A quantitative trait variant in Gabra2 underlies increased methamphetamine stimulant sensitivity

Abstract

Psychostimulant (methamphetamine, cocaine) use disorders have a genetic component that remains mostly unknown. We conducted genome‐wide quantitative trait locus (QTL) analysis of methamphetamine stimulant sensitivity. To facilitate gene identification, we employed a Reduced Complexity Cross between closely related C57BL/6 mouse substrains and examined maximum speed and distance traveled over 30 min following methamphetamine (2 mg/kg, i.p.). For maximum methamphetamine‐induced speed following the second and third administration, we identified a single genome‐wide significant QTL on chromosome 11 that peaked near the Cyfip2 locus (LOD = 3.5, 4.2; peak = 21 cM [36 Mb]). For methamphetamine‐induced distance traveled following the first and second administration, we identified a genome‐wide significant QTL on chromosome 5 that peaked near a functional intronic indel in Gabra2 coding for the alpha‐2 subunit of the GABA‐A receptor (LOD = 3.6–5.2; peak = 34–35 cM [66–67 Mb]). Striatal cis‐expression QTL mapping corroborated Gabra2 as a functional candidate gene underlying methamphetamine‐induced distance traveled. CRISPR/Cas9‐mediated correction of the mutant intronic deletion on the C57BL/6J background to the wild‐type C57BL/6NJ allele was sufficient to reduce methamphetamine‐induced locomotor activity toward the wild‐type C57BL/6NJ‐like level, thus validating the quantitative trait variant (QTV). These studies show the power and efficiency of Reduced Complexity Crosses in identifying causal variants underlying complex traits. Functionally restoring Gabra2 expression decreased methamphetamine stimulant sensitivity and supports preclinical and human genetic studies implicating the GABA‐A receptor in psychostimulant addiction‐relevant traits. Importantly, our findings have major implications for studying psychostimulants in the C57BL/6J strain—the gold standard strain in biomedical research.

Genetic basis of thermal nociceptive sensitivity and brain weight in a BALB/c reduced complexity cross

“…34 This reduced genetic complexity can facilitate the identification of causal genes and variants influencing complex traits by reducing the density of genetic polymorphisms within a chromosomal interval and minimizing variant-variant interactions, given that a single major locus is typically identified for a given trait on an already nearly isogenic, segregating genetic background. [35][36][37] We have had repeated success in using Reduced Complexity Crosses (RCC) between C57BL/6 substrains to map the genetic basis of complex traits ranging from binge-like eating 38 to thermal nociception as measured via hot plate nociceptive sensitivity 39 to methamphetamine stimulant sensitivity, 40 to name a few. Combining behavioral QTL with tissuespecific gene expression QTL analysis can facilitate identification of plausible candidate genes (e.g., Ryr1 for hot plate; Bryant et al, 2019) and even sometimes directly validate causal variants via CRISPR/Cas9 (e.g., Gabra2 for methamphetamine stimulant sensitivity).…”

“…Combining behavioral QTL with tissuespecific gene expression QTL analysis can facilitate identification of plausible candidate genes (e.g., Ryr1 for hot plate; Bryant et al, 2019) and even sometimes directly validate causal variants via CRISPR/Cas9 (e.g., Gabra2 for methamphetamine stimulant sensitivity). 40 As another example of substrains that can be used in a RCC, 36 the BALB/cJ (J) and BALB/cByJ (By) substrains of mice were separated in 1935 after the completion of backcrossing at generation F37 and have been maintained since then as separate inbred substrains. Over time, the fixation of spontaneous mutations and residually heterozygous loci, yielded approximately 8500 SNPs, insertions, and deletions that distinguish the substrains, comprising approximately a 500-fold reduction in genetic complexity compared to C57BL/6J versus most classical inbred strains.…”

Cocaine-Induced Locomotor Activation Differs Across Inbred Mouse Substrains

“…Additionally, a recently developed genotyping array captures polymorphisms between inbred mouse substrains, facilitating rapid and reliable genotyping of RCCs (33). RCCs have been used successfully to identify genetic polymorphisms that impact psychostimulant response, binge eating, binge alcohol consumption, thermal nociception and brain weight (34)(35)(36)(37)(38).…”

Zhx2 is a candidate gene underlying oxymorphone metabolite brain concentration associated with state-dependent oxycodone reward

“…Large phenotypic variance combined with a drastic reduction in the density of genetic polymorphisms facilitates the identification of causal genes/variants that can readily be validated via gene editing (Mulligan et al, 2019). Reduced complexity crosses have been used between multiple mouse substrains (e.g., C57BL/6, DBA/2, BALB/c) to map the genetic basis of complex phenotypes such as psychostimulant sensitivity, binge-like eating, and thermal nociception (Beierle et al, 2022;Goldberg et al, 2021;Harkness et al, 2015;Kirkpatrick et al, 2017;Kumar et al, 2013;Miner et al, 2017;Reed et al, 2018;Shi et al, 2016).…”