Biomedical Sciences ETDs

Michael Baca

Publication Date

12-1-2013

Abstract

Genes and environmental conditions interact in the development of cognitive capacities and each plays an important role in neuropsychiatric disorders such as attention deficit/hyperactivity disorder (ADHD) and schizophrenia. Multiple studies have indicated that the gene for the SNARE protein, SNAP-25 is a candidate susceptibility gene for ADHD, as well as schizophrenia, while maternal smoking is a candidate environmental risk factor for ADHD. In this study, mice heterozygous for a Snap25 allele and deficient in SNAP-25 expression were utilized to model genetic effects in combination with prenatal exposure to nicotine to explore genetic and environmental (G x E) factors and interactions. Striatal long-term depression (LTD) is a form of synaptic plasticity in which there is a reduction in the glutamate released by cortical afferents onto striatal medium spiny neurons (MSNs). The glutamatergic inputs activate ionotropic and group I metabotropic glutamate receptors while dopaminergic inputs from the substantia nigra pars compacta activate type-2 dopamine receptors (D2Rs), which collectively lead to post-synaptic production of endocannabinoids that diffuse in a retrograde manner to activate pre-synaptic type-1 cannabinoid (CB1) receptors on to pre-synaptic terminals. The CB1 receptors initiate signaling cascades that lead to the long-term decrease in glutamate release in pre-synaptic glutamate terminals. Using a high frequency stimulus (HFS) electrophysiological paradigm for LTD induction in striatal MSNs, I first characterized synaptic depression in four G x E groups representing mice prenatal nicotine exposed or not or having Snap25 deficiency or not, that showed responses which could be divided by cluster analysis into populations expressing LTD and short-term depression (STD). STD is characterized by an initial decrease in amplitude in the response to the HFS followed by near full recovery of the response within 30 minutes, while LTD occurs when the initial decrease remains attenuated for at least 30 minutes. I found that prenatal exposure to nicotine in Snap25 heterozygote mice produced a less robust LTD population and less return to baseline in the STD population. Using receptor antagonists in the same HFS electrophysiological paradigm I next examined the roles of dopaminergic D2Rs and cannabinoid CB1Rs, both critical for LTD induction in the striatum. I found that prenatal exposure to nicotine in Snap25 heterozygote mice produced a deficit in the D2R-dependent induction of LTD, although the CB1R involvement in plasticity was not impaired. From these results I developed the hypothesis that the impaired induction of LTD due to prenatal exposure to nicotine in Snap25 heterozygote mice, could be related to changes in D2R affinity and/or changes in the number of D2R receptors. This was initially tested using a [35S]-GTPγS binding assay to measure the agonist-stimulated response of G-protein-coupled D2R receptors. Indeed, the agonist-stimulated response was found to be reduced in Snap25 heterozygote mice prenatally exposed to nicotine, which was consistent with a reduction in affinity of the agonist for the D2R receptor, and/or reduced GPCR coupling to downstream signaling, but not with changes in the number of receptors. Next, a D2R agonist saturation binding assay, using [3H]-quinpirole (a D2R selective agonist) followed by Scatchard analysis, showed that Snap25 heterozygote mice prenatally exposed to nicotine exhibited significantly lower affinity (higher Kd) for D2R binding without a significant change in Bmax, which would reflect a change in the number of receptors. Collectively, these results support individually (G and E factors) and in combination (G x E interactions), the HET genotype and prenatal nicotine exposure lead to an impaired D2R GPCR signaling resulting from decreased agonist affinity and possibly receptor-effector coupling of the D2R receptors These receptor binding observations are consistent with the electrophysiological results showing that, in the presence of a D2R antagonist, cortico-striatal circuits in Snap25 heterozygote mice prenatally exposed to nicotine exhibit a significant deficit in LTD induction. This study demonstrates that genetic conditions present in a Snap25 haplodeficient mouse model together with prenatal nicotine exposure can together alter specific receptor affinity and signaling in dopaminergic D2 receptors (D2Rs) with functional consequences in synaptic long-term depression (LTD). Since SNAP25 is a candidate susceptibility gene for cognitive disorders such as ADHD and schizophrenia and additionally prenatal exposure to nicotine is a candidate environmental risk factor for ADHD, these studies have important translational relevance. The study also presents a general set of experimental procedures by which potential G x E interactions can be explored to determine if they alter receptor function and/or synaptic plasticity in specific brain regions.

Keywords

Neuroscience, Gene-environment interactions, Electrophysiology

Sponsors

National Institutes of Health; Sandia National Laboratories

Document Type

Dissertation

Language

English

Degree Name

Biomedical Sciences

Level of Degree

Doctoral

Department Name

Biomedical Sciences Graduate Program

First Committee Member (Chair)

Wilson, Michael

Second Committee Member

Allan, Andrea

Third Committee Member

James, Conrad

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