of developmental delay in Western countries, no intervention studies addressing this problem could be found. Caregivers of HIV infected children face numerous stressors. Poverty, stigma and their own health care needs make parenting an HIV positive child even more challenging. The needs of caregivers of HIV infected children have not been well researched in the context of developing countries. The aim of this study was therefore to establish whether a basic home stimulation programme would have any impact on the neurodevelopmental status of young children infected with HIV, and on the parenting stress levels of their caregivers. Further objectives of the study were to establish the prevalence and progression of developmental delay in HIV infected children; to monitor the effect of antiretrovirals on neurodevelopment; to determine who the caregivers of HIV infected children were and to determine what factors were predictive of neurodevelopmental status and parenting stress levels. In order to meet these objectives a longitudinal randomized controlled trial was conducted. One hundred and twenty two HIV positive children, under two and a half years of age, were recruited for this study at Harriet Shezi Children’s Clinic at Chris Hani Baragwanath Hospital in Soweto. Children were randomly assigned to a control or an experimental group. The developmental status of all children was monitored over a year using the Bayley Scales of Infant Development II. Parenting stress was monitored with the Parenting Stress Index/Short Form. Children in the experimental group received a basic home stimulation programme, which was updated every three months when they came to visit the clinic, as well as all the usual clinic services. Children in the control group received all the usual services at the clinic but no stimulation programme. Most of the children in the sample were cared for by their biological mothers. They came from poor homes with limited access to common household amenities. Most of the caregivers had not completed 12 years of schooling. The children in the control and experimental groups were well matched for all their baseline measurements and demographic characteristics. At baseline the children were wasted and stunted and had very low CD4 counts. Only 16% of the children were on antiretrovirals at baseline assessment. The children were severely delayed with respect to both motor and cognitive development. The parenting stress levels of the caregivers were very high at baseline. Over the period of one year the children in the experimental group showed a significantly greater improvement in cognitive (p=0.01) and motor (p=0.02) development when compared to children in the control group. Although the children improved, they still had a degree of developmental delay at the end of the study period. The parenting stress levels decreased significantly for caregivers in both the control and the experimental groups (p<0,001), but there was no significant difference between the two groups (p=0.057). The groups were well matched at all time points for anthropometric measures and CD4 counts with no significant differences being found. There was also no difference in the number of children on antiretroviral therapy between the groups at any time. Children who were antiretroviral naïve at the start of the study and then started highly active antiretroviral therapy showed a significant improvement in motor development (p<0.001), but no improvement in cognitive development (p=0.77). A combination of a number of factors was predictive of developmental status. This included growth parameters, CD4 counts and the age of the child. Being in the experimental group and being older at baseline assessment were important predictors of improvement in MDI and PDI over time. Parenting stress was predicted by a number of factors, including educational level of the caregiver, type of housing and the number of children in the household. A decrease in parenting stress was most likely in caregivers who were better educated and who lived in households with fewer adults. These results signify that a basic home programme can significantly improve both the cognitive and motor development of young children infected with HIV. This programme was simple and easily implemented and should become standard practice at paediatric HIV clinics in South Africa. The current protocol for administering antiretrovirals in South Africa allowed for motor, but not cognitive improvement in young children commencing treatment. Parenting stress was not affected by the addition of a basic home stimulation programme.The psychosocial and developmental needs of South African children infected with HIV are complex and multifaceted. Further research is needed to establish the best possible interventions for these children and their families.

G-protein coupled receptor 50 (GPR50) is a genetic risk factor for psychiatric illness. It is a member of the melatonin receptor family, which includes the well characterised melatonin receptors 1 and 2 (MT1 and MT2). However, the ligand for GPR50 remains elusive and little is known about GPR50 signalling pathways. Despite this, GPR50 is known to enhance neurite outgrowth and inhibit the actions of the neurite outgrowth inhibitor NOGO-A. Existing evidence also indicates a role in lipid metabolism; GPR50 knockout mice displayed abnormalities in energy homeostasis and weight control, whilst sequence variants are associated with altered lipid levels in humans. Further, a yeast-2-hybrid screen identified SREBF2 and ABCA2, regulators of lipid homeostasis, as GPR50 interactors. This thesis explores the role of GPR50 in neuronal development and lipid metabolism. The work presented in this thesis shows that GPR50 promotes neuronal differentiation. Overexpression significantly increased the number of neurites per cell in SH-SY5Y cells. Further, dendritic branching was enhanced by GPR50 transfection in hippocampal and cortical neurons (DIV 14). In hippocampal neurons, GPR50 transfection also lead to a shift towards spine maturity although it had no effect on spine morphology, suggesting GPR50 enhances spine development but may not alter synaptic strength. The effect of GPR50 on neuronal morphology may be driven by actin remodelling. Immunocytochemistry showed an enrichment of GPR50 in highly dynamic regions of the membrane, i.e. the lamellipodia and dendritic spines. Overexpression in SH-SY5Y cells also resulted in an increase in WAVE-2 and phosphorylated RAC1/CDC42, key modulators of actin dynamics. Additionally, GPR50 transfection altered the protein level and localisation of α- catenin, another regulator of actin organisation, in HEK293 and SH-SY5Y cells respectively. An involvement of GPR50 in lipid metabolism has also been demonstrated in this thesis. Verification of the Y2H study suggested GPR50 does not physically interact with SREBF2 or ABCA2. However, ABCA2 appears to induce the intracellular localisation of GPR50 in several cell lines. In SH-SY5Y cells, this was mimicked by the inhibition of cholesterol trafficking, suggesting the translocation of GPR50 to the plasma membrane is dependent on cholesterol transport. Further, the depletion of lipoproteins resulted in the downregulation of GPR50, indicating a responsiveness to lipid levels. Finally, GPR50 increased lipid metabolism, as seen by a decrease in intracellular lipid droplets upon GPR50 overexpression. The data presented here extends previous work indicating a role of GPR50 in neurodevelopment. It also highlights a potential mechanism by which GPR50 regulates neuronal morphology, i.e. via actin remodelling. Reports that GPR50 is involved in energy homeostasis is also supported in this thesis, further, results presented here suggest GPR50 is specifically involved in lipid metabolism. These processes are often disrupted in mental illness, thus this work may provide a functional link between GPR50 and psychiatric disorders.

Psychiatric illnesses are disorders that affect millions worldwide. Evidence from quantitative and molecular genetics analysis suggests a strong genetic component to these disorders. There is also evidence that embryonic neurodevelopment is a key period in the progression schizophrenia. The aim of the present study was to use post-mortem human hippocampus from subjects of a variety of psychiatric phenotypes to investigate neurodevelopmentally- relevant gene expression in this region of the adult human brain. Particular interest is paid to schizophrenia risk gene DISC1; it has been shown to exhibit linkage and association to schizophrenia and is highly involved in embryonic and post natal neurodevelopmental processes. The results reported in this study indicate that DISC1 binding partners, and genes used to mark neurogenesis, can be found aberrantly expressed in schizophrenia and bipolar disorder, relative to controls. The results also suggest that DISC1 genotype may predict expression patterns of DISC1 binding partners and neurogenesis markers, irrespective of diagnosis. This may provide clues to the timing and nature of abnormal brain development in this illness and aid in development of treatment strategies.

Student Number : 7809567 - MSc dissertation - School of Therapeutic - Faculty of Health Sciences / HIV/AIDS in Sub-Saharan Africa has resulted in a major increase in the number of HIV infected children and orphans. HIV infected children are at risk of developmental delays and growth impairments which is further compromised by poor living conditions. Institutionalisation is not the preferred method of caring for children in need, however, it does provide a stable environment, shelter, nutrition and medical care. Objective: To compare the anthropometric measurements and neurodevelopment of HIV infected and HIV uninfected children who were vertically infected, not on antiretroviral treatment and residing in institutions in Gauteng, South Africa. Method: A comparative, longitudinal study of 16 HIV infected and 24 HIV uninfected children between the ages of 16 and 42 months. The Bayley Scale of Infant Development II (MDI and PDI) was used to evaluate neurodevelopment. The children’s mean z-scores for weight-for-age, height-for-age, weight-forheight and head circumference-for-age were calculated. Evaluations were carried out at two time points, seven months apart. Results: The HIV infected children scored significantly lower than HIV uninfected children at both time points, in neurodevelopmental (MDI p<0.02 and p<0.00; PDI p<0.00 and p<0.00) and anthropometric measurements for-age (weight p<0.00 and p<0.01; height p<0.00 and p<0.00; head circumference p<0.00 and p<0.07). Both groups (HIV infected and HIV uninfected) showed a significant improvement over time regarding to their weight-for-age (p<0.00; p<0.01) and head circumference-for-age (p<0.01 and p<0.08). The height-forage showed no significant improvement in the HIV infected group (p>0.2) but did in the HIV uninfected group (p<0.03). There was a severe delay in the mental abilities of both the HIV infected and HIV uninfected children and the motor abilities of the HIV infected children, which did not change over time, but the motor abilities of the HIV uninfected children did improve significantly. Conclusion: The HIV virus affects the neurodevelopment and growth of HIV infected children. Both groups showed an improvement over time in their growth particularly weight-for-age indicating that they may have benefited from their institutionalisation.

Background: Docosahexaenoic acid (DHA), choline and uridine-5-monophosphate (UMP) are important brain nutrients which form phosphatidylcholine, the most abundant brain membrane phospholipid. DHA, choline and UMP supplementation increases rodent brain phospholipids, synaptic components, functional brain connectivity and cognitive performance. This novel pilot study supplemented infants at risk of neurological impairment (ARNI) with a nutrient combination containing these neurotrophic compounds. Aims: 1) In a double blind randomised control trial (RCT), investigate if intake of a specific nutrient combination improves neurodevelopmental and visual outcome in infants ARNI. 2) Using novel measures of cortical visual function, investigate the effect of perinatal brain injury severity, gestational age at birth and sex upon visuocognitive development in infants at risk of neurodevelopmental impairment. Method: Recruitment was from UK neonatal units. Eligibility: ≤ 31 weeks, weight < 9th percentile; < 31 weeks with ≥ Grade II intraventricular haemorrhage (IVH) or preterm white matter injury (PWMI); 31-40 weeks with ≥ Grade II IVH or PWMI, ≥ Sarnat Grade II HIE or defined brain MRI abnormalities. Stratification was by sex, gestation and brain injury severity. Randomised infants received neurotrophic supplementation or placebo, for 2 years. Primary outcome was Bayley Scales of Infant Development III (BSID III) composite cognitive score (CCS) after 2 years. Secondary outcomes included BSID III composite language score (CLS) and BSID III composite motor score (CMS). Cortical visual measures were pattern reversal visual event related potential (PR-VERP) latency (transient and calculated), orientation reversal visual event related potentials (OR-VERP), and the Fixation Shift test (FS). Functional behavioural vision was assessed using the Atkinson Battery of Child Development for Examining Functional Vision (ABCDEFV). Local Ethics Committee approval was granted. Results: 62 neonates were recruited. After 2 years, mean CCS in the intervention group was 87.7 (SD 20.4) and 81.6 (SD 18.5) in the placebo group (mean difference = 2.28, p=0.13; -0.2, 18.2). Mean CLS in the intervention group was 91.5 (SD 20.1) and 83.2 (SD 19.6) in the placebo group (mean difference = 2.74, p=0.1; -2.4, 18.3). CMS was similar in both groups. In relation to trial visual outcome measures, more infants in the placebo group gave a statistically significant OR-VERP response than in the intervention group (p=0.03). There were no statistically significant differences between the placebo and intervention on any other trial visual outcome measure. Cohort analyses indicate that transient PR-VERP latency is prolonged in children at risk of neurodevelopmental disability compared to typically developing infants (mean difference = -23.3, p=0.015, 95% CI -42.10 - -4.54). Calculated PR-VERP latency is prolonged to an even greater extent in children at risk of neurodevelopmental disability compared to typically developing infants (mean difference -148.6, p=0.000, 95% CI -179.7- -117.43), and remains prolonged across the age range tested. Conclusions: 1) The difference in CCS and CLS between intervention and placebo groups represents a clinically significant effect size. Use of neurotrophic micronutrient supplementation in infants ARNI warrants exploration in a large multicentre RCT. 2) Calculated PR-VERP latency may be a more appropriate outcome measure of cortical visual function than transient PR-VERP latency in infants at risk of neurodevelopmental disability.

In a heterogeneous environment where temperature influences fitness, individuals must navigate to a thermal optimum to maximize reproductive output and minimize physiological stress. However, the optimal temperature varies among individuals due to genetic and environmental contributions. The neural and genetic basis to such natural variation in behavior has remained elusive in most cases, as the high-throughput genomic, neurodevelopmental, and behavioral techniques were not developed. Using the nematode / 10000-01-01

No description available.

The human brain, as much as it is “unaccountable” in the eyes of Virginia Woolf, is a marvel. It is the evolutionary increase in brain size, especially in the cerebral cortex, that both allowed Mrs Woolf to create and us to perceive the beautiful imagery that exists in her fictional world. The evolutionary increase in brain size in part reflects the increase in the number of neurons generated during neocortical development. This in turn reflects two principal features of cortical expansion, namely, an increase in the number of neural stem and progenitor cells (from here on referred to as progenitor cells) and their neurogenic potential. Strikingly, in order to cater for this increase in progenitor cells and neurogenic potential, there is a significant expansion and diversification of basal progenitors in the subventricular zone (SVZ). Basal progenitors can be divided into three types: basal intermediate progenitors (bIPs), basal radial glias (bRGs) and transit-amplifying progenitors (TAPs). bIPs are the most abundant progenitors in the mouse SVZ. These cells are non-polar and are Pax6 and Sox2 negative, but Tbr2 positive. They have limited proliferative capacity as they can divide only once to produce two neurons. bRGs and TAPs, on the other hand, are able to undergo multiple rounds of division and exist in higher abundance in gyrencephalic brains (for bRG, in humans up to 50% versus mouse 5% at mid-neurogenesis). The morphology of bRGs are reported to be dynamic (fluctuating between states of having process(es) to none), whereas TAPs are generally described to be non-polar during mitosis. bRGs are known to express Pax6 and Sox2 but not Tbr2 while TAPs are known to express both Pax6 and Tbr2. The increase in the proportion of these self-renewing basal progenitors (more specifically bRGs) might allow for cortical expansion. Hence, the main objective of this doctoral work was to generate more bRGs in the mouse dorsal telencephalon, the region that ultimately develops to become the cerebral cortex. To achieve this objective, two approaches were used– (i) a general approach by microinjecting a pool of ferret poly-A+ RNA and (ii) a candidate approach by conditionally expressing the transcription factor Pax6. In the general approach, the microinjection technique was first established and validated in an organotypic slice culture of the mouse dorsal telencephalon. A pool of ferret poly-A¬+ RNA extracted at P1, the developmental stage corresponding to the peak of bRG production, was then microinjected into the dorsal telencephalon. We hypothesized that at the peak of bRG production, the “instructive” messages on how to generate bRG would be at their peak. Hence, by introducing these “instructive” messages into a apical radial glia (aRG), these cells would thus “know” how to generate bRGs. At 24 h after microinjection, only aRGs, the predominant progenitor residing in the ventricular zone during mid-neurogenesis were recovered. At 48 h after microinjection, however, 75% of cells that translated the ferret poly-A¬+ RNA had a morphology reminiscent of bRG. These cells were located away from the ventricular surface and had a basal but not apical process. We conclude from these experiments that we did indeed generate bRG-like cells in the mouse dorsal telencephalon via microinjection of the ferret poly-A¬+ RNA. In the candidate approach, this work aimed to conditionally express Pax6, a transcription factor that has been linked to proliferation and neurogenesis in aRG. More specifically, as there is a significant increase in the number of Pax6 positive cells (bRGs) in the SVZ of gyrencephalic animals during mid-neurogenesis, we wanted to recapitulate this phenomenon in the mouse dorsal telencephalon, where Pax6 is normally downregulated. To achieve this, the Tis21–CreERT2 mouse was used. Tis21 is a pan-neurogenic marker that is switched on once aRG switches from a proliferative division (i.e. 1 aRG⇒2aRG) to a neurogenic division (i.e. 1aRG⇒1aRG+1bIP). Consequently, the neurogenic aRGs and its progeny, bIPs would thus be Tis21 positive. By conditionally expressing Pax6 in Tis21 positive aRGs, the ectopic expression of Pax6 was successfully induced in the SVZ of the mouse dorsal telencephalon. Interestingly, conditional expression of Pax6 increased the percentage of proliferating cells in the SVZ. However, instead of producing more bIPs as predicted by the neurogenic division of Tis21 positive aRGs, these cells had the cell morphology, transcription factor expression profile, and division-type of bRGs and/or TAPs. Thus, using the conditional expression of Pax6 we were able to generate more bRG-like progenitors in the mouse dorsal telencephalon. The fate of these conditionally expressing Pax6 progenitors at a later stage was then investigated. A phenotypic change in the behaviour of neurons generated was observed. Instead of migrating into the cortical plate, cells that were highly expressing Pax6 formed a heterotopia at the SVZ or intermediate zone, suggestive of Pax6 interfering with neuronal migration. Interestingly, of those lowly expressing Pax6 cells that successfully migrated to the CP, a disproportionate majority became upper layer neurons. As the fate of neurons are dependent on their date of birth (i.e early born neurons are normally found in the deep layer while late born neurons are normally found in the upper layer), the increase in the upper layer neurons is consistent with the fact that conditionally expressing Pax6 delayed the birth of these neurons by delaying neurogenesis in order to increase the number of proliferative divisions. Interestingly, this increase in upper layer neurons is consistent with the difference between small- and large-brained species. In conclusion, through this work more bRGs was successfully generated in the mouse dorsal telencephalon through two distinct but complementary approaches.