Micrornas 9a, 9b, 9c And 315 Regulate Expression Of A Reporter For The Neuronal Microtubule-Associated Protein Futsch/map1b

Fragile X syndrome (FXS) is the most common form of inherited mental retardation in humans. FXS is caused by loss of the Fragile X Mental Retardation Protein (FMRP), an important regulator of neuronal mRNA translation. Patients with FXS display cognitive deficits including memory problems. Protein synthesis-dependent long-term changes in synaptic plasticity are involved in the establishment and maintenance of long-term memory. One prevalent theory of FXS pathology predicts that FMRP is required to negatively regulate the translation of important mRNAs at the synapse. We are investigating microRNAs (miRNAs) as a potential regulator of synaptic FMRP-regulated mRNAs that have previously been described as being crucial to the process of synaptic plasticity. The general hypothesis underlying this thesis is that FMRP may negatively regulate the expression of futsch (the Drosophila homologue of the microtubule-associated protein gene MAP1B) via the miRNA pathway. The first step we took in testing this hypothesis was to confirm that futsch is subject to miRNA-mediated translational control. Using in silico target analysis, we predicted that several neuronally expressed miRNAs target the futsch mRNA 3'UTR and repress expression of Futsch protein. Then, using an in vitro luciferase reporter system, we showed that miR-315 and members of the miR-9 family selectively down-regulated futsch reporter translation. We have confirmed by site- directed mutagenesis that the miRNA interaction with the futsch 3'UTR is specific to the miRNA seed region binding site. Interestingly, reduction of FMRP levels by RNAi had no effect on futsch 3'UTR reporter expression. Together, these data suggest regulation of futsch expression by the miRNA pathway might be independent of FMRP activity. However, additional experiments need to be completed to confirm these preliminary results.

Re-Examining the Stop-Signal Task to Test Competing Theories of AD/HD

The current study tested two competing models of Attention-Deficit/Hyperactivity Disorder (AD/HD), the inhibition and state regulation theories, by conducting fine-grained analyses of the Stop-Signal Task and another putative measure of behavioral inhibition, the Gordon Continuous Performance Test (G-CPT), in a large sample of children and adolescents. The inhibition theory posits that performance on these tasks reflects increased difficulties for AD/HD participants to inhibit prepotent responses. The model predicts that putative stop-signal reaction time (SSRT) group differences on the Stop-Signal Task will be primarily related to AD/HD participants requiring more warning than control participants to inhibit to the stop-signal and emphasizes the relative importance of commission errors, particularly "impulsive" type commissions, over other error types on the G-CPT. The state regulation theory, on the other hand, proposes response variability due to difficulties maintaining an optimal state of arousal as the primary deficit in AD/HD. This model predicts that SSRT differences will be more attributable to slower and/or more variable reaction time (RT) in the AD/HD group, as opposed to reflecting inhibitory deficits. State regulation assumptions also emphasize the relative importance of omission errors and "slow processing" type commissions over other error types on the G-CPT. Overall, results of Stop-Signal Task analyses were more supportive of state regulation predictions and showed that greater response variability (i.e., SDRT) in the AD/HD group was not reducible to slow mean reaction time (MRT) and that response variability made a larger contribution to increased SSRT in the AD/HD group than inhibitory processes. Examined further, ex-Gaussian analyses of Stop-Signal Task go-trial RT distributions revealed that increased variability in the AD/HD group was not due solely to a few excessively long RTs in the tail of the AD/HD distribution (i.e., tau), but rather indicated the importance of response variability throughout AD/HD group performance on the Stop-Signal Task, as well as the notable sensitivity of ex-Gaussian analyses to variability in data screening procedures. Results of G-CPT analyses indicated some support for the inhibition model, although error type analyses failed to further differentiate the theories. Finally, inclusion of primary variables of interest in exploratory factor analysis with other neurocognitive predictors of AD/HD indicated response variability as a separable construct and further supported its role in Stop-Signal Task performance. Response variability did not, however, make a unique contribution to the prediction of AD/HD symptoms beyond measures of motor processing speed in multiple deficit regression analyses. Results have implications for the interpretation of the processes reflected in widely-used variables in the AD/HD literature, as well as for the theoretical understanding of AD/HD.