Genetic association between dopamine signaling molecules and striatal presynaptic dopamine

Abstract

Background: Dopamine- and cAMP-regulated phosphoprotein of 32 KDa (DARPP-32) and the D2receptor are central, closely-linked mediators of dopamine signaling, which have been implicated in the pathophyiology of several heritable neuro-psychiatric illnesses, including schizophrenia. In medium spiny neurons of the striatum, D2 receptor activation obstructs phosphorylation of DARPP-32, curtailing its ability to inhibit protein phosphatase 1, thereby affecting the phosphorylation status of a multitude of downstream targets. The genes coding for DARPP-32 and D2 receptor contain frequent, functional variations in humans, which have demonstrated associations with dopamine-dependent cognitive measurements, striatal activity and connectivity by functional magnetic resonance imaging (MRI), and, in select experiments, modest association with schizophrenia. These variants are a 7- marker haplotype in the gene coding for DARPP-32 which predicts altered mRNA expression in postmortem prefrontal cortical samples, and a single nucleotide polymorphism (SNP) in the D2 receptor gene (rs6277), which affects protein folding and stability. Though the growing literature of association studies examining dopamine-related neurocognitive phenotypes forward interpretation that these DARPP-32 and D2 receptor gene variants alter dopamine system physiology, there is little direct, consistent evidence for this from in vivo human studies. Methods: As a well-validated method of assaying striatal presynaptic dopamine synthesis, 18F-FDOPA PET was employed in 82 healthy Caucasian adults under 50 years of age who also underwent genotyping for DARPP-32 and D2 receptor variants with Taq-Man 50-exonuclease assay. After carbidopa pretreatment to prevent peripheral tracer degradation and at least 6 hours of fasting to limit tracer competition for central nervous system access, 8-16 mCi of 18F-DOPA were injected intravenously and 90 minutes of dynamically binned images were acquired. These images were attenuation-corrected, realigned, co-registered to a structural MRI obtained in a separate session, warped to standard space using ANTS software and smoothed. The Patlak method was adopted to calculate the specific uptake constant Ki using an occipital reference region. Striatal region of interest and voxel-wise regression analyses queried Ki maps for genotype and gene-gene interaction effects, controlling for sex. Results: Genotype groups were well-matched for age. The non-risk genotypes for DARPP-32 (haplotype non-homozygotes) and D2 receptor (T homozygotes) were associated with greater Ki in the caudate (p=3.7x10-3 uncorrected, and p=1.5x10-2 FWE-corrected, respectively). When examined together, there existed a gene-by-gene interaction in the caudate head, where subjects who were both DARPP-32 non-homozygotes and D2 receptor T homozygotes had far greater Ki than other groups (p=1.0x10-2, FWE-corrected). Conclusions: In line with previous data from animal studies suggesting an influence of DARPP-32 and D2 receptor genes on presynaptic dopaminergic characteristics, our results indicate that the studied variants - or those in linkage disequilibrium with them - may bias striatal dopamine synthetic function in the living human brain. Though their direction does not support a straightforward presynaptic mechanism of schizophrenia risk, these findings provide preliminary evidence in favor of a dopaminergic explanation to previous neurocognitive and neurophysiological associations with these genetic polymorphisms. The described epistasis suggests that in the cognitive caudate, presynaptic dopaminergic tone may be regulated by a network of proteins extending at least to post-synaptic terminals, and future work is needed to comb this understudied pathway for novel targets in the treatment of dopamine-related neuropsychiatric disease.